USF Libraries

Evaluation of microbial removal at a water reclamation facility

MISSING IMAGE

Material Information

Title:
Evaluation of microbial removal at a water reclamation facility
Physical Description:
viii, 107 leaves : ill. ; 29 cm.
Language:
English
Creator:
Riley, Kelley R.
Publisher:
University of South Florida
Place of Publication:
Tampa, Florida
Publication Date:

Subjects

Subjects / Keywords:
Water reuse -- Virginia   ( lcsh )
Water -- Microbiology   ( lcsh )
Dissertations, Academic -- Marine Science -- Masters -- USF   ( fts )

Notes

General Note:
Thesis (M.S.)--University of South Florida, 1998. Includes bibliographical references (leaves 78-84).

Record Information

Source Institution:
University of South Florida
Holding Location:
Universtity of South Florida
Rights Management:
All applicable rights reserved by the source institution and holding location.
Resource Identifier:
aleph - 025977416
oclc - 41930692
usfldc doi - F51-00140
usfldc handle - f51.140
System ID:
SFS0044191:00001


This item is only available as the following downloads:


Full Text

PAGE 1

EVALUATION OF MICROBIAL REMOVAL AT A WATER RECLAMATION FACILITY by KELLEY R. RILEY A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science Department of Marine Science University of South Florida December 1998 Major Professor: Joan B Rose, Ph. D

PAGE 2

Graduate School University of South Florida Tampa, Florida CERTIFICATE OF APPROVAL Master s Thesis This is to certify that the Master's Thesis of KELLEY R. RILEY with a major in Marine Science has been approved by the Examining Committee on April 23 1998 as satisfactory for the thesis requirement for the Master of Science degree Examining Committee : Professor: Rose Ph. D Member : JbhD H Ph D Member : Richard 0.' Mines, Jr. P.E.

PAGE 3

DEDICATION To my husband Darryl and my children Kristopher and Morgen for their support and the sacrifices they made so that I could obtain this degree I will forever be grateful. To my mom and sister for their constant support and encouragement and finally to my dad for instilling in me the value of education

PAGE 4

ACKNOWLEDGMENTS I would like to acknowledge the staff at UOSA for their assistance in sample collection and analyses I would also like to acknowledge my advisor and friend Joan Rose for her assistance support and the opportunity she gave me. I thank the people in the lab that I had the pleasure of working with and especially to John Lisle, for his guidance and wisdom

PAGE 5

TABLE OF CONTENTS LIST OF TABLES iii LIST OF FIGURES v ABSTRACT vii CHAPTER 1 : 1 Water Reclamation and Reuse 1 History of Water Reclamation and -Reuse 2 Current Indirect Potable Reuse Projects in the United States 5 Pilotand Demonstration-Scale Projects 7 Summary 11 Microorganisms in Wastewater 13 Current Indicator Microorganisms for Evaluating Water Quality and Treatment 15 Alternative Indicator Microorganisms 16 Development of the Upper Occoquan Sewage Authority 18 CHAPTER 2. RESEARCH OBJECTIVES 20 CHAPTER 3 MATERIALS AND METHODS 22 Sampling Sites 22 Microbiological Sampling 25 Bacteria 25 Protozoa 26 Human Viruses 27 Coliphage 28 Pilot Studies 29 CHAPTER 4 RESULTS 34 UOSA Treatment Processes 34 Bacteria 36 E Human Viruses 47 Coliphage 47 Pilot Studies 55

PAGE 6

CHAPTER 5 DISCUSSION CHAPTER 6 CONCLUSION LIST OF REFERENCES APPENDICES APPENDIX A APPENDIX B APPENDIXC 63 76 78 85 86 99 104 i i

PAGE 7

LIST OF TABLES Table 1 Description of Sampling Sites within UOSA Reclamat i on Facil i ty 24 Table 2 Summary of Microorgan i sm and Method 24 Table 3 Clostrid i um perfringens 39 Table 4 Enterococci 40 Table 5 Total coliform 41 Table 6 Fecal coliform 42 Table 7 Cryptosporidium 48 Table 8 Giard i a 49 Table 9 Enterov i ruses 50 Table 1 0 Col i phage 51 Table 11 Pilot Study on the Removal of Beads and Phage by Chemical Lime Treatment at a F l ow Rate of 7 gpm 57 Table 12 Pilot Study on the Removal of Beads and Phage by Chem i cal Lime Treatment at a Flow Rate of 3 gpm 57 Table 1 3 Pilot Study on the Removal of Cryptosporidium and Phage by Chemical Lime Treatment at a Flow Rate of 7 gpm 58 Table 14 Pilot Study on the Removal of Cryptosporidium and Phage by Chem i cal Lime Trea t ment at a Flow Rate of 3 gpm 58 Table 15 Co rr elation Matrix of Ind i cator and Pathogenic Mi croorgan i sms for Ent i re Treatment Process 70 iii

PAGE 8

Table 16 Table 17 Table 18 Correlation Matrix of Indicator and Pathogenic Microorganisms after Lime Treatment Removal and Inact i vation of Cryptosporid ium, Beads and Phage by Chemical Lime Treatment by Pilot Studies Compared to Monitoring Data Comparison of the UOSA Final Effluent to the Final Effluent Reservoir Water Quality 7 0 71 75 iv

PAGE 9

LIST OF FIGURES Figure 1 Schematic of UOSA Process Description 23 Figure 2 Schematic of UOSA Pilot Plant 30 Figure 3 Average Levels for Positive Samples through the Treatment Train for Bacteria 43 Figure 4 Percentage of Samples Positive through the Treatment Train for Bacteria 44 Figure 5 Log1o Removal of Sites Compared to Site 001 45 Figure 6 Average Levels for Positive Samples for Enteroviruses Protozoa and Coliphage 52 Figure 7 Percentage of Samples Positive for Enteroviruses Protozoa and Coliphage 53 Figure 8 Log1o Removal of Sites Compared to Site 001 54 Figure 9 Fluorescent Bead Removal by Chemical Lime Treatment 59 Figure 10. Cryptosporidium Removal by Chemical Lime Treatment 60 Figure 11. MS2 Removal by Chemical Lime Treatment 61 Figure 12 PRD1 Bacteriophage Removal by Chemical Lime Treatment 62 Figure 13 Comparison of Fecal Coliform Removal to Pathogen Removal throughout the Treatment Plant 64 Figure 14 Comparison of Clostridium Removal to Pathogen Removal throughout the Treatment Plant 65 v

PAGE 10

Figure 15 Comparison of Enterococci Removal to Pathogen Remova l throughout the Treatment Plant 66 Figure 16. Comparison of Enteroviruses and Coliphage in Posit i ve Samples 68 Figure 17 Comparison of Enteroviruses and Clostridium in Postive Samples 69 vi

PAGE 11

EVALUATION OF MICROBIAL REMOVAL AT A WATER RECLAMATION FACILITY by KELLEY R. RILEY An Abstract Of a thes i s subm i tted in partial fulfillment of the requirements for the pegree of Master of Science Department of Marine Science University of South Florida December 1998 Major Professor : Joan B. Rose, Ph D vii

PAGE 12

Water reclamation and reuse have become an important consideration for many communities experiencing increased growth and demand on water resources. Research has been focused on using this reclaimed water for indirect potable reuse, in other words, supplementation of surface waters or groundwaters currently used for drinking water supplies. The Upper Occoquan Sewage Authority (UOSA) Water Reclamation Plant has been reclaiming wastewater and discharging into the Occoquan Reservoir since 1978 This reservo i r serves as a drinking water supply for approximately one million people in Northern Virginia A study was initiated to monitor the bacteria protozoa and viruses entering the water reclamation plant and to evaluate the unit processes for the removal of these microorganisms Eight sites within the plant were monitored monthly for a year for Enterococci, Clostridium, total and fecal coliforms, coliphage, Giardia, Cryptosporidium and enteroviruses Chemical lime treatment with second stage recarbonation and disinfection were the most efficient barriers to the passage of microorganisms Of all the indicators Clostridium and coliphage best reflected the removal of enteroviruses for the chemical treatment system and the disinfection process Abstract Professor : Joan B Rose, Ph.D 6fessor Marine Science Date Approved : Q3) JC19'(: viii

PAGE 13

CHAPTER 1 INTRODUCTION Water Reclamation and Reuse Water reclamation and reuse have become an important consideration and reality for conserving existing potable water supplies due to increased demands on water resources for domestic, commercial, industrial and agricultural uses Water reclamation involves treating wastewater with advanced treatment processes to a high quality so that it can be reused again A wide variety of reclaimed water usages include : landscape and agricultural irrigation ; i ndustrial process water, power plant cooling water ; toilet flushing car washing and augmentation of recreational water bodies (D Angelo 1996) Recla i med water (properly treated wastewater effluent) has been successfully used in the United States for decades to meet nonpotable water needs. Recent research has focused on the feasibility of using reclaimed water for supplementing surface water or groundwater drinking water suppl i es There are two types of potable reuse: indirect and direct. Indirect potable reuse involves treating wastewater to a quality equal to or better than the current water supply and then purposely reintroducing it into a surface water or groundwater that will ultimately be used as a potable water supply for a population Direct 1

PAGE 14

potable re u se involves treating wastewater to a dri n k ing water quality and then purposely introducing it directly into a water treatment plant or potable water distribution system (McEwen and Richardson 1996) The cont i nued depletion of potable supplies has increased the interest in using highly treated reclaimed water to augment potable water resources Currently four planned indirect potable reuse full-scale projects in the United States are found in Virginia Texas Georgia and California (Asano 1995 Asano and Levine 1996 Pia et al1996, McEwen and R i chardson 1996) Tampa Bay, Florida i s currently considering the use of reclaimed water to supplement the existing surface and groundwater supplies A planned direct potable reuse full sca l e faci l i ty is located in South Africa (van Leeuwen 1996, Hattingh and Bourne 1988 Grabow and Isaacson 1978 Hrudey et al 1-991, Asano 1995 Asano and Levine 1995). History of Water Reclamation and Reuse Wastewater reclamat i on and reuse have their roots in the early water and wastewater systems of the Minoan civilization in ancient Greece The use of wastewater for agricultural irrigation dates back 5 000 years (Angelakis and Spyridakis 1996, Asano and Levine 1996) During the nineteenth century large scale wastewater carriage systems were used for discharge i nto surface waters which resulted in the unplanned indirect use of sewage and other effluents for potable water supplies (Asano and Levine 1996) This indirect reuse caused 2

PAGE 15

epidemics of Asiatic cholera and typhoid during 1840-1850 and resulted i n the discovery that the water supply was causing the epidem i cs Improved water treatment techniques, inc l uding the introduction of water filtration the development of alternative water sources using reservoirs and aqueducts and the relocation of water intakes upstream from wastewater discharges allowed some protection of potable water systems During the 20th century programs were developed in the United States for the planned reuse of wastewater The State of California was the first to promote water reclamation and reuse and promulgated the first regulations for reuse in 1918 (Asano and Levine 1996 Crook and Surampalli 1996). Some of the in i t i al reuse programs were developed to provide water for irrigation in Arizona and California in the late 1920s In 1940 chlorinated wastewater was used for steel processing In 1960 urban water reuse systems were developed for Colorado and Florida (Asano and Levine 1996 ) Currently in the 1990s there is increased pressure to develop new sources of water especially i n water poor areas such as the West and the Southwest. Wastewater treatment and pur i fication processes are currently available that can produce water of any quality ; therefore water reuse has become a factor in the planning and efficient use of water resources (Asano and Levine 1996 Crook and Surampall i 1996) The longest history of potable reuse is i n Namibia where potable reuse has provided 10-20% of Windhoek's water supply since 1969 (Asano and Levine 1995 Grabow and Isaacson 1978 Hattingh and Bourne 1989 Asano 1995 van Leeuwen 1996) The South African Council for Scientific and Industrial 3

PAGE 16

Research (CSIR) conducted resea r ch on the technology of water reclamation from secondary effluent since the early 1960s and the Stander Water Reclamat i on Plant was built in 1970 (van Leeuwen 1996) By the end of the 1970s the processes used at the Stander Water Reclamation Plant included coagulation and flocculation, settling ozonation sand filtration, biological granular activated carbon chlorinat i on and stabilization Lack of further funding and difficulties supplying the water from the plant to the consumers caused the project to end but the plant continued to operate exclusively for research purposes The Windhoek Water Reclamation Plant was constructed in 1968 during a devastating drought and followed the design of the Stander Water Reclamation processes This plant was built to supplement the drinking water supply to Windhoek South West Africa and was the first plant in the world to initially reclaim wastewater for the direct supplementation of the city s drinking water supply (van Leeuwen 1996 Grabow 1991 Grabow and Isaacson 1978 Stander and Clayton 1977). Extensive microbiological analyses including enteric viruses parasite ova total and feca l coliforms fecal streptococci Pseudomonas aeruginosa Staphylococcus aureus, and Clostridium pettringens were performed on all water sources and treated supplies when the reclaimed water was first introduced into the system (Grabow and Isaacson 1978 Nupen 1970) and these evaluations were continually expanded Since 1973 epidemiological studies have been coord i nated by the South African Institute for Medical Research to evaluate the health aspects of the reclaimed water. Continuous health monitoring bioassays and epidemiological studies have 4

PAGE 17

proven that the reclaimed water is as safe as the other conventional water supplies (Grabow and Isaacson 1978 van Leeuwen 1996) The first documented case of indirect potable reuse of treated wastewater in the United States was short term and occurred during a severe drought from 1952-1957 at Chanute Kansas when treated wastewater was mixed with water stored in the river channel behind the water treatment dam Chlorinated secondary effluent was collected behind the dam on the river and used as intake water (Asano 1995) The treated water met bacteriological standards for drinking water but was pale yellow and had an unpleasant taste and odor, foamed when agitated and contained a h i gh level of dissolved minerals and organic chemicals Many technological advancements have occurred in wastewater treatment since that time which can produce a high quality water Current Indirect Potable Reuse Projects in the United States The current full-scale operating planned indirect potable reuse projects in the United States are in Texas Virginia Georgia and California These projects demonstrate how indirect potable reuse can be used to augment water resources by incorporating the multiple barrier approach to treatment (Asano and Levine 1995 Asano and Levine 1996 Asano 1995, McEwen and Richardson 1996 Pia et al 1996) Two to five advanced unit processes are specifically used to remove pathogenic microorganisms and trace organics. 5

PAGE 18

The Whittier Narrows Recharge Project (County Sanitation Districts of Los Angeles County) has been surface spreading secondary effluent for infiltration to an underground potable water supply since 1962 The amount of reclaimed water recharged annually averages 16% of the total inflow into the basin The population is estimated to be exposed to 0 to 23 percent of reclaimed water An independent scientific advisory panel to the State of California concluded that the groundwater replenishment project was as safe as the surface water supplies (Asano and Levine 1995, Asano and Levine 1996 Asano 1995 McEwen and Richardson 1996 Pia et al 1996) Water Factory 21 (Orange County Water District) has been blending reclaimed water with deep-well groundwater and using this mixture for deep injection into a heavily used aquifer to prevent salt water intrusion since 1976 (Asano 1995 McEwen and Richardson 1996 Pia et al 1996) The treatment includes lime clarification recarbonation filtration and then half of the flow receives carbon adsorption, the other half of the flow receives reverse osmosis and the entire flow is disinfected Viruses were monitored from 1975 to 1982 in the final effluent and it was demonstrated that the final effluent was essentially free of measurable levels of viruses" and no total coliforms were detected in 179 samples of the effluent tested in 1988 (Crook et al 1989) A planned surface water augmentation project in Georgia is the Clayton County project which utilizes conventional secondary treatment followed by land treatment involving overland flow The treated effluent becomes part of the inflow to a stream that serves as a drinking water source 6

PAGE 19

The Fred Harvey Water Reclamation Plant (EI Paso Public Service Board) recharged reclaimed water to the Hueco Bolson drinking water aquifer since 1985 (Asano 1995 McEwen and Richardson 1996 Pia et al 1996) The water travels to the potable well fields to become part of the potable water supply Treatment of the raw wastewater involves primary treatment act i vated sludge/powdered activated carbon treatment for organic removal nitrification and denitrification lime treatment, recarbonat i on filtration ozonation and granu lar activated carbon adsorption An increase in total dissolved solids content in the aquifer has been observed (McEwen and Richardson 1996). Pilotand Demonstration-Scale Projects Numerous studies have been conducted in an attempt to evaluate the health effects of using reclaimed water in order to supplement existing water supplies or replace the potable supply completely (Crook et al 1989 Asano 1995 McEwen and Richardson 1996 Pia et al 1996 Asano and Levine 1995 Asano and Levine 1996) Health effects refer to the large number of possible effects that can occur after consuming drinking water that is not treated properly Microbiological contaminants such as bacteria, viruses and protozoa can cause acute and chronic health effects (McEwen and Richardson 1996) Currently, toxicological testing is most commonly used to evaluate the effect of complex organic chemical mixtures that may be present in the water 7

PAGE 20

The Pomona Virus Study and the Monterey Wastewater Reclamat i on Study for Agriculture (MWRSA) provided evidence that alternative tertiary treatment systems could effectively remove viruses and potentially pathogen-free effluent could be produced using tertiary treatment and extended disinfection with chlorine (Asano and Levine 1996 Asano and Levine 1995 Asano et al 1992, Yanko 1993 Asano and Mujeriego 1988) The Whittier Narrows Groundwate r Replen i shment Project has been us i ng a m i xture of recla i med water stormwater and surface water to recharge the drinking water aquifer in Los Angeles County since 1962. The amount of recla im ed water in the extracted potable water supply is 0 to 11 percent. A Health Effects Study' begun in 1978 and the major findings published in 1984 stated that both the groundwater and the reclaimed water met Federal Drinking Water Standards ; no viruses were detected in either types of water ; and trace organ i c chem i cals did not exceed the theoretical lifetime risk value (McEwen and Richardson 1996, Asano and Lev i ne 1 996) This landmark study to evaluate the health effects associated with groundwater recharged with recla i med water provided an opportunity to identify the impacts of water reuse on water quality and human health Another objective of the study was to use the data to develop statewide wastewater reclamation criteria for groundwater recharge The study conducted toxicological and chemical stud i es percolat i on studies hydrogeologic studies and epidemiological studies. No measurab l e adverse i mpacts on the groundwater or the health of the populat ion drink i ng the water were found (Asano 8

PAGE 21

and Levine 1995 Asano and Levine 1996 Pia et al 1996 McEwen and Richardson 1996, Nellor 1985) The Potomac Estuary Experimental Water Treatment Plant evaluated the use of the Potomac Estuary as a source of drinking water Approximately 50% of the estuary may be comprised of treated wastewater during drought conditions The treatment plant was operated between 1981 and 1983 Finished water quality was compared to product water from three other treatment plants Microbiological parameters, metals and organics were monitored and two in-vitro toxicological tests were performed. The Ames Salmonella microsome test was used to assess the chemical mutagenesis and mammalian cell transformation assay was used to assess the potential mutagenesis of the reclaimed wastewater. It was determined that the reclaimed water compared favorably with the other product water in terms of the toxicological tests performed (McEwen and Richardson 1996) The City of Denver's Direct Potable Water Reuse Demonstration Project a 5 year project was started in 1985 to determine the feasibility of converting secondary wastewater to drinking water quality and comparing it to the current drinking water supply. Initial research evaluated many treatment processes to determine the optimum treatment sequence This sequence was utilized over a 2 year period to determine if the reclaimed water could meet Federal Drinking Water Standards The reclaimed water and the current drinking water were compared for chron i c toxicity and carcinogenicity in rats and mice A two generation reproductive toxicity study was also conducted No adverse health 9

PAGE 22

effects were reported (Asano 1995 Asano and Levine 1995 McEwen and Richardson 1996 Pia et al 1996 Lauer 1991, Lauer and Rogers 1996 Cond i e et al1994). The San Diego Total Resource Recovery Project began in 1988 to determine if raw sewage could be treated to a quality comparable to the existing Miramar reservoir raw water supply The findings indicated there was no difference in viral concentration in the two waters : the microbiological quality of the reclaimed water was better than the Miramar surface water with lower total coliform counts ; Giardia cysts were not detected in the reclaimed water or the Miramar reservoir ; and the reclaimed water met the microb i ological criteria for recreational waters without disinfection The study concluded that the health risk associated with the use of the reclaimed wa ter was less than or equal to the existing raw water supply (Asano 1995 Asano and Levine 1995 McEwen and Richardson 1996 Gagliardo et al 1996 Danielson et al 1996 de Peyster et al 1993) Tamp a Water Resource Recovery Project is a pilot project that began in 1986 to determine if a reclaimed secondary effluent could be produced to a quality that could be blended with existing surface and groundwaters that are currently used for potable supplies The reclaimed water met all primary and secondary drinking water standards, no organic chemicals were present and it was determined that the reclaimed water was an acceptable raw water supply (McEwen and Richardson 1996 Asano 1995 Final Report CH2M Hill 1993 Hemmer 1994) 10

PAGE 23

Summary The potential for indirect potable reuse is being seriously considered by many communities in the United States and other countries due to the diminishing supply of potable water resources An American Water Works Association (AVWVA) policy on reuse was described by Bergman who notes there are still many obstacles to be overcome, including acceptance of non potable reuse for edible crops and for use inside buildings" (Bergman 1994 ) A review article by Smith (1995) however notes that reuse programs are regulated on a state-by-state basis and more attention is being focused on reclaiming wastewater for potable uses In summarizing various demonstration and pilot projects conducted, six research projects were identified as the most useful for advancing the knowledge of water reclamation and reuse of wastewater. These projects included microbial risk assessment monitoring identification of new indicators of pathogenic microorganisms evaluation of effects of process selection on particle size distribution seasonal storage for reclaimed water nonpotable water management, and evaluation of metals and synthetic organic chemicals in irrigation water (Crook 1994). Van Riper and Geselbracht (1996) stated that the initial success of reclaimed water projects will be sustained if the public perceives the reuse of wastewater is healthy and desirable Federal regulations for water reuse do not exist in the United States However the United States Environmental Protection Agency (EPA) published 11

PAGE 24

Guidel i nes for Water Reuse in 1992 including a rev iew of sta t e standards ( C r oo k and Surampalli 1996 US EPA 1992) Th i s has led to the deve l opment of c riteri a by individual states which has resulted i n a variation of regulations among the d i fferent states The most common parameters monitored i nc l ude b i ochem i ca l oxygen demand (BOD) turbidity or total suspended solids (TSS), total or fecal coliform bacteria nitrogen and chlorine contact time and res i dual. If there is likely to be public contact with the reclaimed water tert i ary treatment i s required to produce a finished water that is virtually pathogen-free ( State of California 1978 Florida Department of Env i ronmental Regulat i on 1990 State of Ari zona 1991 ) Variations among states include the use of the tota l co li form as the indicator organism in California while the fecal coliform is used as the ind i cator organism in Florida Texas and Arizona Florida is the only state that requires mon i toring for TSS ; the other 3 states monitor for turb i dity California and Florida specify treatment processes wh i le Texas and Ar i zona do not. Californ i a and Florida have developed the most comprehensive regula ti ons address i ng t h e many uses of reclaimed water There is still concern regarding the use of highly treated wastewater to supplement water supplies One of the concerns is the emergence of waterborne pathogens 12

PAGE 25

Microorganisms in Wastewater Wastewater contains a wide variety of microbial pathogens, i e bacteria protozoa and viruses (Bitton 1994) The pathogenic protozoa of concern include Cryptosporidium and Giardia. Giardia is a common cause of wate r borne disease in the United States (Craun 1988) and in 1985, Cryptosporidium caused the largest waterborne disease outbreak in Milwaukee which affected over 400 000 people (MacKenzie e t al 1994) Cryptosporidium an enteric coccidian protozoan, has been recognized as a pathogen in humans since 1980 (Madore et al1987, Rose 1988 Lisle and Rose 1995 Marshall et al 1997) Cryptosporidiosis an parasitic infection principally of the intestinal tract, causes profuse watery diarrhea abdominal pain nausea vomiting and fever The disease is se l f-limiting in immunocompetent individuals but can be fatal for immunocompromised ind i viduals The oocyst (the environmental stage of the organism) is extremely resistant to conventional disinfection processes and continues to be implicated in waterborne outbreaks throughout the world (Campbell et al1982, Korich et al1990, Peeters et a l 1989) Cryptosporidium oocysts are ubiquitous in the water environment. Rose et al ( 1988) indicated that 91% of the sewage samples examined contained varying levels of oocysts. The oocysts have been shown to survive in flow-through chambers of river water and tap water after 176 days with die-off rates approximately 95% (Lisle and Rose 1995) The fecal material may protect the oocysts from desiccation thus prolonging the viability of the organism. Sewage 13

PAGE 26

discharge could be a significant source of contamination of oocysts in the environment. Giardia, a flagellated protozoa has been recognized as one of the most common parasites of humans in the United States and the most common cause of waterborne outbreaks (Rose et al 1991 Rose et al 1989, Rodgers et al 1995, Hibler and Hancock 1990) Giardiasis causes diarrhea, abdominal distension flatulence and malaise The Giardia cyst is commonly found in raw sewage in fairly large numbers and has also been shown to be resistant to conventional disinfection processes (LeChevallier et al 1991, Hibler and Hancock 1990) Enteroviruses are a group of human viruses that replicate initially in cells of the intestinal tract These small (22 nanometers) viruses include polio virus, coxsackie virus and echo viruses Another enteric virus of concern is hepatitis A virus The enteric viruses cause illnesses such as diarrhea aseptic meningitis, conjunctivitis myocarditis and hepatitis and are found on a routine basis in untreated wastewater (Gerba and Rose1990). The water industry has become sensitive to protozoan contamination of water supplies. The recent promulgation of the Information Collection Rule (ICR) by the Environmental Protection Agency (EPA) will require drinking water facilities serving over 100,000 population to monitor their finished water for Cryptosporidium Giardia and Enteroviruses (USEPA 1994) This has caused a renewed interest in the enteric protozoa and other microbial contaminants in wastewater which may impact water supplies and their control by advanced treatment processes. Cryptosporidium has been found in raw wastewater at 14

PAGE 27

concentrations of 850-13 700 oocysts/L and Giardia at levels of 3375 cysts/L (Rose et al 1996 Madore et al 1987) Wastewater facilities discharging into watersheds serving as drinking water supplies may need to ensure that the effluent is nearly pathogen-free or that levels of these microorganisms have been reduced to some level of acceptability. Current Indicator Microorganisms for Evaluating Water Quality and Treatment Current regulations in most states for discharg i ng wastewater are based on the concentration of coliform bacteria in the final wastewater effluent (Crook and Surampalli 1996 US EPA 1992) Coliforms are classified as total or fecal coliforms Total coliforms are any aerobic or facultat i ve anaerobic gram negative non-spore fo r ming bacillus that ferment lactose and gas at 37 C after 24 hours (Standard Methods for the Examination of Water and Wastewater 1992) Total col i forms are the presumpt i ve ind i ca tor of fecal contamination and are used in drinking water as a standard for d i s i nfection The fecal coliform a subgroup of the total coliform group, are normal inhabitants of the human and animal intestines. Fecal coliforms are differentiated from total coliforms by incubation at an elevated temperature of 44. 5 C (Standard Methods for the Examination of Water and Wastewater 1992) The presence of fecal coliforms confirms fecal contamination and indicates the increased possibility of water contamination by enteric pathogens The absence of fecal coliforms does not 15

PAGE 28

necessarily guarantee that the pathogens are not present. Coliform bacteria have been the indicators of choice for evaluating disinfection processes during water treatment. Numerous researchers have demonstrated that the coliform standard is not adequate for evaluating the effi cacy of treatment primarily disinfection of viruses and protozoa (Baker and Hegarty 1997 Snowdon and Cliver 1989 Fujioka and Shizumura 1985 Dutka 1973, Funderberg and Sorber 1985 Gerba et al 1979 Metcalf 1978 Cabelli 1977) Viral and protozoan waterborne outbreaks have occurred with drinking water supplies that met current U.S. EPA standards for total coliforms and turbidity (Gerba and Rose 1990 Rose et al 1985 MacKenzie et al 1994 LeChevallier et al 1991 Seligmann and Reitler 1965 Boring et al 1971 Keswick et al 1985, Payment and Armon 1989) Few studies have been done in wastewater in an attempt to correlate indicator coliform bacteria with viruses or protozoa Alternative Indicator Microorganisms Alternative microorganisms such as Enterococci Clostridium perfringens and F-specific coliphage, have been proposed as better indicators of water quality, fecal pollution and public health risks (Cabelli 1977 Fujioka and Shizumura 1985, Armon and Kott 1996 Snowdon and Cliver 1989 Grabow 1990) However little data are available on the use of alternative indicator microorganisms (Enterococci Clostridium perfringens and coliphage) compared 16

PAGE 29

to the conventional indicator microorganisms (total and fecal coliforms) for indicating the presence of pathogens in wastewater effluent. Enterococcus is a subgroup of the fecal streptococci bacterial cocc i of fecal orig i n found i n both an i mals and humans Taxonomically these bacteria possess the group D antigen and conform to the Sherman criteria (C l ausen et al 1977) The enterococcus group includes Streptococcus faecium S faecalis S durans and related biotypes (Clausen et al 1977) Enterococcus generally appears to be more pers i stent than either bacterial pathogens or fecal coliforms (Cohen and Shuval 1973, Dav i es-Galley et al 1994 Sinton et al 1994) C perfringens is an enteric gram posit i ve anaerobic spore-fo r ming pathogenic bacterium found in feces. Although there were considerable controversies about using Clostrid i um as a water quality indicator (Cabelli 1977) more recently a number of scientists (Fujioka and Shizumura 1985 Payment and Franco 1 993) recommend C perfringens as a valuable supplement to other water quality tests due to its spore-forming property particularly in situations where detection of viruses or remote fecal pollution i s desirable Th i s microorganism is consistently present in wastewater at concentrations of 1 03 to 104 colony-forming units (CFU)/1 00 ml, and its resistance to chlorination and other environmental factors is similar to the enter i c viruses and protozoa (Fuj i oka and Shizumura 1985 Payment and Franco 1993) F-specific coliphage is a virus that infects E. coli bacteria and can be found in fecally contaminated water These coliphages contain RNA and one of the host specific characterist i cs is the i r adsorption to long filamentous structures 17

PAGE 30

the F-pili on bacteria (Snowdon and C l iver 1989) Coliphages are acellular and approximately the same size as Enteroviruses so t hey have been suggested as adequate model organ i sms for enteric viruses in water (Have l aar et al 1993 Funderberg and Sorber 1985) Development of the Upper Occoquan Sewage Authority In the late 1960 s increased populat i on growth in the Occoquan Watershed led to the degradation of the Occoquan Reservo i r The V i rginia Water Control Board developed a Policy for Waste Treatment and Water Quality Management in the Occoquan Reservoir' in 1971 because of this degradation This policy mandated the construction of the Upper Occoquan Sewage Authority (UOSA) Water Reclamat i on Plant a state of t he art treatment facility to reclaim all the wastewater generated in the watershed (Robbins 1993) The Upper Occoquan Sewage Authority Water Reclamation Plant has been reclaiming wastewater and discharging to the Occoquan Reservoir s i nce 1978 (Robb i ns 1 985). Th i s reservoir serves as a potable water supply for approximately one million people in Northern Virginia (Asano and Levine 1995, Asano and Levine 1996 Asano 1995 McEwen and Richardson 1996 Pia et al1996). This is one of two planned surface wate r augmentation projects currently in operation in the United States at this time ( Pia e t al1996) Ten to fifteen percent of the reservo ir i s compr i sed of recla i med water on average but during t i mes of drought as much as 90% of the flow into the reservo i r comes from the plant d i scharge (McEwen 18

PAGE 31

and Richardson 1996) Treatment of the water includes primary and secondary treatment along with five advanced wastewater treatment processes including chemical treatment with high lime and recarbonation multimedia filtration granular activated carbon filtration ion exchange and chlorination/dechlorination Treated water is blended with the supply in the reservoir and later treated in a conventional water treatment plant before delivery to customers in that area. Although in operation for many years no studies had been undertaken on the removal of pathogenic microorgan i sms from this reclamation plant. 19

PAGE 32

CHAPTER2 RESEARCH OBJECTIVES The objectives of this project were to evaluate the removal of microorganisms commonly found in wastewater through processes at the UOSA advanced water reclamation facility The specific objectives of this study were to: 1) Examine the removal of bacteria (rout i ne indicators and alternative indicators) protozoa human viruses and coliphage and determ i ne which unit process demonstrated the greatest reduction of microorganisms 2) Determine i f the use of alternative microorgan i sms provided data to better reflect the occurrence of pathogens in wastewater. 3) Evaluate a pilot plant for the removal of Cryptosporidium This study took place at the Upper Occoquan Sewage Authority (UOSA), a 27 million gallons per day (mgd) Water Reclamation Plant located in Northern Virginia. An evaluation of the treatment processes within UOSA was necessary to determine if microbial levels, particularly protozoa and viruses, were reduced in the final effluent. The comparison of the treated effluent with the water quality in the Occoquan reservoir was a major 20

PAGE 33

objective of the study The impact of future regulations such as the Information Collection Rule (ICR), was an important issue in initiating this study 21

PAGE 34

Sampling Sites CHAPTER 3 MATERIALS AND METHODS UOSA treatment system consists of a series of barriers : conventional treatment followed by five advanced treatment processes (Figure 1 ) Samples were collected once per month for one year (from April 1995 through April 1996 with the exception of December 1995) from the eight sites within the UOSA Water Reclamation Plant. A high flow event associated with a large rainfall was sampled in January 1996 Each of eight sites were mon i tored for bacteria, viruses and protozoa. Sampling sites included the headworks untreated wastewater after preliminary screening (001 ) ; secondary effluent (011 ) ; second stage recarbonation effluent (020) ; multimedia filter influent after passage through the open ballast ponds (017); multimedia filter effluent (021 ) ; ion exchange bed effluent (023) ; the dechlorinated final effluent (033) ; and UOSA's final effluent reservoir (060) (Table 1 ) The ion exchange system did not operate in the ammonia-removal mode during this study but did operate as a post GAC treatment filter The data collected were used to assess the levels of microbial contaminants entering the plant the levels after the various treatment processes, and the reduction of microorganisms through the unit processes. The methods used are summarized in Table 2 and described in detail thereafter 22

PAGE 35

N w Figure 1. Schematic of UOSA Process Description ,,.. Conventional Tre a tment Advanced 001 Sertlnlng and Grit Biolog i cal Rttyc le filler Headwork p, ..... Olgut er l Compo ttlng a .......; obil Compost Filter Prttlltt 021 H .. dworke l OeweCtrtd Soli d to Land Dltpo181 or Lendllll .. Exchlngt Flntl Elllutnt RtttiYOir Chlorlnttlon/ Otchlorlnttlon 060 033

PAGE 36

Table 1 Description of Sampling Sites within UOSA Reclamation Fac i lity SITE UNIT PROCESS 001 Headworks untreated wastewater post preliminary screening 011 Secondary effluent 020 Second stage recarbonation post lime treatment (high pH 11. 3) 017 Filter influent post carbonation post passage through open ballast pond 021 Multi-media filtered effluent 023 Granular activated carbon (GAC) contactor effluent 033 Final plant effluent post dechlorination 060 Final effluent reservoir Table 2 Summary of Microorganism and Method ORGANISM Enterococci Total col i forms Fecal coliforms Clostridium METHOD Membrane filtration mE agar/EIA agar Membrane filtration mEndo agar Membrane filtration mFC agar Membrane filtration mCP agar REFERENCE Standard Methods for the Examination of Water and Wastewater 9230C Standard Methods for the Examination of Water and Wastewater 92228 Standard Methods for the Examination of Water and Wastewater 92220 Armon and Payment 1988 ................ Enterovirus 1 MDS filter US EPA 1994 Adsorption elution BGM cells -c/YiJiaspandlum .. ---cart"rra9e-ti"itraiiO n-------Ros eaTaf 1 9 9 f--IFA Giardia Cartridge filtration Rose et al 1991 IFA 24

PAGE 37

Microbiological Sampling Bacteria For each sampling event a single one-liter grab sample was collected in a sterile 1 liter sample bottle from each site for bacterial and d i rect coliphage analyses Samples were analyzed upon collection for the bacteria at the UOSA laboratory (Table 2). The membrane filtration technique detailed in Standard Methods for the Examination of Water and Wastewater was used to determine Enterococci densities (Standard Methods for the Examination of Water and Wastewater 1992 Method 9230C). One-hundred ml aliquots of each sample were assayed in duplicate and up to 200 ml volumes were sampled for disinfected water Dilutions of the samples were used as necessary for s i tes 001, 011 and 017 The membrane filters were p l aced on mE agar (Difco Detroit Ml) and incubated for 48 hours at 41C The membranes were then transferred to EIA agar (Difco, Detroit Ml) and incubated at 41 C for 20 minutes. The pink to red colonies were evaluated for a black or reddish-brown precipitate on the of the filter and identified as enterococci. A membrane filtration technique was used to determine Clostridium perfringens densities (Payment and Franco 1993 Cabelli 1977) One-hundred ml aliquots of each sample were assayed in duplicate as previously described The membrane filters were placed on mCP agar (Acumedia Baltimore MD) 25

PAGE 38

and incubated at 45C in an anaerobic gas-pak jar for 18-24 hours. The plates were placed in a ziploc bag containing a petri dish filled with ammonium hydroxide for 1 minute to identify C perfringens colonies Colonies that turned a bright pink were identified as C perfringens The membrane f i ltration technique detailed in Standard Methods 92228 and 9222D (Standard Methods for the Examination of Water and Wastewater 1992) was used to determine total coliform and fecal coliform densities respectively One-hundred ml aliquots of each sample were assayed in duplicate as previously described The membrane filters were placed on M Ende medium (Difco Detroit Ml) for total coliforms and incubated at 35 C for 24 hours Red colonies that produced a metallic green sheen were identified as total coliforms The membrane filters were placed on M -FC medium (Difco Detroit Ml) for fecal coliforms and incubated in a water bath at 44. 5C for 24 hours Blue colonies were identified as fecal coliforms Protozoa A slightly modified version of the ICR protozoan protocol was utilized for the collection and detection of Cryptosporidium oocysts and Giardia cysts (Federal RegisterNol. 59, No 28/February 10, 1994 Appendix C to Subpart M Proposed ICR Protozoan Method for Detecting Giardia cysts and Cryptosporidium oocysts in Water by Fluorescent Antibody Technique) Protozoan samples were collected by filtration through a 1.0 um nominal 26

PAGE 39

porosity 10-inch yarn wound cartr i dge filter (Microwynd AMF Cuno Balt i more MD) The volume of water filtered was monitored by attached flow meters After collect i on the filters were put in ziploc baggies and placed on ice for transport to the University of South Florida Upon receipt at the University of South Florida the filter was cut and washed to recove r the accumulated debris and density gradients were used to separate the oocysts and cysts from the sediments. The final concentrate was filtered onto 0.22 urn cellulose acetate membrane filters in duplicate and stained with monoclonal antibodies. These monoclonal antibodies are tagged with a f l uorescent label fluorescein isothiocyanate (FITC), which specifically binds to the oocyst and cyst wall and when examined using epifluorescen t microscopy the oocyst or cyst fluoresce green. Equ i valent volumes from the concentrates which were examined under the microscope were calcu l ated and the concentration of cysts and oocysts per 100 L were determined Human Viruses All samples were analyzed for enterov i ruses by Dr Sam Farrah at the University of Florida The Proposed Virus Monitoring Protocol was utilized for the collection and detection of human enterovi r uses (Federal RegisterNol. 59 No 28/February 10, 1994 Appendix D to Subpart M Proposed Virus Monitoring Protocol) Samples were collected by filtration through a 1 0-inch pos i tively charged pleated cartridge filter designed to capture viral particles (1 MDS 27

PAGE 40

George Edwards Company Pelham AL) The volume of water filtered was monitored by attached flow meters After collection the filters we r e secured in ziploc baggies and placed on ice for transport to the Univers i ty of F l orida. The human enteric viruses were eluted from the filters usi n g beef extract and concentrated by a flocculation method. The v i ruses were grown on BGM cell culture i n 75 cm2 flasks The flasks were evaluated daily for cell destruction caused by viruses Positive and negat i ve cells were passaged once into new cells for confirmation An MPN method was used to enumerate the concentration of enteroviruses Coliphage An aliquot of water f rom the 1 liter grab samp l e was ana l yzed using an aga r overlay technique described by Adams 1959 Escherichia coli (E. coli American Type Culture Collection ( ATCC) #15597 Rockville MD ) was used as the bacterial host and was grown to stationary phase 24 hours before each coliphage assay. A one to two -ml aliquo t of the sample was added to a tube containing 3 ml of tryptic soy broth (Difco Detroit Ml) containing 1 5% agar (Difco Detro i t Ml) kept liquid at 48 C then 0 1 ml of host bacteria was added mixed and poured onto a tryptic soy agar petri plate. Replicate plates were set up in order to assay 8 0 ml from most sites. Dilutions ( made with sterile phosphate buffer solution ) of the samples were necessary for untreated water The plates were incubated at 37 C for 24 hours After 24 hours the petri plates 28

PAGE 41

.were removed from the incubator and examined for the presence of plaques (clearings in the bacterial lawn) Plates containing less than 300 plaque-forming units (PFU) were counted Pilot Studies A pilot facility representing the chemical treatment process was built at UOSA and two separate challenges were conducted (Figure 2) Initially the pilot facility was evaluated for detention time using a fluorogenic dye It was determined that the flow rate of 3 gpm simulated the surface loading rate and the flow rate of 7 gpm simulated the hydraulic detention time of the full-scale system. The actual and theoretical detention times were 130 and 150 minutes respectively for the 3 gpm flow rate and 54 and 64 minutes respectively for the 7 gpm flow rate. The pilot plant influent water was secondary effluent drawn from the line feeding the full-scale facility This was seeded during the first trial with fluorescent beads (3 um in diameter as a surrogate for Cryptosporidium oocysts) and two bacterial viruses, MS2 coliphage and PRD1 bacteriophage Fifty (50) ml each of MS2 stock and PRD1 stock (1011/ml) and one ml of approximately 109 fluorescent beads were added to 1 L of secondary effluent in a 3 L carboy. The influent was injected over a 6-minute time period with a peristaltic pump An influent sampling port was set up and samples were collected every 30 seconds to 1 minute Effluent samples were collected in 500-ml bottles every 30 minutes 29

PAGE 42

w 0 Isolation Valve From Rapid Mix In Figure 2. Schematic of Pilot Plant Flowmeter Roll Two 90 ELS Sample Wlthdrawl Teo w/flex, tygon & pinch clamp Polymer InJection Filling Sludge Drain w/Valva Sampling Tee w/ Valve to Plant Waste

PAGE 43

for 2 to 3 hours. The effluent samples were neutralized with 1 N sulfuric acid immediately upon collection The second pilot study was seeded with MS2 coliphage and PRD1 bacteriophage and formalinized Cryptospor i dium parvum oocysts. One hundred(100) ml each of MS2 stock and PRD1 stock (1 011/ml) were added to 1 L of secondary effluent in a 3 L carboy. Five ml of 1 08/ml formalinized Cryptosporidium parvum oocysts was added approximately 1 minute after the injection of the phage to prevent the inactivation of MS2 and PRD1 due to the formalin present in the oocyst stock solution The samples were collected as previously described MS2 coliphage (ATCC catalog number 15597-81) were propagated for use in the pilot studies by inoculating a 1 L flask containing 200 ml of tryptone yeast extract (TYE) with 2 0 ml of the host bacteria. Escherichia coli (E. coli) (ATCC catalog number 15597) The culture flask was _placed in a shaking incubator maintained at 37C When the bacterial density reached approximately 1 08/ml colony forming units/ml -(CFU/ml) (wh.ich had been previously determined) an aliquot of the virus stock (approximately 1 012/ml PFU/ml), was added to provide a multiplicity of infection (MOl) of 0 1 The culture was shaken continuously unt i l the host cells lysed Then 02 g. of lysozyme and 6 0 ml of sterile 0 2M ethylenediaminetetraacetic acid (EDTA) were added to the culture to lyse the host cells and release tbe virus (bacteriophage) and the sample was incubated for an additional 30 minutes i n a shaking incubator at 37 C. The propagated virus and cellular debris were then centrifuged for 20 minutes at 3600 31

PAGE 44

rpm and filtersterilized using a 0.45 urn sterile membrane filter. The resulting stock was titered by the agar overlay technique and refrigerated at 4C until needed PRD1 bacteriophage was propagated in the same manner with the host bacteria Salmonella Pilot influent samples were diluted and effluent samples were assayed directly for the bacteriophage using an agar overlay technique with each of the respective hosts as previously described (Adams 1959). Plaques were enumerated after 24 hours incubation at 3JOC. Samples containing oocysts were filtered and stained with monoclonal antibodies The inf l uent samples were assayed directly (0.1 to 1.0 ml) for the oocysts by filtering the sample onto 0 .22 urn cellulose acetate membrane filters (Sartorius) using a Hoefer manifold and stained with monoclonal antibodies (Crypt-a-Gio, Waterborne Inc. New Orleans LA). These monoclonal antibodies are tagged directly with a fluorescent label, fluorescein isothiocyanate (FITC) which specifically binds to the oocyst wall and when examined using epifluorescent microscopy the oocysts fluoresce green. The membrane filters were mounted onto a glass slide using 2% 1 ,4-Diazabicyclo[2 2 2) octane (DABCO) and a coverslip and the entire membrane filter was counted under 400X epifluorescent microscopy. A similar procedure was used for the fluorescent bead samples except the monoclonal antibodies were not necessary The sample was filtered onto a membrane filter mounted onto a glass slide with DABCO and examined microscopically The effluent samples were concentrated by centrifugation at 1050 x g if necessary or 1 0 to 20 ml of sample were filtered 32

PAGE 45

onto 0 22 um cellulose acetate membrane filters in duplicate and then stained with monoclonal antibodies The effluent samples for the fluorescent beads wer e processes similarly with the exception of staining with monoclonal antibodies Fluorescent beads and oocysts/ml were calculated in the influent and the effluent. 33

PAGE 46

UOSA Treatment Processes CHAPTER4 RESULTS The UOSA treatment system consists of a series of processes : conventional and secondary activated sJudge treatment followed by five advanced treatment processes (Robbins 1993) Primary treatment consists of screening grit removaJ, and primary clarificat i on. Secondary treatment is provided by a complete-mix activated sludge system comprised of biological treatment clarification and return activated sludge pumping The act iv ated sludge system includes pre-aeration basins (selectors) that promote a more efficient microbial culture. In the biological reactors microorganisms (activated sludge) decompose organic pollutants and form a biological floc which is separated from the water by settling in the secondary clarifie r s A portion of the settled floc is returned to the selectors to activate the biological reactions The remainder is combined with primary sludge and transferred to the anaerobic digestion system The digested sludge is currently dewatered and composted. The conventional system is followed by high pH chemical treatment, filtration, granular activated carbon (GAC) adsorption ion exchange 34

PAGE 47

and disinfection using sodium hypochlorite followed by dechlorination using sulfur dioxide. The high pH chemical treatment system includes rap i d mix basins flocculation basins chemical clarification first stage recarbonation clarification, and second stage recarbonation. Calcium hydroxide is the principal coagulant and carbon dioxide is used for recarbonation Mechanical mixers in the rapid mix basins completely mix the coagulant with the wastewater The pH increases to 11. 3 thereby enhancing inactivation of some microorganisms and precipitating phosphorus heavy metals and suspended solids (Grabow et al 1978) The small coagulated particles (floc) are then slowly stirred in the flocculation basins to promote aggregation into larger particles, which settle more readily in the chemical clarifiers Following chemical clarification the pH is restored to neutral by two stage recarbonation with intermediate clarification In first stage recarbonation, the pH is adjusted to 9 7 which causes precipitation of carbonates and other materials. The carbonates form a small dense floc which settle readily in the recarbonation clarifiers After recarbonation clarification more carbon dioxide is added to adjust the water to pH 7 The chemical treatment effluent flows into ballast ponds for flow equalization and is pumped at a uniform rate through the remaining advanced treatment processes Multimedia pressure filters are used to remove suspended sol i ds and turbidity A backwash equalization tank prevents hydraulic surges Granular activated carbon (GAC) adsorption performed in an upflow expanded bed removes a wide range of synthetic organic compounds GAC loses its adsorptive 35

PAGE 48

capacity over time and is periodically regenerated in an on-site multiple-hearth furnace The ion exchange columns remove ammonia and activated carbon fines UOSA S unoxid i zed nitrogen standard (1. 0 mg/L) is normally met by nitrification in the secondary treatment systems ; but during drought conditions nitrogen is removed in ammonia form by the ion exchange system The ion exchange system does not operate in the ammonia-removal mode very often but can operate as a post GAC treatment filter Disinfection is accomplished by chlorination Sodium hypochlorite is applied to the ion exchange effluent at the chlorinat i on mix station The mixing station is capable of breakpoint chlorinating ammonia remaining after nitrification or ion exchange treatment. The chlorinated water then flows through the chlorine contact chamber The contact chamber usually contains the effluent for 30 minutes of contact time and the average total residual chlorine (TRC) after contact time is 1 5 mg/L with a standard deviation of 0.3 mg/L and the free residual chlorine (FRC) average is about 70% of the TRC. The water is then dechlor i nated with sulfur dioxide prior to discharge into UOSA s final effluent reservoir. Bacteria Clostridium perfringens were detected at an average of 3. 7 x 104 and 4.4 x 1 03 CFU/1 00 ml in the untreated and secondary wastewater respectively The 36

PAGE 49

second stage recarbonation had an average of 5.1 CFU/1 00 ml, and the multi media influent averaged 16 5 CFU/100 ml. The multi-med i a effluent and the carbon adsorption effluent both averaged 1 8 and 2.2 CFU/1 00 ml ( Table 3 ). Clostridium perfringens was detected in the final effluent post dechlorination at an average concentration of 3 5 CFU/1 00 ml in 1 0% of the samples and at an average concentration of 4 9 CFU/1 00 ml in 78% of the final effluent reservoir samples (Figures 3 and 4) Clostridium was reduced by the high pH chemica l treatment by approximately 3 log10 (99. 9%) and the overall reduction for Clostridium was 5 log10 (99 999%) (Figure 5) The enterococci were detected i n the untreated wastewater at an average of 5 0 X 105 CFU/100 ml, an"d lhe secondary effluent averaged 2 2 X 103 CFU/1 00 ml. Enterococci were reduced to average levels of 23. 5 1 06.4 12 and 10. 8 CFU/1 00 ml in the second stage recarbonation multi-media influ e nt and effluent and carbon adsorption effluent respectively (Table 4) Ente r ococci were detected in the final effluent post dechlorination at an average concentration of 2 7 CFU/1 00 ml in 17% of the samples and at an average concentration of 10 1 CFU/1 00 ml in 73% of the final effluent reservoir samples (Figures 3 and 4). The overall reduction for Enterococci was 6 log10 (99 9999%) (Figure 5) Total coliforms were detected in untreated wastewater and secondary effluent at averages of 2.4 x 107 and 1 7 x 105 CFU/1 00 ml, respectively. Levels were reduced from 121 CFU/100 ml after second stage recarbonation to an average of 1.6 CFU/1 00 ml in the final effluent postdechlorination (Table 5) 37

PAGE 50

Four of 11 samples were positive (36%) for total coliforms in the final effluent postdechlorination and average levels increased in the final effluent reservoir to an average of 180 3 CFU/1 00 mL in 100% of the samples (Figures 3 and 4) The overall reduction for total coliforms was 7 log10 (99 99999%) (Figure 5) Fecal coliforms were detected in the untreated wastewater and secondary effluent at averages of 9 0 x 105 and 7 8 x 103 CFU/1 00 mL respectively (Table 6) Levels were reduced from 23 CFU/1 00 mL to <0 5 CFU/1 00 mL in the final effluent dechlorination Levels increased to 568 CFU/1 00 mL prior to the multimedia sand filter (post passage through the open ballast pond) and also increased in the final effluent reservoir to an average of 22 9 CFU/1 00 mL in 100% of the samples (Figures 3 and 4) Conventional treatment and chemical treatment achieved a total 4 6 log10 reduction of fecal coliforms The overall reduction for fecal coliforms was 6.2 log10 (>99 9999%) (Figure 5) 38

PAGE 51

w \0 Table 3 Clostridium perfringens (CFU/100 mL) Sample site ----> 001 011 020 017 021 023 033 060 # of samples 9 9 10 10 10 10 10 9 # positive 9 9 9 9 7 5 1 7 % samples positive 100% 100% 90% 90% 70% 50% 10% 78% Sensitivity Limits < 0 5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 Minimum CFU/100 mL 15,000 311 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 Maximum CFU/100 mL 57,000 28,000 12 95 6.0 6.0 3. 5 12.5 Mean (arithmetic) : Positives only 36,667 4,452 5.1 16.5 1.8 2.2 3.5 4.9 All samples 36,667 4,452 4.6 14.9 1.3 1.1 0.35 3.8 REMOVAL EFFICIENCY UNIT PROCESS % 87.85 99.9 -224 91.28 15.4 68.18 N / A CUMULATIVE % 87.85 99.99 99.96 99.996 99.997 99.999 N/A Clostridium perfringens; a gram positive, spore forming, capsulated, gas producing, nonmotile, anaerobic bacillus and is a normal inhabitant of the intestinal tract of man and animals. Food poisoning is caused by c perfringens, in rare cases intestinal gas gangrene. The data show the greatest absolute removal of C. perfringens was in conventional treatment, but the high pH chemical treatment provided the greatest log reduction. Removal efficiencies for processes following chemical treatment were perhaps underestimated to some extent by very low influent counts. ,. ,. L

PAGE 52

0 Table 4 Enterococci (CFU/100 mL) Sample site ----> 001 011 020 017 021 023 033 060 # of samples 12 12 11 12 12 12 12 11 # positive 12 12 10 10 8 10 2 8 % samples positive 100% 100% 91% 83% 83% 83% 17% 73% Sensitivity Limits < 0.5 < 0.5 < 0.5 < 0.5 < 0 5 < 0.5 < 0.5 < 0.5 I I Minimum CFU/100 mL 50,000 233 < 0.5 < 0.5 < 0.5 < 0 5 < 0.5 < 0.5 Maximum CFU/100 mL 870,000 11,000 80 370 45.5 59 4.5 61 Mean (arithmetic) : Positives only 500,000 2,183 23.5 106.4 12 10.8 2 7 10. 1 All samples 500,000 2,183 21.4 88.7 10 9 0.45 7.4 REMOVAL EFFICIENCY UNIT PROCESS % 99.56 99.0 -314 88.73 10.0 95. 0 N/A CUMULATIVE % 99.56 99.996 99.98 99.998 99.998 99.9999 N/A Enterococci is a name commonly used for some bacteria within the genus Streptococcus. These organisms are generally found in human and animal feces. Enterococcus appears to be more persistent in the environment than either bacterial pathogens or fecal coliform. The data show conventional treatment ( 011) and chemical treatment (020) had about equal removal efficiency for enterococci. Together these two unit processes achieved a 4.4 log1 0 reduction of enterococci. On average, the UOSA plant achieved a 6 log1 0 reduction of enterococci during this study. N/A -Not applicable

PAGE 53

..... Table 5 Total coliform (CFU/100 mL) Sample site ----> 001 011 020 017 021 023 033 060 I # of samples 10 10 11 10 11 11 11 6 # positive 10 10 11 10 11 11 4 6 % samples positive 100% 100% 100% 100% 100% 100% 36% 100 Sensitivity Limits < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 <0.5 Minimum CFU/100 mL 2.4*106 24,000 0.5 20 5.5 2.0 < 0.5 17 Maximum CFU/100 mL 42*106 610,000 800 >16 1 000 >1, 6oo >1,600a 4.0 600 Mean (arithmetic) : Positives only 24*106 170,000 121 1,822 178 166 1.6 180 All samples 24*106 170,000 121 1,822 178 166 0.58 180 REMOVAL EFFICIENCY UNIT PROCESS % 99.29 99.93 -1,701 90.23 6.74 99.65 CUMULATIVE % 99.29 99.999 99.992 99.999 99.9993 99.9999 99 Total coliform are any aerobic or facultative anaerobic, gram negative, non spore forming, bacilli that ferment lactose to acid and gas at 37 C after 24 hours. Most of the total coliform species are widespread in the environment. Some total coliform species reside mainly in the intestines of human and animals, and are short lived in the environment. Total coliform are the presumptive indicator of fecal contamination and are used in drinking water as the standard for disinfection. Conventional treatment and high pH chemical treatment had similar removal efficiency for total coliform. These two processes achieved a total 5 log10 reduction of total coliform. The total plant achieved a 7.6 log10 reduction of total coliform on average during this study. Samples were too numerous to count at dilutions used; analyzed by MPN method

PAGE 54

"-' Table 6 Fecal col i f orm (C FU/100 mL) -----Sample site ----> 001 011 020 017 021 023 033 060 # of samples 11 11 10 11 11 10 11 8 # positive 11 11 9 11 10 9 0 8 % samples positive 100% 100% 90% 100% 91% 90% 0% 100 Sensitivit y Limits < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 Minim u m CFU/100 m L 90,000 1,200 0.5 1.0 < 0.5 0.5 < 0.5 3 0 I Maximum CFU/100 mL 5.7*106 41,000 130 5,000 240 130 < 0.5 71 I Mean (arithmetic) : Positives only 900,000 7,764 23 568 37 21 < 0 5 22.9 All samples 900,000 7,764 20.7 568 33.6 18.9 < 0.5 22.9 REMOVAL E FFICIENCY I UNIT PROCESS % 99.14 99.73 -2,744 94.08 43.75 > 97.6 N/A CUMULATIVE % 99.14 99.998 99.937 99.996 99.9979 >99.99999 N/A Fecal coliform, a sub group of total coliform, are normal (usually non-pathogenic) inhabitants of human and animal intestine. Fecal coliform are differentiated from total coliform by incubation a t an elevated temperature of 44.5 C and fermentation of lactose. Fecal coliform is the confirmation indicator for fecal contamination, and indicates a good chance the water may be contaminated by enteric pathogens. Absence of fecal coliforms does not guarantee absence of viruses and protozoa. Conventional treatment and chemical treatment achieved a total 4 6 log10 reduction of fecal coliform. Fecal coliform were not detected in the dechlorinated effluent. N/A -Not applicable -'

PAGE 55

+:w _J E 0 0 ::> lL 0 Figure 3. Average Levels for Positive Samples through the Treatment Train for Bacteria 1 .000E+081 .000E+07 1,000,000 1 0 000 1 ,000. 100-10 1 001 011 020 .... 017 '* 021 023 Sampling Sites 033 ..... Enterococci ->K C lo s t ridium +Tota l Co liform s +Fec al Coliform s 12 samples No fecal coliforms detected in final effluent 060

PAGE 56

""'" ""'" 120-100 Q) > :E 80 (/) 0 0... 60 1 ...... c Q) 0 Q> 40 -0... 20 0 Figure 4. Percentage of Samples Positive through the Treatment Train for Bacteria 001 011 020 017 021 023 033 Sampling Sites """ Enterococci *Clostridium +Total Coliforms Fecal Colifo rms 12 samples Clostridium 1 0% positive in the final effluent Fecal Coliform not detected in final effluent 060

PAGE 57

.!> V1 Figure 5 Log 10 Removal of Sites Com pared to Untreated Wastewater Log10 Removal 10r-------------------------------------. 8 6 4 2 0 011 020 017 021 023 033 S a mpling Sites C l ostrid ium Q h otal Coliforms 0 Fecal Coliforms

PAGE 58

Protozoa Cryptosporidium oocysts were detected in the u n treated wastewater at an average of 1 ,484/1 00 L. No oocysts were detected in the secon d ary effluent but two of 12 samples in the second stage recarbonation were pos i tive at an average concentrat i on of 3.45 oocysts/100 L (Table 7). Cryptosporidium oocysts were detected in one of the 12 samples (8%) in the final effluent post dechlorination a t a concentrat i on of 0.44 oocysts/1 00 L. One of 11 samples were positive for Cryptosporidium at a co n centrat i on of 5. 7 oocysts/1 00 L in the final effluent reservoir (Fi gures 6 and 7) Convent i onal and h i gh pH chem i cal treatment reduced the oocysts by 99.96% Overall reductions for Cryptosporidium were at least 41og10(99.99%) (Figure 8). Giardia cysts were detected in the untreated wastewater and secondary effluent at averages of 4 9 x 104 and 2 3 x 103 CFU/1 00 L, respectively (Table 8). After second stage recarbonation one of 12 samples (8%) in the mult i media effluent was positive for Giardia cysts at a concentration of 61. 3 cysts/1 00 L. Two of 12 samples (17%) were pos i tive for Giardia cysts in the final effluent post dechlorination at an average concentration of 6 6 cysts/1 00 L Levels increased in the final effluent reservo i r to an average of 42 9 cysts/1 00 L i n 18% of the samples (Figures 6 and 7) Conventional treatment and high pH chemical t r eatment achieved a 2.7 log10 reduction and the overall plant achieved a 4.6 log10 reduction (Figure 8) 46

PAGE 59

Human Viruses Enteroviruses were detected in untreated wastewater and secondary effluent at averages of 1 085 and 23.6 PFU/1 00 L. Levels were reduced to 0.2, 0.3 0 116 and 0 123 PFU/1 00 L in the second stage recarbonation, multi-media i nfluent and effluent and GAC effluent respectively (Table 9) Enteroviruses were not detected after chlorination or in the final effluent reservoir (Figures 6 and 7). Overall plant reduction of enterovirus was greater than 41og1D(99 99%) (Figure 8). Coliphage Coliphage were detected in the untreated wastewater at an average of 3 8 x 105 PFU/1 00 ml and 1 8 x 103 PFU/1 00 ml in the secondary effluent. Coliphage were reduced to an average of 28, 142 33, and 25 PFU/1 00 ml through the second stage recarbonation multi-media influent and effluent and carbon adsorption effluent respectively (Table 1 0) Coliphage were not detected in the final effluent postdechlorination but levels increased to 20. 6 PFU/1 00 ml in 45% of the samples in the final effluent reservoir (Figures 6 and 7) Overall reductions across the plant were 4.4 log10 (Figure 8) 47

PAGE 60

.1:' Table 7 Cryptosporidium (Oocysts/100 L) Sample site ----> 001 011 020 017 021 023 033 060 # of samples 12 12 12 12 12 12 12 11 # positive 2 0 2 2 1 2 1 1 % samples positive 100% 0% 17% 17% 8.3% 17% 8.3% 9.1% Sensitivity Limits < 15.0 <12.4 < 1.0 < 1.0 < 0.42 < 0.9 < 1.2 < 1.95 Min oocysts/100 L 277.8 <12.4 2.1 6.4 < 0.42 < 0.9 < 1.2 < 1.95 Max oocysts/100 L 2,690 <2500 4.8 56.3 0.9 2.82 0.44 5.7 Mean (arithmetic) : Positives only 1,484 N / A 3.45 31.3 0.9 2.7 0.44 5.7 All samples 1,484 <312.2 0.575 5.217 0.075 0.45 0.037 0.52 REMOVAL EFFICIENCY UNIT PROCESS % 78.96 99 .816 -807 98.56 -300 0.8370 N / A CUMULATIVE % 78.96 99.961 99.65 99.995 -99.97 99.998 N/A Cryptosporidium is the name assigned to a fairly large variety of very small protozoan oocysts. The oocysts do not normally occur in high concentrations in the natural environment Each oocyst has the potential to release a maximum of four viable Cryptosporidium sporozoites which can initiate infection. Thus, the potential for infection from ingesting one oocyst is quite high. Some oocysts are pathogenic to man, causing chronic diarrhea which in severe cases in the immunocompromised can b e fatal. The data indicate the combination of conventional and high pH chemical treatment reduced the plant influent oocyst population by 99.96%. On average the total plant achieved a 99.998% (4.6 log1 0 ) reduction. N / A -Not applicable

PAGE 61

1.0 T able 8 Giardia (C ysts/100 L) Sample site ----> 0 0 1 0 1 1 020 017 021 023 033 060 # of samples 12 12 1 2 12 12 12 12 11 # positive 12 8 6 2 1 0 2 2 % samples positive 100% 67% 50% 17% 8.3% 0% 17% 18.2% Sensitivity L imits < 15.0 < 10. 0 < 3.0 < 4.6 < 1. 0 < 1.2 < 1.2 < 5.7 M in. cysts/100 L 2,246 24. 5 < 3 0 < 4.6 < 1.0 < 1.2 < 1.2 < 5 7 Max cysts/100 L 142,631 17,671 728 326.8 61.3 < 1.2 12. 8 7.5 Mean (arithmetic): 48,691 2,297 170 220 61.3 < 1.2 6 6 42. 9 Positives onl y 48,691 1,531 8 5 3 6.7 5.1 < 1.2 1.1 7 8 All samples REMOVAL EFF I CIENCY UNIT PROCESS % 96.856 94.45 56.82 86.10 76. 4 > 8.3 N/A CUMULATIV E % 96.856 99.825 99.925 99.989 99.9975 99.998 N / A Giardia is the name for a group of single-celled, flagellated, pathogeni c protozoa found in a variety of vertebrates, including, mammals, b irds, and reptiles. These organisms exist as trophozoites (active or feeding stage form) inside the host intestinal tract or as cysts excreted in the feces, depending on the stage of their life cycle. Giardia lamblia (causes diarrhea) is the clas sical exampl e of the grou p associated with humans a n d its cysts are founc in wastewater in fairly large numbers. Conventional treatment and high pH chemical treatment a chieved similar removal efficiencies for Giardia lamblia. Combined, these t wo processes achieved a 2 7 log10 reduction. On average t h e total p lant achieved a 4.6 log1 0 reduction. N/A -N ot ap p licab l e

PAGE 62

1..11 0 Table 9 Enteroviruses (MPN/100 L} Sample site ----> 001 011 020 017 021 023 033 060 # of samples 1 2 12 12 12 12 12 12 11 # positive 12 1 2 1 3 3 3 0 0 % samples positive 100% 100% 8.3% 25% 25% 25% 0% 0 % Sensitivity Limits < 1.0 < 1. 0 < 0.012 < 0.03 5 < 0 .043 < 0.056 < 0.05 < 0.14 Minimum CFU/100 mL 150 1.4 < 0.012 < 0.035 < 0.043 < 0.056 < 0.05 < 0 .14 Maximum CFU/100 mL 4,600 120 0.22 0.7 0.27 0.18 < 0 .13 < 0.91 Mean (arithmetic} : Positives only 1,085 23.6 0.22 0.3 0.116 0.123 < 0.085 < 0.28 All samples 1,085 23.6 0.018 0.1548 0.09 0.097 < 0.085 < 0 .28 REMOVAL EFFICIENCY UNIT PROCESS % 97.8 99.92 -760 41.86 -7. 8 > 12.4 N/A CUMULATIVE % 97.8 99.998 99.99 99.992 99.991 > 99.992 N/A .. Enteroviruses are a group of viruses that replicate initially in cells of the intestinal tract. These small (22 nm.} viruses include polio virus, coxsackie and echo viruses, and Hepatitis A virus. They are ribonucleic acid (RNA} v iruses. I n this study a cumulative 99.998% rem o val (3. 38 log reduction} of enteroviruses was achieved by the combination of conventional and c h emical treatment. Enteroviruses were not detected in dechlorinated final effluent. The plant overall removal efficiency was 99.999% or a 4.3 log10 reduction. N/A -Not applicable

PAGE 63

VI ._. Table 10 Coliphage (PFU/100 mL) Sample site ----> 001 011 020 017 021 023 033 060 # of samples 12 12 12 12 12 12 12 11 # positive 12 12 4 8 3 1 0 5 % samples positive 100% 100% 33% 67% 25% 8.3% 0% 45% Sensitivity Limits < 12.5 < 12.5 < 12.5 < 12.5 < 12.5 < 12. 5 < 12.5 < 12.5 Minimum CFU/100 mL 60,000 200 < 12.5 < 12.5 < 12.5 < 12. 5 < 12.5 < 12.5 Maximum CFU/100 mL 860,000 5,500 75 790 75 25 < 12.5 50 I Mean (arithmetic) : Positives only 380,000 1,821 28 142 33 25 < 12.5 20. 6 All samples 380,000 1,821 9.3 94.6 8.3 2.1 < 12.5 9.4 REMOVAL EFFICIENCY UNIT PROCESS % 99.52 99.49 -1017 91.226 74.7 50 N / A CUMULATIVE % 99.52 99.998 99.975 99.998 99.999 99.999 N / A Coliphage are viruses that use Escherichia coli bacteria as their host. The direct method detected large numbers of coliphage in the plant influent. The method did not detect Coliphage in the dechlorinated effluent, thus the reported mean values ( < 12. 5 PFU/100 milliliter) are a function of the method sensitivity. The data seem to suggest direct method assay may be more appropriate for highly contaminated water than for clean water. The p lant achieved a 4.4 log10 reduction of coliphage as measured by the direct assay method. N/A -Not applicable

PAGE 64

lJl N Figure 6 Average Levels for Positive Samples for Enteroviruses, Protozoa and Coliphage* 1 000 000 _J 100,000 0 0 T" 10, 000 (/) ... (/) >. 0 0 1 000 0 100 (/) (/) >. 0 10 lL 0... 1 -. x x. .... . 0 1 001 011 020 017 021 023 Sampling Si tes *Phage *Ente r ovi ru s X Cryptosporidium Giardia Coliphage results PFU/1 00 ml No enteroviruses detected in final effluent or reservoir No coliphage d ete c t ed in final e ffluent ........ x-033 ... ... 060

PAGE 65

1.11 w Q) ;!::: en 0 a.. ..... c Q) 0 .... Q) a.. Figure 7. Percentage of Samples Posit i ve for Enteroviruses, Protozoa and Coliphage 120 100. \ \ \ 80 \ \ \ \ ..... 60 40 20 --.:_ 0 ,, 001 011 020 017 021 023 033 Sampling Sites *Coliphage Enterovirus XCryptosporidium Giardia 12 samples 060

PAGE 66

Vl Figure 8. Log1 0 Removal of Sites Compared to Untreated Wastewater Log1 0 Removal 6.-------------------------------------, 5 4 3 2 1 0 011 020 017 021 023 033 Sampling Sites .Phage [8) Ent e r ovirus Q Cryptosporidium !ZIGiardia

PAGE 67

Pilot Studies Tables 11 and 12 show the results of the fluorescent bead and bacteriophage experiments at 7 gpm and at 3 gpm. Tables 13 and 14 show the results for the oocyst and bacteriophage experiment at the same two flow rates The influent total numbers were calculated by multiplying the average influent levels (#s/ml) with the total time Figures 9 1 0, 11, and 12 diagram the removal of fluorescent beads Cryptosporidium oocysts MS2 and PRD1 bacteriophages respectively by chemical lime treatment at 7 gpm and at 3 gpm. The influent was injected over 4 minutes and monitoring showed levels of 1 x 1 04 fluorescent beads/ml after 1 minute (Figure 9) The retention time in the pilot faciliity was 2 hours Beads were first detected in the effluent after 30 minutes and ranged in levels between 1 and 36/ml. After 3 hours the level dropped off to zero. Cryptosporidium oocysts were detected in the influent at concentrations of 6 x 1 02/ml after 3 minutes (Figure 1 0) Oocysts were also first detected after 30 minutes and ranged from 0 02 to 0 .22/ml. Similar results were observed for MS2 and PRD1 bacteriophage with influent concentrations of 1 x 1 08 PFU/ml after 1 minute (Figures 11 and 12) PRD1 bacteriophage was detected first in the effluent after 5 minutes and at higher concentrations than MS2 The bacteriophage were inactivated by the high pH which was maintained at 11. 2 throughout the experiments MS2 coliphage was more sensitive to inactivation with 99.9999% reduction as compared to PRD1. A large difference was observed between the two sets of experiments for PRD1. PRD1 was inactivated 55

PAGE 68

99. 999% in the first experiment ; only 94 to 95% inactivation was observed i n the second experiment. This may have been due to better attention to the neutralization of the high pH immediately upon collect i on of the sample in the second experiment. The removal of the formalinized oocysts ranged from 99% to 99.65% while the beads at either flow rate were removed by approximately 98% 56

PAGE 69

Table 11 Pilot Scale Study on the Removal of Beads and Phage by Chemical Lime Treatment at a Flow Rate of 7 gpm Fluorescent MS2 Phage PRD1 Phage Beads Average Influent 4 25 X 10;j 9 3 X 101 5 88 X 101 levels (#s/mL) Time (minutes) 5.17 5 17 5 17 Influent Total 2 197 X 104 4 .82 X 108 3 04 X 108 Numbers Average Effluent 6 67 0 0 1 15 X 10 levels (#s/mL) Time (minutes) 65 65 30 Effluent Total 433 150 3.45 X 10 Numbers Percent 98.02 > 99 99996 98 86 Reduct i ons Table 12 Pilot Scale Study on the Removal of Beads and Phage by Chemical Lime Treatment at a Flow Rate of 3 gpm Fluorescent MS2 Phage PRD1 Phage Beads Average Influent 1 13 X 104 2 3 X 10 3.47 X 10 levels (#s/mL) Time (minutes) 5 75 5 75 5 .75 Influent Total 6 .5x104 1 33 X 109 1.997 X 10 Numbers Average Effluent 14.4 85.4 10 levels (#s/mL) Time (minutes) 210 210 210 Effluent Total 3024 1 79 X 104 2100 Numbers Percent 95 35 99.998 99.9989 Reductions 57

PAGE 70

Table 13 Pilot Scale Study on the Removal of Cryptosporidium and Phage by Chemical Lime Treatment at a Flow Rate of 7 gpm Cryptosporidium MS2 Phage PRD1 Phage Average Influent 296 1 59 X 105 1 03 x 101 levels (#s/ml) Time (minutes) 7 3.5 3 5 Influent Total 2072 5.55 X 105 3.6 X 107 Numbers Average Effluent 0.12 9 56 2 .74x104 levels (#s/ml) Time (minutes) 60 60 60 Effluent Total 7 2 573.75 1 .64 X 10 Numbers Percent 99 65 99 99989 95.4 Reductions Table 14 Pilot Scale Study on the Removal of Cryptosporidium and Phage by Chemical Lime Treatment at a Flow Rate of 3 gpm Cryptosporidium MS2 Phage PRD1 Phage Average Influent 513.2 1.9x 105 7.95 x 101 levels (#s/ml) Time (minutes) 6 5 3.5 3 5 Influent Total 3335 8 6 65 X 105 2 78 x 105 Numbers Average Effluent 0 22 1 0 1.15x10 levels (#s/ml) Time (minutes) 150 150 150 Effluent Total 33 67 150 1.73 X 101 Numbers Percent 99 .01 99.99998 93.8 Reductions 58

PAGE 71

V1 \0 _J E en "0 Cd Q) Ill Figure 9 Fluorescent Bead Removal by Chemical Lime Treatment 100,000.-----------------------, 1,000 100 10 \ ,.,.-.,<. 0 .Q. '\.. ',"o.o-o, \ a. 1 0 1 ('I') ...l() ('I') C\J lO lO f'-. ('I') ('I') l() .ql() l() C\J l() 0 0 lO ll> ll> ll> l() 0 oo C\JW ...-...-C\J C\J ('I') l() ...-...-...-C\J Time (Minutes) *Influent 3gpm 0 Effluent 3gpm .s:;;.l nfluent 7gpm *Effluent 7gpm

PAGE 72

"' 0 _J E -en ..... en >-0 0 0 Figure 1 0 Cryptosporidium Removal by Chemical Lime Treatment 1 ,000. 100 10 1 0 1 0 .01 0.001 I I I }J I ., I 0 -o--o ...... 0 -...... o ... ., .,
PAGE 73

0\ ,_. .....J E :J u. a.. 1 .000E+09 1. 000E+08 1 0 0 0 E + 0 7 1,000,000 100,000 10,000 1 ,0001 1 oo I 101 1 F i gure 11. MS2 Remova l by Chem i cal Lime T r eatme n t I I 0 0 I ' ' o o o-o 0 1 C') -yo-10 C') (\J 10 10 ,...._ C') C') 10 .q-10 1.{) (\J 10 0 0 10 1.0 I{) I{) 10 0 00 NOO .... .., -.-.-.-(\J -.--.-(\J (\J C') 1.{) Time (Minutes) Infl uent 3gpm 0 Effluent 3gpm -Q-I n fluent 7gpm ... Eff lu en t 7gpm

PAGE 74

0\ N _J E ::J LL a.. Figure 12 PRD1 Bacteriophage Removal by Chemical Lime Treatment 1 .000E+09 1 .000E+08 1 .000E+07 1 ,000,000 100,000 1 0 ,000 1 ,000 100 10 1 <) I , I I ... 0o o o o ... Time (Minutes) *Influent 3gpm ()-Effluent 3gpm 7gpm .... Effluent 7gpm

PAGE 75

CHAPTERS DISCUSSION Five of the unit processes that removed the m i croorganisms included acti vated sludge treatment chemical lime t reatment filtration GAC and chlorina t ion Microbial removal was at least 99. 9% (3 log10) through chemical lime treatment and further reducti on occurred after chlor i nation The coliform bacteria as expected, were less resistant to the un i t processes than the pathogens (Figure 13) While the fecal coliform is the indicator used for discharge requ i rements the data clearly show that Giardia cysts were not removed to the same level as the fecal coliforms through the unit processes. Alternative indicators such as Clostridium and Enterococci demons t rated increased res i stance to the unit processes and showed sim i lar removals to the protozoa (Figures 14 and 15) Clostridium appears to be the best conservative indicator because it can still be found after reduction through the unit processes Possibly the spore-forming properties of this microorganism are similar to the oocyst or cyst stage which allows for increased resistance against phys i cal and chemical treatment processes Coliphage have been suggested as a superior indicator for human viruses particularly for d i sinfection efficacy The percent of samples positive for 63

PAGE 76

0\ Figure 13. Comparison of Fecal Coliform Removal to Pathogen Removal throughout the Treatment Plant Log1 0 Removal 6 5 4 3 2 1 0 Act. Sludge Chemical Lime Filtration Un i t Processes GAC Chlori nation &.1 Fecal Colif o rms Cryptosporidium ID Giardia

PAGE 77

(j\ V1 Figure 14. Comparison of Clostridium Removal to Pathogen Removal throughout the Treatment Plant Log1 0 Removal 5 4 3 2 1 0 Act. Sludge Chemical Lime Filtration Unit Processes GAG Chlorination Cryptosporidium OJ Giardia l3J Enterovirus

PAGE 78

0\ 0\ Figure 15. Comparison of Enterococci Removal to Pathogen Removal throughout the Treatment Plant Log1 0 Removal 6 5 4 3 2 1 0 Act. Sludge Chemical Lime Filtration Unit Processes GAC Chlorination Cryptosporidium []Giardia I8J Enterovirus

PAGE 79

Enteroviruses and coliphage concentrations throughout the unit processes were similar It is interesting to note how closely coliphage indicates the presence of viruses in wastewater (Figure 16) Coliphage concentrations were higher than enteroviruses in both unit processes that demonstrated the greatest removal of microorganisms (high pH chemical treatment and disinfection). Clostridium concentrations were also higher than the Enteroviruses (Figure 17) The ability of coliphage to indicate the presence of pathogens in the reclaimed water provides evidence of its potential for serving as an additional alternative indicator microorganism. Clostridium showed increased resistance to the unit processes as compared to both total coliforms and fecal coliforms This supports research that has demonstrated the use of coliphage and possibly Clostridium for assessing the reliability of treatment processes (Rose et al 1996, Grabowet al1978, Baker and Hegarty 1997 Armon and Kott 1996 Fujioka and Shizumura 1985, Cabelli 1977) Analysis of the entire data set by Pearson s Correlation Coefficient was performed and the results were somewhat misleading, due to the high correlations suggested (Table 15) This may be due to the fact that the concentrations of microorganisms were high at the primary and secondary sampling sites. Analysis of the data after chemical lime treatment by Pearson s Correlation Coefficient showed no correlations with any of the indicator microorganisms to Cryptosporidium The strongest correlations for Enteroviruses were shown with Enterococci Clostridium and coliphage with values of 0.5456, 0 5295 and 0 3523, respectively The strongest correlations for Giardia were 67

PAGE 80

0\ 00 120100 -Q) 80 "(j) 0 a.. 60 c Q) 0 .... Q) 40-a.. 20 o 001 12 samples Figure 16 Comparison of Enteroviruses and Coliphage in Positive Samples 011 020 017 021 023 033 Sampling Sites *Coliphage *Entero viru s 060

PAGE 81

0\ \0 120-100 Q) 80 (/) 0 a.. 60 -c Q) 0 L. Q) 40 a.. 20 o 001 12 samples Figure 17. Comparison of Enteroviruses and Clostridium in Positive Samples ** "* "* 011 0 2 0 017 021 0 2 3 033 Sampli n g S i tes >IE C l os t rid i um Enter o v i rus 060

PAGE 82

demonstrated with Clostridium and coliphage with values of 0.3819 and 0 3538 respectively (Table 16) Table 15 Correlation Matrix of Ind i cator and Pathogenic Microorganisms for Entire Treatment Process Organism Total Fecal Entero Clostri Crypto Giardia Coliphage Virus Fecal 0.9498 1 .000 0 9378 0.8359 0 1142 0.8006 0 8929 0 8762 coliforms Enterococci 0.9052 0 9378 1 .0000 0.8362 0 1366 0 8404 0 8834 0 8982 Clostridium 0.8689 0 8359 0.8362 1 0000 0.1531 0 .7455 0.8330 0 .7935 Crypto 0 1223 0 1442 0 1366 0 1531 1.0000 0.1894 0 1565 0 1721 Giardia 0 .7572 0 8006 0 8404 0 7455 0.1894 1.0000 0.7941 0.8705 Coliphage 0 8593 0.8929 0 8834 0 8330 0 1565 0 .7941 1 .0000 0 .8473 Enterovirus 0.8454 0 8762 0 8982 0.7935 0 1721 0 8705 0 8473 1 0000 Table 16 Correlation Matrix of Indicator and Pathogenic Microorganisms after Lime Treatment Organism Total Fecal Entero Clostri Crypto Giardia Coliphage Virus Fecal 0 2448 1.000 0 6915 0.4324 0.1603 0 5084 0 5269 coliforms 0.0606 Enterococci 0.3063 0 .6915 1 0000 0 5461 0.0204 0.2474 0 3948 0.5456 Clostridium 0 4786 0 4324 0.5461 1.0000 0.0438 0.3819 0 4473 0 .5295 Crypto 0.0426 -0 0606 0 0204 0 0438 1.0000 0.2177 0 0830 0.0202 Giardia 0 2446 0.1603 0 2474 0 3819 0 2177 1.0000 0 3538 0 .3799 Coliphage 0.2970 0.5084 0 3948 0 4473 0 0830 0.3538 1 0000 0.3523 Enterovirus 0.2679 0.5269 0 5456 0.5295 0 .0202 0.3799 0.3523 1 0000 Pilot scale studies of the chemical lime treatment process demonstrated removals of Cryptosporidium oocysts by 99% which was also observed during the full-scale process For the coliphage the indigenous monitoring showed a 99% reduction and represents the assay of a heterogenous population of 70

PAGE 83

coliphage some of which may be very sens it ive to the high pH (such as MS 2) and some of which may be more resistant to the high pH (such as PRD1 ). More information should be gathered regarding the resistance of PRD1, which may be a more valid surrogate for the high pH chemical treatment for the human enteroviruses which are known to be more resistant to h i gh pH than the coliphage Table 17 compares the removals of the pilot studies and the removals estimated by the i ndigenous monitoring program Table 17 Removal and Inactivation of Cryptosporidium Beads and Phage by Chemical Treatment by P il ot Studies Compared to Monitoring Data Condition Oocyst Bead MS2 PRD1 Indigenous Evaluated Removal Removal Removal/ Removal/ Coliphage (%) Inactivation Inactivation Removal/ Inactivation Pilot Scale 98.99 99.53 99. 9999 93. 8 NA 3gpm 99. 9999+ 99 999+ Pilot Scale 99.65 98. 02 99. 9999 95.4 NA 7gpm 99 99999+ 99 999+ Full-Scale 99.8 NA NA NA 99.5 Monitoring +Experiment 1, perhaps problems with the neutralization of the sample Removal efficiencies and log reductions were greatest through secondary treatment and high pH chemical treatment. This was similar to results reported by Grabow and Isaacson (1978) that excellent reductions were observed for microorganisms at an operational pH of 11. 2 After secondary and chem i cal treatment system detection of indicator bacteria coliphage enteroviruses and 71

PAGE 84

protozoa were sporadic and the levels were near the limits of detection of the various methods The results in this study were similar to those for enterovirus removal through the reclamation plant as described by Yanko (1993) and Rose et al (1996). Analysis of 10 years of enteric virus monitoring from 6 tertiary treatment water reclamation plants i n California found only 1 sample out of 590 samples was positive for enteric viruses in the final effluent (Yanko 1993). Removals of viruses through secondary treatment was at least 99. 8%. The primary barrier in these plants was disinfection In the California studies chlorine contact times averaged approximately 90 minutes with final total residuals of 4-5 mg/L. The virus monitoring data from this 1 0 year period confirmed the results of the Pomona Virus Study and suggest that the viral risks associated with the use of reclaimed water are within acceptable levels These studies also found that male specific coliphage were more resistant to standard disinfection practices A study conducted at a water reclamation plant by Rose et al (1996) in St. Petersburg, Florida evaluated filtration and disinfection at 4 mg/L for a minimum of 30 minutes. This study demonstrated a 5 log10 removal for enteroviruses but 8% of the samples were positive for enteroviruses in the storage tank at low levels (approximately 1/500 L) Asano and Mujeriego state that for efficient virus removal and inactivation two major operating criteria must be met: (1) the effluent must be low in suspended solids and turbidity prior to disinfection to reduce shielding of viruses and chlorine demand and (2) sufficient disinfectant dose and contact time provided for the wastewater (Asano and Mujeriego 1988) 72

PAGE 85

Giardia cysts and Cryptosporidium oocysts were reduced by 4.1 and 3 3 log10, respectively but 25% and 16% of the samples were positive for Giardia cysts and Cryptosporidium oocysts in the storage tanks (Rose et al1996). UOSA has two major barriers for viruses : chemical lime treatment and disinfection. This is different than other studies and probably is superior since no human viruses were detected in the final effluent. The true removal (only >) cannot be calculated Enteroviruses were not detected in any of the samples from the post dechlorination effluent at an average total residual chlorine of 1 5 mg/L or in the final effluent reservoir but levels of bacteria and coliphage were higher unless the residual was kept above 1 9 mg/L. Total coliforms enterococci, Clostridium, Cryptosporidium and Giardia were all detected in the final effluent. UOSA treatment processes achieved approximately 4 log10 reduction with 17% and 8% of the samples posit i ve for Giardia cysts and Cryptosporidium oocysts respectively, in the final effluent postchlorination. Levels of the microorganisms increased slightly (with the exception of enteroviruses and Giardia) after passage of the effluent through the open ballast pond prior to multi-media filtration which was not suprising with the wild geese population at the plant. The percentage of samples positive and the concentrations were compared for the treated reclaimed water and the final effluent reservoir water. In every case the treated final effluent was of better quality than the water in the UOSA effluent reservoir (Table 18) The concentration and percent of samples positive increased for the indicator microorganisms and the pathogens in the final 73

PAGE 86

effluent reservoir The naturally occurr ing indicator bacteria and protozoa detected in the reservoir were most likely contr i buted from animals No human viruses were detected in the effluent reservoir during th i s study 74

PAGE 87

-....j V1 Table 18 Comparison of the UOSA Final Effluent to the Final Effluent Reservoir Water Quality Microorganism Percentage of Average I Samples Concentration for Positive Positive Samples Effluent Final Effluent Effluent Final Effluent Reservoir Reservoir Clostridium1 10 78 3 5 4.9 Total coliforms 1 36 100 1 6 180.3 Fecal coliforms 1 0 100 < 0.5 22 9 Enterococci1 17 73 2.7 10.1 Coliphage;l 0 45 < 12.5 20.6 Enterovirus;, 0 0 < 0.085 < 0 .28 Cryptosporidium4 8.3 9.1 0 .44 5.7 Giardia5 17 18.2 6 6 42.9 CFU/1 00 ml, ;lPFU/mL, "PFU/1 00 L, qOocysts/1 00 L, Cysts/1 00 L

PAGE 88

CHAPTER 6 CONCLUSION This is the first major study to examine comprehensively bacteria protozoa, alternative indicators and viruses through a full-scale advanced wastewater treatment facility. One year of monitoring for a total of 96 samples supported the following conclusions: Chemical lime treatment with second stage recarbonation is the most efficient barrier to the passage of microorganisms. Pilot scale studies of the chemical lime treatment process demonstrated removals of Cryptosporidium oocysts by 99% which was also observed during the full-scale process The removal mechanism for protozoa after the chemical treatment system appears to be physical removal since other experiments conducted but not presented in this research showed no decrease in viability of oocysts after exposure to high pH or high pH and disinfection combination Of all the indicators, Clostridium and coliphage best reflected the removal of enteroviruses for the chemical treatment system and the disinfection process The percentage of samples positive and the concentrations for the bacteria coliphage and protozoa were compared for the treated reclaimed water and the final effluent reservoir water receiving the effluent. In every case the 76

PAGE 89

treated water was of better quality that the ambient water in the UOSA final effluent reservoir. This study has developed an extens i ve database for the concentrations of bacteria protozoa and viruses present in the raw wastewater and the removals of these microorganisms through the various unit processes in an advanced water reclamation fac i lity The data that have been collected on microbial removals from this study provides i nformation necessary for determining the impact of water quality and potential health risks for surface water augmentation or groundwater recharge of drinking water supplies through planned indirect potable reuse This study also provides in i t i a l data for the use of alternative indicator microorganisms in addition to the conventional fecal coliform to indicate the presence of pathogens i n water. Thi s may have greater reliabil i ty in the future for ensuring the protection of drinking water supplies 77

PAGE 90

REFERENCES 1. Adams, M.H. 1959 Bacteriophages lnterscience Publishers New York 1959 2. Armon and Kott. 1996. Bacteriophages as Indicators of Pollution Critical Reviews i n Environmental Science and Technology 26(4) : 299-335. 3. Armon, R., and Payment, P. 1988 A modified m-CP medium for enumerating Clostridium perfringens from water samples Can J Microbial. 34:78-79 4 Asano, T 1995 Drinking Repurified Wastewater. Journal of Environmental Engineering 121: 548 5 Asano, T. and Mujeriego, R. 1988 Pretreatment for Wastewater Reclamation and Reuse In Pretreatment in Chemical Water and Wastewater Treatment (ed H H Hahn and R Klute) Springer-Verlag Berlin Heidelberg : 347356 6. Asano, T., Leong, L Y.C. Rigby, M.G., and Sakaji, R.H. 1992 Evaluation of the California Wastewater Reclamation Criter i a using Enteric Virus Monitoring Data Wat. Sci. Tech 26 : 1513 1524 7 Asano, T. and Levine A.D 1995 Wastewater reuse : a valuable link in water resources management. Water Quality International 4 : 20-24 8. Asano, T. and Levine, A.D. 1996. Waste-water Reclamation Recycling and ReusePast Present and Future. Water Science and Technology 33 : 1-14 9 Baker, Katherine H. and Hegarty, J.P. 1997 Detection and occurrence of i ndicator organisms and pathogens Water Environment Research 69 : 403-415. 10. Bitton, G. 1994 Wastewater Microbiology Wiley-Liss New York, 77 100. 11. Boring, J.R. Ill, Martin, W.T and Elliot, L.M. 1971 Isolat i on of Salmonella typhimurium from municipal water Riverside Calif 1965 American Journal of Epidemiology 93 : 49-54 78

PAGE 91

12 Cabelli, V.J. 1977. Clostridium perfringens as a water quality indicator In Bacteria/Indicators/Health Hazards Associated With Water ASTM STP 635 A.W. Hoadley and B J Dutka (Ed ) American Society for Testing and Materials 1977 : 247-264 13. Clausen, E.M., Green, B.L., and Litsky, W. 1977 Fecal streptococci : indicator of pollution In Bacteria/Indicators/Health Hazards Associated With Water ASTM STP 635 A.W Hoadley and B J Dutka (Ed.) American Society for Testing and Materials, 1977:247-264 14. Cohen, J., and Shuval, H.l. 1973 Water Air and Soil Pollution 2:85-95. 15 Condie, L.W., Lauer, W.C., Wolfe, G.W., Czeh, E.T., and Bums, J.M. 1994 Denver Potable Water Reuse Demonstration Project. Food and Chemical Toxicology 32 : 1021-1030 16 Craun, G. F. 1988. Surface water supplies and health. Journal American Water Works Association 80:40-52 17 Crook, J. 1994 Assessment of Water Reclamation and Reuse Research Needs Proc 1994 AWWAIWEF Water Reuse Symp Dallas Texas :371. 18 Crook, James, Asano, Takashi, and Nellor, Margaret. 1990 Groundwater Recharge with Reclaimed Water in California. Water Environment and Technology 2:42-49. 19. Crook, James and Surampalli, Rao Y. 1996 Water Reclamation and Reuse Criteria in the U S Wat. Sci. Tech 33:451-462. 20 d' Angelo, Salvatore 1996 A\!INVA and WEF Prepares Guidelines For Using Reclaimed Water to Augment Potable Water Resources. Proc 1996 AWWAIWEF Water Reuse Symp., San Diego, California : 55-58 21. Danielson, R.E., Pettegrew, L.A., Soller, J.A., Olivieri, A W., Eisenberg, D.M., and Cooper, RC. 1996 A Microbiological Comparison of a Drinking Water Supply and Recla i med Wastewater for Direct Potable Reuse Proc. 1996 AWWAIWEF Water Reuse Symp San Diego Califomia : 727-734. 22 Davies-Colley, R.J., Bell, R.J., and Donnison, A.M. 1994. Sunlight inactivation of enterococci and fecal coliforms in sewage effluent diluted in seawater Applied Environmental Microbiology 60:2049-2058 23 de Peyster, Ann, Froines, John R., Olivieri, Adam W., and Eisenberg, Don M. 1993 Aquatic Biomonitoring of Recla i med Water for Potable Use : The San Diego Health Effects Study Journal of Toxicolc>gy and Environmental Health 39:121-141 79

PAGE 92

24. Dutka, B.J. 1973. Col i forms are an inadequate index of water quality Journal of Environmental Health 36 : 39 25. Florida Department of Environmental Regulation. 1990. Reuse of Reclaimed Water and Land Application Chapter 17-610. Florida Administrative Code, Florida Department of Environmental Regulation, Tallahassee Florida 26. Fujioka, R.S. and Shizumura, L.K. 1985 Clostridium perfringens a reliable indicator of stream water quality Journal Water Pollution Control Federation 57 : 986-992. 27. Funderberg, S.W. and Sorber, C.A. 1985 Coliphages as indicators of enteric viruses in activated sludge Water Research 19:547. 28. Gagliardo, P.F., Findley, P., Richardson, T.G., and Weinberg, K. 1996 Optimization of Reclamation and Repurification at San Diego North City In Proc AWWAM/EF Water Reuse Conference San Diego California 29. Gerba, C.P., Goyal, S M., LaBelle, R.L. et al. 1979 Fa i lure of indicator bacteria to reflect the occurrence of Enteroviruses inmarine waters American Journal of Public Health 69 : 1116-1119 30. Gerba, C.P. and Rose, J.B. 1990 Viruses in source and dri nking water In Drinking Water Microbiology, (ed G A McFeters) Springer-Verlag, New York: 380-396. 31. Grabow, W O.K. 1990 Microbiology of Drinking Water Treatment: Reclaimed Wastewater In Drinking Water Microbiology (ed G A McFeters), Springer-Verlag, NewYork: 185-203 32 Grabow, W.O.K., and Isaacson, Margaretha. 1978. Microbiological Quality and Epidemiological Aspects of Reclaimed Water. Prog. Wat. Tech 10: 329-335 33. Grabow, W.O.K., Middendorff, lrmela G and Sasson, Nerine C. 1978 Role of Lime Treatment in the Removal of Bacteria Enteric Viruses and Coliphages in a Wastewater Reclamation Plant. Applied and Environmental Microbiology 35 : 663-669. 34. Grabow, W.O.K., Burger, J.S., and Nupen, E.M. 1980. Evaluation of Acid Fast Bacteria, Candida albicans Enteric Viruses and Conventiona l Indicators for Monitoring Wastewater Reclamation Systems Prog Wat. Tech. 12: 803-817 35. Grabow W.O.K., Bateman, B.W., and Burger, J.S. 1978. Microbiological Quality Indicators for Routine Monitoring of Wastewater Reclamation Systems. Prog Wat. Tech. 10:317-327. 80

PAGE 93

36 Hattingh, W.H.J., and Bourne, D.E. 1989 Research on the health implications of the use of recycled water in South Africa SAMJ 76 : 7-10. 37. Havelaar, A.H., Olphen, M.V., and Drost, Y.C. 1993 F-specific RNA bacteriophages are adequate model organisms for enteric viruses in fresh water Applied Environmental Microbiology 59 : 2956 2962 38 Hemmer, J. et al. 1994. Tampa Water Resource Recovery Project. Proc. 1994 AWWAIWEF Water Reuse Symp Dallas Texas : 557. 39. Hibler, C. and Hancock, C . Waterborne Giardiasis 40. Hrudey, Steve E., Hrudey, Elizabeth J., and Shaw, Nola J. 1991 Health Effects associated with waste treatment, disposal and reuse. Research Journal WPCF 63: 437-444 41. Lauer, William C. 1991. Water 9 Quality for Potable Reuse. Water Science and Technology 23 : 2171-2180 42. Lauer, W., and Rogers, S.E. 1996 The Demonstration of Direct Potable Water Reuse : Denver's Pioneer P r oject. Proc AWWANVEF Water Reuse Conference San Diego Ca l ifornia. 43 LeChevallier, M.W., Norton, W.D., and Lee, R.G 1991 Giardia and Cryptosporidium spp in surface water supplies Applied Environmental Microbiology 57 :261 0-2616 44. LeChevallier, M.W., Norton, W.O. and Lee, R.G 1991. Giardia and Cryptosporidium spp in Filtered Drinking Water Supplies Applied Environmental Microbiology 57 : 2617-2621 45. Lisle, J.T. and Rose, J B 1995 Cryptosporidium contamination of water in the USA and UK: a mini-review. J. Water SRTAqua 44 : 103-117. 46. MacKenzie, William R., Hoxie, Neil J Proctor, Mary E., Gradus, M. Stephen, Blair, Kathleen A., Peterson, Dan E., Kazmierczak, James J Addiss, David G., Fox, Kim R., Rose, Joan B., and Davis, Jeffrey P 1994 A massive outbreak in Milwaukee of Cryptosporidium infection transmitted through the public water supply The New England Journal of Medicine 331(3) : 161-167 47 Madore, M.S., Rose, J.B., Gerba, C.P., Arrowood, M J and Sterling, C.R 1987 Occurrence of Cryptosporidium oocysts in sewage effluents and selected surface waters Journal of Paras i tology 73: 702-705. 81

PAGE 94

48. Marshall, Marilyn M., Naumovitz Donna, Ortega, Ynes, and Sterling, Charles R. 1997 Waterborne Protozoan Pathogens Clin Microbial. Rev. 10: 68-85 49. McEwen, Brock and Richardson, Tom. 1996. Indirect Potable Reuse : Committee Report Proc. 1996 AWWAM/EF Water Reuse Symp., San Diego California : 485-503. 50. Metcalf, T.G. 1978. Indicators for viruses in natural waters In Water Pollution Microbiology 2 (ed. R. Mitchell), Wiley-lnterscience New York : 301-325 51. Neller, Margaret H., Baird, Rodger B., and Smyth, John R. 1985. Health Effects of Indirect Potable Water Reuse Journal AWWA : 88-96 52 Olivieri, A.W., Eisenberg, D.M., Cooper, R.C., Tchobanoglous, G., Gagliardo, P. 1996 Recycled Water a Source of Potable Water : City of San Diego Health Effects Study Water Science and Technology 33 : 285-296 53. Payment, P. and Franco, E. 1993. Clostridium perfringens and somatic coli phages as indicators of the efficiency of dri nking water treatment for viruses and protozoan cysts. Applied Environmental Microbiology 59:2418-2424. 54. Payment, Pierre, and Armon, Robert. 1989. Virus Removal by Drinking Water Treatment Processes Critical Reviews in Environmental Contro l19:15-31. 55 Pia, Michelle M., Grebbien, Virginia, and Gaston, John M. 1996 Potable Reuse and the Emerging Conflicts with Drinking Water Regulations 1996 AWWANVEF Water Reuse Symp., San Diego Califomia : 715-721. 56. Robbins, M.H.1993 Supplementing a surface water supply with recla i med water. In Proceedings of the AVVWA Annual Conference and Exposition June 610, San Antonio TX, AWWA, Denver CO. 57. Rodgers, Mark R., Flanigan, Debbie J. and Jakubowski, Walter. 1995 Identification of Algae Which Interfere with the Detection of Giardia Cysts and Cryptosporidium Oocysts and a Method for Alleviating This Interference Applied and Environmental Microbiology 61: 3759-3763 58 Rose, J.B. 1988 Occurrence and Significance of Cryptosporidium in water Journal Amer i can Water Works Association 80 : 53-58. 59. Rose Joan B., Landeen, LeeK., Riley Kelley R., and Gerba, Charles P. 1989 Evaluation of Immunofluorescence Techniques for Detection of Cryptosporidium Oocysts and Giardia Cysts from Environmental Samples Applied and Environmental Microbiology 55:3189-3196. 82

PAGE 95

60 Rose, J.B., Gerba, C.P., and Jakubowski, Walter. 1991. Survey of Potable Water Supplies for Cryptosporidium and Giardia. Environmental Science and Technology 25 : 1393-1400 61. Rose, Joan B., Robbins, Millard, Friedman, Debra, Riley, Kelley, Farrah, Samuel R., and Hamann, Carl L. 1996 Evaluation of Microbiological Ban iers at the Upper Occoquan Sewage Authority 1996 AWWA/WEF Water Reuse Symp., San Diego, California : 291-305. 62. Rose, Joan B., Dickson, Linda J Farrah, Samuel R., and Carnahan, Robert P. 1996. Removal of Pathogenic and Indicator Microorganisms by a Full Scale Water Reclamation Facility. Water Research 30 : 2785-2797 63. Seligmann, R. and Reitler, R. 1965 Enteropathogens in water with low Escherichia coli titer. Applied Environmental Microbiology 57 : 1572 157 4. 64. Sinton, L.W., Davies-Colley, R.J., and Bell, R.G. 1994 Inactivation of enterococci and fecal coliforms from sewage and meatworks effluents in seawater chambers. Applied Environmental Microbiology 60:2040-2048 65. Smith, Robert G. 1995. Water reclamation and reuse Water Environment Research 67:488-495 66. Snowdon, Jill A. and Cliver, Dean 0. 1989 Coliphages as Indicators of Human Enteric Viruses in Groundwater. Critical Reviews in Environmental Control 19: 231-249 67. Standard Methods for the Examination of Water and Wastewater. 1992 APHA AWWA, WEF, 18th Edition, Washington, D C 68. State of Arizona. 1991. Regulations for the Reuse of Wastewater. Arizona Administrative Code Chapter 9, Article 7, Arizona Department of Environmental Quality, Phoenix, Arizona. 69. State of California. 1978. Wastewater Reclamation Criteria. California Administrative Code, Title 22, Division 4 California Department of Health Services, Sanitary Engineering Section Berkeley, Ca l ifornia 70. Stander, G.J., and Clayton, A.J. 1977 Planning and construction of wastewater reclamation schemes as an integral part of water supply. In Water; Wastes and Health in Hot Climates, (ed. R Feachem M. McGarry and D Mara) Wiley, London : 383-391. 71. U.S. EPA 1992. Guidelines for Water Reuse U S Environmental Protection Agency, Center for Environmental Research Information, Cincinnati Ohio. 83

PAGE 96

72. USEPA, Monitoring Requirements for Public Drinking Water Supplies ; Proposed Rule, Federal Register, 59(28) : 6332-6429 1994 56. USEPA, Monitoring Requirements for Public Drinking Water Supplies ; Proposed Rule Federal Register 59(28) : 6430-6440 1994 73. van Leeuwen, J. (Hans). 1996 Recla i med water-an untapped resource Desalination 106:233-240 7 4 Van Riper, C. and Geselbracht, J. 1996 Water reclamation and reuse Water Environment Research 68 : 516-520 75. Yanko, William A. 1993 Analysis of 10 years of virus monitoring data from Los Angeles County treatment plants meeting California wastewater reclamation criteria. Water Environ. Res 65 : 221-226 84

PAGE 97

APPENDICES 85

PAGE 98

APPENDIX A 86

PAGE 99

Table Al 4/28/95 SAMPLE ID SITE DESCRIPTION 001 I UNTREATED WASTEWATER I 011 SECONDARY EFFLUENT 020 S ECOND STAGE RECARB. 017 MULTIMEDIA. INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT oJj FINAL EFF POSTDECIIL. 060 FINAL EFFL. RESERVOIR NA -Sample was not analyzed. co --...! ENTEROCOCCI 6. 5 X 10' 1 0 X 10 3 0 < 10.0 < 10 0 1.0 < 0 5 < 1.0 TOTAL/FECAL PHAGE COLIFORMS 11/100 ll/ 100mL m t. NA 6.5 X 10' NA. 200 NA. < 12.5 NA. < 12 5 NA. < 12 5 NA. < 12 5 NA < 12 5 NA < 14 ) ENTEROVIRUS CLOSTRIDIUM CRYPTOSPORIDIUM GIARDIA MPN/100 L ll/ 100mL II/100L II/100L 690 0 NA < 1123 22 4 6 14 0 NA. < 33.9 67 9 < 0 .37 NA. 2 1 4 22 < 0 .10 NA < 0 .74 < 0.74 < 0 .069 NA < 0.42 < 0 42 < 0.077 NA 2.82 < 0 94 < 0 061 NA 0 44 0 44 < 0 .14 NA < 1.95 1. 9 5

PAGE 100

Table A2 5/24/95 SAMPLE 10 SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF POSTDECHL. 060 FINAL EFFL. RESERVOIR Plate too numerous to count (a)Problem with analysis 00 00 ENTEROCOCCI ll/ 100mL 5 0 X 104 400 1.5 5.5 0.5 < 0.5 < 0 5 < 0 5 TOTAL/FECAL PHAGE COLI FORMS lll100mL 111100 mL I 3. 2 X 105 1 8 X 10' I 1 2 X 101 l 5 X 101 15 0 I 3 0 < 12 5 92 0 I 57.0 25 0 52 0 I 11.0 < 1 2 5 2.0 I (a) < 12 5 4 .01<1.0 < 12.5 < 1 0 I 19.0 < 12.5 ENTEROVIRUS CLOSTRIDIUM MPN/100 L lll 100mL 2 4 X 10' 3 1 < 0 12 2 0 < 0 067 2 5 < 0 045 0 5 < 0 11 0.5 < 0 11 < 0.5 < 0 21 < 0 5 CRYPTOSPORIDIUM III100L 277.8 < 1 9 8 < 3 9 6.4 0.9 < 0 9 < 2 1 < 3.7 GIARDIA II/100L 22500 59 5 < 3 9 < 6 4 < 0.9 < 0 9 < 2 1 7 5 'd (1) :::1 p.. t-' X ,...... (") 0 :::1 rt t-' :::1 (1) p.. ..._,

PAGE 101

Table A3 6/27/95 SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 C? \.() MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF POSTDECHL. 060 FINAL EFFL RESERVOIR Problems with analysis. ENTEROCOCCI 11/lOOmL 7 1 X 10' 4 5 X 101 88 5 15. 5 3 0 < 0.5 16.0 TOTAL/FECAL PHAGE COLIFORMS 11/100 l#/100mL mL 2. 4 X 10' I 8.6 X 105 2. 5 X 101 1.5 X 10'/ 1.6 X 101 3 8 X 101 7 o I lal < 12.5 114.0 I 118.0 < 12.5 41. o I 29 o < 12 5 37.o I 9.o < 12.5 1.0/cl.O < 12.5 I < 12.5 -ENTEROVIRUS CLOSTRIDIUM MPN/100L 11/lOOmL 1 0 X 101 3 0 X 104 1.2 X 101 4 5 X 101 < 0.012 < 0 5 < 0.035 4 0 0 .037 < 0 5 0.120 < 0 5 < 0.069 < 0.5 < 0.14 < o s CRYPTOSPORIOIUM II/100L < 1. 15 X 101 < 24.5 < 2.6 < 5 0 < 10.3 < 4 7 < 4.6 5.7 GIARDIA II/100L 1.22 X 10' 24.5 2 6 < 5.0 < 10.3 < 4 7 < 4 6 < 5.7 > 't:l 't:l (I) :::1 r:L ..... :< > ....... () 0 :::1 rt ..... g (I) 0.. -......;

PAGE 102

Table A4 7/24/95 I SAMPLE ID I SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECHL. 060 FINAL EFFL. RESERVOIR ENTEROCOCCI ll/100mL 7 5 X 10' 600. 0 6.0 92 0 6 0 0 5 4 5 0 5 o17 Echovirus 11 ; 023 -Echovirus 11 b Too numerous to count \.0 0 TOTAL/FECAL PHAGE COLIFORMS 11/100 ll/ 100mL mL 2. 2 X 10' I 5.5 X 101 3 6 X 10 2 4 X 10'/ 5.5 X 10' l 2 X 1 01 6.0 I 2 0 < 12 5 b I 202 0 < 16 7 17 0 I 8 o < 12 5 8 0 I 8 0 < 25 0 <1.0 I <1.0 < 12.5 b I 3 o < 12. 5 ENTEROVIRUS CLOSTRIDIUM MPN/100 L ll/100mL 260 0 4 5 x 1 o 4.32 1.4 X 10' < 0.097 1.0 07 2.5 0.043 2.5 0 068. 6 0 < 0.047 3 5 < 0.23 10.0 CRYPTOSPORIDIUM II/100L < 603.8 < 12 4 < 5.1 < 5 0 < 4 9 < 4 6 < 1.2 < 11 3 GIARDI A II/ 1 00 L 5 1 X 10' 37.2 5 1 < 5. 0 < 4.9 < 4 6 < 1.2 < 11.3 '"0 (I) l:l 0.. > ,....._ (') 0 l:l rt ..... g (I) 0.. .......-

PAGE 103

Table AS 8/30/95 SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECHL. 060 FINAL EFFL RESERVOIR Too numerous to count "Not determined <.0 ...... ENTEROCOCCI lll100mL 8 2 X 10' 2.7 X 10' 11.5 130.0 15.5 5.0 1.0 4.0 TOTAL/FECAL PHAGE CO!, I F ORMS llllOOOIL 111100 mL 4 5 X 10' I 6 3 X 10' 4 1 X 104 1 5 X 10'1 2 8 X 10' 1 0 X 1 04 10. 5 I 3.5 < 12.5 44o.o I 275.0 12.5 6o. o I 3o.o 12.5 55. 0 I 17. 5 < 12. 5 <0.5 I < 0 5 < 12. 5 I 4 5 < 12.5 ENTEROVIRUS C LOSTRIDIUM MPNI100 L llllOOmL 1. 3 X 101 b 47. 0 < 0.103 < 0 .081 b < 0 .043 < 0 .056 < 0 .055 < 0.39 CRYPTOSPORIDIUM III100L <2959 < 375.8 < 11. 0 < 4.6 < 1.0 < 4.6 < 2 0 < 42.4 GIARDIA III100L 44386 375. 8 < 11. 0 < 4 6 < 1 0 < 4 6 < 2.0 <42. 4 .g-'d ro ::1 p.. ,..-... (") 0 :::J rt t::l s:: ro p.. ..._,

PAGE 104

Table A6 9/26/95 SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND SThGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECHL. 060 FINAL EFFL. RESERVOIR 0 20 Coxsackie 85 <..0 N ENTEROCOCCI ll/100mL 8 7 X lO s 850. 0 4 5 5 370. 0 17 0 16 5 < 0.5 6.5 TOTAL/FECAL PHAGE COLIFORMS 11/100 11/lOOmL mL 3 3 X 101 / 5 9 X lOs 2 9 X 10' 3 7 X 10'/ 2 8 X 10' 7 6 X 10' 160. 0 I 10.0 12 5 825 0 I 480.0 75 0 10. 0 I 27 5 < 12 5 46 5 I 11 0 < 12 5 <0 5 I <0.5 < 12.5 245. 0 I 16.5 12 5 ENTEROVIRUS CLOSTRIDIUM MPN/100L 11/lOOmL 1 2 X 10' 5, 7 X 104 57. 0 1 1 X 10' 0 22 7.0 0.18 5 0 < 0 12 0 5 < 0 09 < 0 5 < 0 09 < 0.5 < 0.23 2.5 CRYPTOSPORIDIUM II/100L 2690 < 79 1 4.8 < 12.5 < 1.0 < 0.9 < 4 9 < 10 6 GIARDIA II/100L 3 9 X 104 79.1 4 8 < 12.5 < l. 0 < 0 9 < 4.9 < 10.6 'd ::1 0. X > ......... (') 0 ::s rt g 0. ........

PAGE 105

\.() w Table A7 10/24/95 SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER I 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFI.UENT 033 FINAL EFF. POSTDECHL. I 060 FINAL EFFL. RESERVOIR ENTEROCOCCI ll/100mL 8 7 X lOs 1 4 X 101 17 5 23.5 1.0 3 0 < 0.5 61.0 TOTAL/FECAL PHAGE COLIFORMS 11/100 ll/100m L mL 2. 8 X 10' I 5.5 X lOs 5 7 X 10' 2 4 X lOS I 1.3 X 101 4 1 X 10' 155 I 57. 5 < 12 5 335 I 49 5 12 5 20.0 I 9.5 < 12 5 17 5 I 4 0 < 12 5 o 5 I < o 5 < 12 5 6oo. o I 55.0 J7. 5 ENTEROVIRUS CLOSTRIDIUM MPN/100 L 11/lOOmL 150 0 1 6 X 10' 1.4 833 0 < 0 14 0 5 < 0 .086 < 0.5 < 0 .051 < 0 5 < 0.066 < 0 5 < 0.061 < 0.5 < 0.15 0 5 CRYPTOSPORIDIUM II/100L < 1. 4 X 101 < 66.2 < 1. 1 < 20 2 < 1. 1 < 0.9 < 2 3 < 22 5 GIARDIA II/100L 9 95 X 10' < 66.2 < 1 1 < 20.2 < 1 1 < 0 9 < 2 3 < 22.5 .6" 'd (IJ ::l 0.. ...... X > -(') 0 ::l rt ...... ::;1 (:::: 11> 0..

PAGE 106

Table AS 11/28/95 1..0 .:-' SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECHL. 060 FINAL EFFL. RESERVOIR ENTEROCOCCI ll/100mL 2 6 X 10' 233.j 4 0 14.5 < 0 5 < 0.5 < 0 5 < 0.5 TOTAL/FECAL PHAGE COLIFORMS 11/100 ll/100mL mL 3. 5 X 10' I 1 4 X 1il5 3 5 X 10' 6 3 x 1 o 1 2. 0 X 101 1.6 X 101 58. 5 I 4 0 75. 0 3oo.o I 52.5 175. 0 43. 0 I 10.5 75.0 25. 0 I 1 0 25. 0 1.0 I < o 5 < 12. 5 4o.o I 11.0 50. 0 ENTEROVIRUS CLOSTRIDIUM MPN/100 L ll/100mL 320.0 3. 2 X 10' 2.5 311.0 < 0.14 3 5 < 0 .069 11. 0 < 0.063 < 0.5 < 0 .062 < 0.5 < 0.049 < 0.5 < 0 .24 4 5 CRYPTOSPORIDIUM II/100L < 4 5 X 101 < 63.3 < 10.9 < 9 6 < 10.1 < 27. 5 < 5.7 < 120. 0 GIARDIA II/100L 6. 3 X 10' < 63.3 < 10.9 < 9.6 < 10.1 < 27. 5 < 5.7 < 120. 0 .fl '0 ro ::l p.. 1-' !I> .--.. n 0 ::l rt 1-' ::l ro p.. '-'

PAGE 107

Table A9 1/23/96 SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF POSTDECHL. OVERFLOW 'MPN method u sed t..<.) V1 ENTEROCOCCI ll/100mL 3.1 X 10' 2. 7 X 101 80.0 325 0 45. 5 59. 0 < 0 5 350.0 TOTAL/FECAL PHAGE COLIFORMS 11/100 ll/100mL mL' > 1 6 X 10' I 6. 4 X 10' 9.0 X 10' 9 0 X 10' I 2 1 X 101 5 0 X 101 BOO I 130 12 5 > 1 6 X 10' I 790.0 5. 0 X 10' >1.6xlO'I 12 5 240 > 1 6 X 10' I < 12. 5 130 <1.11<1.1 < 12.5 eo.o I 9.0 25 0 ENTEROVIRUS CLOSTRIDIUM MPN/100 L ll/100mL 187.0 1. 5 X 10' 1.5 888. 9 < 0.39 9 0 0.66 95.0 0 .27 6 0 0 .18 2.5 < 0.13 < 0 5 0 39 27.0 CRYPTOSPORIDIUM II/100L <1.1x101 < 354 7 < 31.6 < 10.9 < 5 3 < 25.5 < 29.9 < 10. 4 GIARDIA II/100L 6. 5 X 10' < 354. 7 727. 8 326.9 < 5 3 < 25 5 < 28.9 239.2 "d 10 1:1 p.. r' :>< > r-.. (") 0 ::I rt r' g 10 p.. '-'

PAGE 108

Table AlO 2/6/96 I \\) 0\ SAMPLE ID SIT E DESCRIPTION 001 UNTRE A T E D WAS T EWATER 011 SECONDARY EFFLUENT 0 20 SECOND STAGE RECARB. 0 1 7 MULT I M E DIA INFLUENT 0 2 1 MULTI-MEDIA EFFLUENT 02 3 GAC E FFLU ENT 033 FINAL E FF. P O S TOECHL. 060 F INAL EFFL RESERVOIR ENTEROCOCCI ll/100mL 1 6 X 1 05 288. 9 < 0 5 3 0 < 0.5 0 5 < 0 5 1 5 TOTAL/FECAL PHAGE COL I FORM S 11/ 100 #/100mL m L 3.2x10'/ 6.4 X 10' 1 3 X 10 5 8 X 10'1 4 00 1.3 X 10' o 5 I < 0 5 < 12.5 9 6 5 I 10 0 12 5 38.0 I 2 5 < 25.0 20.5 I 2 0 < 25 0 < o 5 I< o 5 < 1 2 5 15 o o I n.o 2 5 0 --E NTEROV IRUS CLOSTRIDIUM MPN/1 00 L ll/100mL 4 6 X 1 03 2 1 X 10' 6 9 338 9 < 0 15 1.5 < 0 14 5 0 < 0 1 5 0 5 < 0.12 < 0 5 < 0.13 < 0 5 < 0 9 1 9 0 -------C R Y PTOSPORIOIUM GIARDI A #/100L #/100L < 2 4 X 103 9 6 X 10' < 155. 3 < 15 5 3 < 25.1 275 7 56 3 112 7 <' 1 0 0 < 10. 0 < 29.6 < 2 9 6 < 25 1 < 2 5 1 < 9 5 5 < 9 5 5 .6" "d (1) :::1 A. > ........ (") 0 ::s rt .... ::s c (1) p.. ._,

PAGE 109

Table All 3/5/96 I I \...0 -.....! SAMPLE ID SITE DESCRIP TION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB 01 7 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECIIL. 060 FINAL EFFL RESERVOIR ENTEROCOCCI ll/100mL 5.9 X 10' l.1X 10' 54 5 62.0 26 0 19.0 < 0 5 14 0 TOTAL/FECAL PHAGE COLIFORMS 11/100 11/lOOmL mL 4.2x10'/ 7.2 X 104 1.2 X 1 01 6 1 X 10'/ 1. 3 X 10' 9.1 X 10' 11 5 I o.5 12.5 20.0 I 1.0 37. 5 11 0 I 2.0 < 12 5 6.0 I o.5 < 12.5 < o 5 I < o s < 1 2 5 11.0 I 6 5 25.0 ENTEROVIRUS CLOSTRIDIUM CRYPTOSPORIDIUM MPN/100 L ll/100mL II/100L 490.0 2.6 X 104 < 9508 7 21.0 2. 8 X 10' < 2524.4 < 0.20 12 0 < 11. 0 < 0.24 1 2 5 < 38.4 < 0 09 1.5 < 10.2 < 0 14 1.5 2 6 < 0 14 < 0 5 < 12 8 < 0 25 12.5 < 92 2 GIARDIA II/100L 14263 1 17671 < 11.0 < 38. 4 61.3 < l. 3 12 8 < 92 2 'd 0. > ,....._ (') 0 ::I rt t-' g (l) 0. '-"'

PAGE 110

Table A12 4/9/96 \,!) 00 SAMPLE ID SITE DESCRIPT ION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECAAB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECHL. 060 FINAL EFFL RESERVOIR ----------ENTEROCOCCI TOTAL/FECAL PHAGE ll/100mL COLIFORMS 111100 lll 100mL mL 2 8 X lOs 1 6 x10'/ 6.0 X 10' 1.8 X 10' 972.2 2 8 X 650. 0 3 3 X 101 8 0 14.5/1.5 < 12 5 10 0 3o.o I 5.o < 12 5 < 0 5 5 5 I < < 12 5 0 5 0 5 8 0 I < 0 5 < 12.5 < 0 5 < 0 5 I < 0.5 < 12.5 5 5 30 o I 9 5 < 12 5 ENTEROVIRUS CLOSTRIDIUM CRYPTOSPORIDIUM MPN/100 L ll/100mL II/100L 4 4 X 10' 5 3 X 10' < 1 3 X 101 4 9 2 7 X 10' < 61.3 < 0 .17 9 5 < 22 3 < 0.13 11 0 < 65.2 < 0 066 1 0 < 11.2 < 0.074 0 5 < 9.2 < 0 082 < 0 5 < 12 8 < 0 19 4 5 < 61 7 GIARDIA II/100L 1 3 X 10' 61 3 < 22 3 < 65.2 < 11.2 < 9 2 < 12 8 < 61 7 I i .&" '1:l II) ::l p. t-' > ,....... (') 0 ::l t-' ::l c II) p. '-"'

PAGE 111

APPENDIX B 99

PAGE 112

Experiment 1 9/26/95 3gpm TIME (sec) i SAMPLE : Beads/mL i MS2 phage/mL 60 I 1-1 4 60E+03 I 1.10E+08 I 120 j 1-2 ; 3.98E+04 I 5.70E+08 150 1-3 I 2 .11E+04 I 7 80E+08 180 : 1-4 ; 3 78E+04 I 6.30E+08 210 i 1-5 : 8.20E+03 I 1.50E+08 235 i 1-6 : 4 00E+02 I 5 90E+07 1 270 i 1-7 4 40E+02 8.80E+06 295 1-8 ; 1 12E+02 1 80E+06 320 : l-9 5.50E+01 8 60E+05 345 1-10 5 20E+01 I 4.80E+05 Omin E-1 < 1 0 3 10E+01 30 min E-2 3 60E+01 I 1 00E+01 60min E-3 : 4 30E+01 I 5 40E+02 95 min : E-4 9.00E+OO i 1.10E+01 125 min I E-5 1 .20E+01 1.10E+01 155 min E-6 1 20E+01 I 1 1 0E+01 185 min E-7 2 .00E+OO l 1 40E+01 210 min : E-8 1.00E+OO I 8 60E+01 Background I < 1 0 1 < 1 0 I Seed ; 1 44E+04 I 3 00E+09 PRD1 phage/ml 3 00E+06 5 .00E+ 07 9 90E+07 1 60E+08 2 90E+07 4 50E+06 1.20E+06 3 30E+05 2.30E+05 1 10E+05 1 50E+03 > 10. 0 > 10. 0 : > 10. 0 > 10. 0 > 10. 0 > 10 0 > 10 0 < 1 0 4 40E+08 100

PAGE 113

Appendix B (Continued) TIME (sec) 30 50 75 110 135 160 200 235 270 310 Omin 30min 65 min 95min 125 min 155 min Background : Seed Experiment 2-9/26/95 7gpm SAMPLE : Beads/ml : MS2 phage/mL 1-1 2 70E+03 1.50E+07 1 2 1.08E+04 : 2 50E+08 ; 1-3 6 40E+03 i 2 50E+08 ; 1-4 1 80E+03 l 2.40E+07 1-5 : 6 40E+03 : 1 50E+08 1-6 1 00E+04 I 1.80E+08 1-7 4.40E+03 : 5.70E+07 ' 1-8 1.82E+02 ; 4.70E+06 1-9 3 20E+01 6.20E+05 1-10 2 .00E+OO : 1 90E+05 E-1 1.00E+OO ; < 1 0 E-2 1 70E+01 < 1 0 I E-3 2 .00E+OO I < 1 0 E-4 < 1 0 < 1 0 E-5 < 1 0 < 1.0 I I E-6 < 1 0 < 1.0 i < 1.0 1 20E+01 I I 2.41E+04 : 2 90E+09 i I PRD1 phage/mL 3 00E+08 7.10E+06 1.20E+08 5 90E+07 3 40E+06 8 40E+07 2.00E+07 1 10E+06 1.30E+05 4 20E+04 1 10E+02 7 90E+01 < 1 0 < 1 0 < 1 0 < 1.0 < 1 0 4 40E+08 101

PAGE 114

Appendix B (Continued) Experiment 1 3/5/96 7gpm TIME (sec) SAMPLE i Oocysts/ml. MS2 phage/ml ; 0 1-1 : I 1.00E+06 30 ; 1-2 I I 2 80E+08 ' 60 I 1 3 ; I 2 90E+08 i ' 90 i 1-4 i I 3 .20E+08 i I 120 1-5 i 2 90E+08 I I 150 i 1-6 I 6.50E+07 I 180 j 1-7 i I 4 .00E+06 I 210 I 1-8 i j 1 .90E+07 I 270 ; 1-10 6.55E+02 I I 300 : 1-11 3 .60E+02 I I 330 i 1-12 4 .45E+02 i i 390 i 1-13 : 1.90E+01 I 420 i 1-14 1.00E+OO i i 1 min I E-1 < 0.01 < 1.0 I i Smin E-2 i < 0 .01 I 1.00E+OO I 10 min i E-3 ; < 0.01 < 1.0 15 min E-4 i < 0 .01 I 1.00E+OO 20 min E-5 < 0 .01 I 2 20E+01 I 25 min I E-6 0 .02 5.00E-01 I : 30min : E-7 I 1.20E-01 < 1 0 i 60 min I E-8 : I 0.22 i 2.50E+OO I Background-! 1 ! 3.50E+OO I I Background-E : I 3.00E+OO I Seed I i 5.40E+09 PRD1 phage/ml 1.00E+06 2 30E+07 1 80E+07 2 20E+07 1 40E+07 2 80E+06 2 20E+05 1 30E+06 2 00E+01 1 20E+01 1.40E+01 2.30E+02 4 80E+03 3 10E+04 1 00E+05 8 30E+04 < 1.0 < 1 0 2 60E+08 1.02

PAGE 115

Appendix B (Continued) Experiment 2 3/5/96 3gpm TIME (se c) SAMPLE : Oocysts/mL MS2 phage/mL 0 i 1-1 : 1 00E+06 30 I 1-2 ; i 3 50E+08 60 I 1-3 I 3.80E+08 90 1-4 I i 3 70E+08 120 1-5 I I 2.70E+08 150 1-6 I I 1.60E+08 180 1-7 : I 5.30E+06 I 210 1-8 I I 3.20E+06 240 I 1-9 5 85E+02 ' : 270 1-10 8 60E+02 300 !-11 i 8 20E+02 330 1-12 6.05E+02 ; 360 i 1-13 2 06E+02 390 I 1-14 3 .00E+OO o min E-1 l < 0 .01 I S.OOE-01 10 min ; E-2 : < 0 .01 ; 1.00E+OO 20 min E-3 i 1 00E-02 ; 2.00E+OO 30 min E-4 1.20E-01 I 1.00E+OO 40 min : E-5 0.41 I 1 .50E+OO I 50 min E-6 ; S .OOE-01 1 .50E+OO 60 min E-7 I 0 45 < 1 0 90 min E-8 i 0 .27 l S.OOE-01 120 min E-9 0 15 1.00E+OO 150 min E-10 0 .11 I 1 .00E+OO Background I I i 7 30E+01 Background-E : 2.50E+OO Seed I 4 10E+03 2 20E+08 PRD1 phage/mL 1.00E+06 1.40E+08 1.30E+08 1 50E+08 1 .50E+08 : 6 .00E+07 3 .30E+06 1 .50E+06 : : 5 .50E+OO : : 1 00E+01 9.30E+03 > 100. 0 2 .20E+05 2 .60E+05 i 2.30E+05 1 .80E+05 1 .50E+05 1 .10E+05 3 50E+01 1 70E+01 1.20E+08 103

PAGE 116

APPENDIX C 104

PAGE 117

..... 0 VI Bacter i a and Protozoa H it s afte r Second Stage Recar b o n at i on ( 020 ) 1 .00E+ 02 ..----------------. -l 1 .00E+01 E 0 0 T::::> u. 0 1 .00E+00 I / \ / \ / .... \ / ... . I I . r . \. / \ I :' : v . . . . . . .. I f . . ..... ) 1 ,000 1 00 10 1 1 .00E-01' It H I It Ji H H j.j *0.1 A P R M A Y JUN JUL AUG SEP OCT NOV JAN FEB MAR APR 1995 1 1996 _J 0 0 TC/) ..... C/) >-(.) 0 0 "0 c: ro C/) ...... C/) >-(.) ... E ntero c o c ci ... C perfring en s "*" Crypt ospo r id ium ,. Giardia

PAGE 118

,_. 0 (J'\ Bacteria and Viral Hits after Second Stage Recarbonation (020) _J 1 .00E+01 E 0 0 or:::::> LL 0 1.00E+00 \ \ \ 1 I tl / I I 'I I I 10 1 .00E-01' i i APR MAY JUN JUL AUG SEP OCT NOV JAN FEB MAR APR 1995 1 1996 _J 0 0 or:::::> LL a. Enterococci ... C perfringens _,._Coliphage ..... Enterovirus :x> "0 "0 (I) ::I p. 1-' :< 0 r-.. n 0 ::I rt 1-' ::I c (I) p. ......-

PAGE 119

..... 0 -...J Bacteria and Protozoa Hits in the F i nal Effluent (033) 1.00E+01 100 __J 0 l 0 10 __J I\ en E +"" en 0 I \ >. 0 (.) 1.00E+00 I 0 0 ::J I \ lL '0 0 I \ c: 1 I \ en +"" I en >. I (.) I I 1.00E-01' * I I o. 1 APR MAY JUN JUL AUG SEP OCT NOV JAN FEB MA R APR 1995 1 1996 -Enterococci C. perfringens Cryptosporidium ...... Giardia !l> '0 '0 (l) p.. ..... >: (") ----n 0 rt f-' c (l) p..



PAGE 1

EVALUATION OF MICROBIAL REMOVAL AT A WATER RECLAMATION FACILITY by KELLEY R. RILEY A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science Department of Marine Science University of South Florida December 1998 Major Professor: Joan B Rose, Ph. D

PAGE 2

Graduate School University of South Florida Tampa, Florida CERTIFICATE OF APPROVAL Master s Thesis This is to certify that the Master's Thesis of KELLEY R. RILEY with a major in Marine Science has been approved by the Examining Committee on April 23 1998 as satisfactory for the thesis requirement for the Master of Science degree Examining Committee : Professor: Rose Ph. D Member : JbhD H Ph D Member : Richard 0.' Mines, Jr. P.E.

PAGE 3

DEDICATION To my husband Darryl and my children Kristopher and Morgen for their support and the sacrifices they made so that I could obtain this degree I will forever be grateful. To my mom and sister for their constant support and encouragement and finally to my dad for instilling in me the value of education

PAGE 4

ACKNOWLEDGMENTS I would like to acknowledge the staff at UOSA for their assistance in sample collection and analyses I would also like to acknowledge my advisor and friend Joan Rose for her assistance support and the opportunity she gave me. I thank the people in the lab that I had the pleasure of working with and especially to John Lisle, for his guidance and wisdom

PAGE 5

TABLE OF CONTENTS LIST OF TABLES iii LIST OF FIGURES v ABSTRACT vii CHAPTER 1 : 1 Water Reclamation and Reuse 1 History of Water Reclamation and -Reuse 2 Current Indirect Potable Reuse Projects in the United States 5 Pilotand Demonstration-Scale Projects 7 Summary 11 Microorganisms in Wastewater 13 Current Indicator Microorganisms for Evaluating Water Quality and Treatment 15 Alternative Indicator Microorganisms 16 Development of the Upper Occoquan Sewage Authority 18 CHAPTER 2. RESEARCH OBJECTIVES 20 CHAPTER 3 MATERIALS AND METHODS 22 Sampling Sites 22 Microbiological Sampling 25 Bacteria 25 Protozoa 26 Human Viruses 27 Coliphage 28 Pilot Studies 29 CHAPTER 4 RESULTS 34 UOSA Treatment Processes 34 Bacteria 36 E Human Viruses 47 Coliphage 47 Pilot Studies 55

PAGE 6

CHAPTER 5 DISCUSSION CHAPTER 6 CONCLUSION LIST OF REFERENCES APPENDICES APPENDIX A APPENDIX B APPENDIXC 63 76 78 85 86 99 104 i i

PAGE 7

LIST OF TABLES Table 1 Description of Sampling Sites within UOSA Reclamat i on Facil i ty 24 Table 2 Summary of Microorgan i sm and Method 24 Table 3 Clostrid i um perfringens 39 Table 4 Enterococci 40 Table 5 Total coliform 41 Table 6 Fecal coliform 42 Table 7 Cryptosporidium 48 Table 8 Giard i a 49 Table 9 Enterov i ruses 50 Table 1 0 Col i phage 51 Table 11 Pilot Study on the Removal of Beads and Phage by Chemical Lime Treatment at a F l ow Rate of 7 gpm 57 Table 12 Pilot Study on the Removal of Beads and Phage by Chem i cal Lime Treatment at a Flow Rate of 3 gpm 57 Table 1 3 Pilot Study on the Removal of Cryptosporidium and Phage by Chemical Lime Treatment at a Flow Rate of 7 gpm 58 Table 14 Pilot Study on the Removal of Cryptosporidium and Phage by Chem i cal Lime Trea t ment at a Flow Rate of 3 gpm 58 Table 15 Co rr elation Matrix of Ind i cator and Pathogenic Mi croorgan i sms for Ent i re Treatment Process 70 iii

PAGE 8

Table 16 Table 17 Table 18 Correlation Matrix of Indicator and Pathogenic Microorganisms after Lime Treatment Removal and Inact i vation of Cryptosporid ium, Beads and Phage by Chemical Lime Treatment by Pilot Studies Compared to Monitoring Data Comparison of the UOSA Final Effluent to the Final Effluent Reservoir Water Quality 7 0 71 75 iv

PAGE 9

LIST OF FIGURES Figure 1 Schematic of UOSA Process Description 23 Figure 2 Schematic of UOSA Pilot Plant 30 Figure 3 Average Levels for Positive Samples through the Treatment Train for Bacteria 43 Figure 4 Percentage of Samples Positive through the Treatment Train for Bacteria 44 Figure 5 Log1o Removal of Sites Compared to Site 001 45 Figure 6 Average Levels for Positive Samples for Enteroviruses Protozoa and Coliphage 52 Figure 7 Percentage of Samples Positive for Enteroviruses Protozoa and Coliphage 53 Figure 8 Log1o Removal of Sites Compared to Site 001 54 Figure 9 Fluorescent Bead Removal by Chemical Lime Treatment 59 Figure 10. Cryptosporidium Removal by Chemical Lime Treatment 60 Figure 11. MS2 Removal by Chemical Lime Treatment 61 Figure 12 PRD1 Bacteriophage Removal by Chemical Lime Treatment 62 Figure 13 Comparison of Fecal Coliform Removal to Pathogen Removal throughout the Treatment Plant 64 Figure 14 Comparison of Clostridium Removal to Pathogen Removal throughout the Treatment Plant 65 v

PAGE 10

Figure 15 Comparison of Enterococci Removal to Pathogen Remova l throughout the Treatment Plant 66 Figure 16. Comparison of Enteroviruses and Coliphage in Posit i ve Samples 68 Figure 17 Comparison of Enteroviruses and Clostridium in Postive Samples 69 vi

PAGE 11

EVALUATION OF MICROBIAL REMOVAL AT A WATER RECLAMATION FACILITY by KELLEY R. RILEY An Abstract Of a thes i s subm i tted in partial fulfillment of the requirements for the pegree of Master of Science Department of Marine Science University of South Florida December 1998 Major Professor : Joan B. Rose, Ph D vii

PAGE 12

Water reclamation and reuse have become an important consideration for many communities experiencing increased growth and demand on water resources. Research has been focused on using this reclaimed water for indirect potable reuse, in other words, supplementation of surface waters or groundwaters currently used for drinking water supplies. The Upper Occoquan Sewage Authority (UOSA) Water Reclamation Plant has been reclaiming wastewater and discharging into the Occoquan Reservoir since 1978 This reservo i r serves as a drinking water supply for approximately one million people in Northern Virginia A study was initiated to monitor the bacteria protozoa and viruses entering the water reclamation plant and to evaluate the unit processes for the removal of these microorganisms Eight sites within the plant were monitored monthly for a year for Enterococci, Clostridium, total and fecal coliforms, coliphage, Giardia, Cryptosporidium and enteroviruses Chemical lime treatment with second stage recarbonation and disinfection were the most efficient barriers to the passage of microorganisms Of all the indicators Clostridium and coliphage best reflected the removal of enteroviruses for the chemical treatment system and the disinfection process Abstract Professor : Joan B Rose, Ph.D 6fessor Marine Science Date Approved : Q3) JC19'(: viii

PAGE 13

CHAPTER 1 INTRODUCTION Water Reclamation and Reuse Water reclamation and reuse have become an important consideration and reality for conserving existing potable water supplies due to increased demands on water resources for domestic, commercial, industrial and agricultural uses Water reclamation involves treating wastewater with advanced treatment processes to a high quality so that it can be reused again A wide variety of reclaimed water usages include : landscape and agricultural irrigation ; i ndustrial process water, power plant cooling water ; toilet flushing car washing and augmentation of recreational water bodies (D Angelo 1996) Recla i med water (properly treated wastewater effluent) has been successfully used in the United States for decades to meet nonpotable water needs. Recent research has focused on the feasibility of using reclaimed water for supplementing surface water or groundwater drinking water suppl i es There are two types of potable reuse: indirect and direct. Indirect potable reuse involves treating wastewater to a quality equal to or better than the current water supply and then purposely reintroducing it into a surface water or groundwater that will ultimately be used as a potable water supply for a population Direct 1

PAGE 14

potable re u se involves treating wastewater to a dri n k ing water quality and then purposely introducing it directly into a water treatment plant or potable water distribution system (McEwen and Richardson 1996) The cont i nued depletion of potable supplies has increased the interest in using highly treated reclaimed water to augment potable water resources Currently four planned indirect potable reuse full-scale projects in the United States are found in Virginia Texas Georgia and California (Asano 1995 Asano and Levine 1996 Pia et al1996, McEwen and R i chardson 1996) Tampa Bay, Florida i s currently considering the use of reclaimed water to supplement the existing surface and groundwater supplies A planned direct potable reuse full sca l e faci l i ty is located in South Africa (van Leeuwen 1996, Hattingh and Bourne 1988 Grabow and Isaacson 1978 Hrudey et al 1-991, Asano 1995 Asano and Levine 1995). History of Water Reclamation and Reuse Wastewater reclamat i on and reuse have their roots in the early water and wastewater systems of the Minoan civilization in ancient Greece The use of wastewater for agricultural irrigation dates back 5 000 years (Angelakis and Spyridakis 1996, Asano and Levine 1996) During the nineteenth century large scale wastewater carriage systems were used for discharge i nto surface waters which resulted in the unplanned indirect use of sewage and other effluents for potable water supplies (Asano and Levine 1996) This indirect reuse caused 2

PAGE 15

epidemics of Asiatic cholera and typhoid during 1840-1850 and resulted i n the discovery that the water supply was causing the epidem i cs Improved water treatment techniques, inc l uding the introduction of water filtration the development of alternative water sources using reservoirs and aqueducts and the relocation of water intakes upstream from wastewater discharges allowed some protection of potable water systems During the 20th century programs were developed in the United States for the planned reuse of wastewater The State of California was the first to promote water reclamation and reuse and promulgated the first regulations for reuse in 1918 (Asano and Levine 1996 Crook and Surampalli 1996). Some of the in i t i al reuse programs were developed to provide water for irrigation in Arizona and California in the late 1920s In 1940 chlorinated wastewater was used for steel processing In 1960 urban water reuse systems were developed for Colorado and Florida (Asano and Levine 1996 ) Currently in the 1990s there is increased pressure to develop new sources of water especially i n water poor areas such as the West and the Southwest. Wastewater treatment and pur i fication processes are currently available that can produce water of any quality ; therefore water reuse has become a factor in the planning and efficient use of water resources (Asano and Levine 1996 Crook and Surampall i 1996) The longest history of potable reuse is i n Namibia where potable reuse has provided 10-20% of Windhoek's water supply since 1969 (Asano and Levine 1995 Grabow and Isaacson 1978 Hattingh and Bourne 1989 Asano 1995 van Leeuwen 1996) The South African Council for Scientific and Industrial 3

PAGE 16

Research (CSIR) conducted resea r ch on the technology of water reclamation from secondary effluent since the early 1960s and the Stander Water Reclamat i on Plant was built in 1970 (van Leeuwen 1996) By the end of the 1970s the processes used at the Stander Water Reclamation Plant included coagulation and flocculation, settling ozonation sand filtration, biological granular activated carbon chlorinat i on and stabilization Lack of further funding and difficulties supplying the water from the plant to the consumers caused the project to end but the plant continued to operate exclusively for research purposes The Windhoek Water Reclamation Plant was constructed in 1968 during a devastating drought and followed the design of the Stander Water Reclamation processes This plant was built to supplement the drinking water supply to Windhoek South West Africa and was the first plant in the world to initially reclaim wastewater for the direct supplementation of the city s drinking water supply (van Leeuwen 1996 Grabow 1991 Grabow and Isaacson 1978 Stander and Clayton 1977). Extensive microbiological analyses including enteric viruses parasite ova total and feca l coliforms fecal streptococci Pseudomonas aeruginosa Staphylococcus aureus, and Clostridium pettringens were performed on all water sources and treated supplies when the reclaimed water was first introduced into the system (Grabow and Isaacson 1978 Nupen 1970) and these evaluations were continually expanded Since 1973 epidemiological studies have been coord i nated by the South African Institute for Medical Research to evaluate the health aspects of the reclaimed water. Continuous health monitoring bioassays and epidemiological studies have 4

PAGE 17

proven that the reclaimed water is as safe as the other conventional water supplies (Grabow and Isaacson 1978 van Leeuwen 1996) The first documented case of indirect potable reuse of treated wastewater in the United States was short term and occurred during a severe drought from 1952-1957 at Chanute Kansas when treated wastewater was mixed with water stored in the river channel behind the water treatment dam Chlorinated secondary effluent was collected behind the dam on the river and used as intake water (Asano 1995) The treated water met bacteriological standards for drinking water but was pale yellow and had an unpleasant taste and odor, foamed when agitated and contained a h i gh level of dissolved minerals and organic chemicals Many technological advancements have occurred in wastewater treatment since that time which can produce a high quality water Current Indirect Potable Reuse Projects in the United States The current full-scale operating planned indirect potable reuse projects in the United States are in Texas Virginia Georgia and California These projects demonstrate how indirect potable reuse can be used to augment water resources by incorporating the multiple barrier approach to treatment (Asano and Levine 1995 Asano and Levine 1996 Asano 1995, McEwen and Richardson 1996 Pia et al 1996) Two to five advanced unit processes are specifically used to remove pathogenic microorganisms and trace organics. 5

PAGE 18

The Whittier Narrows Recharge Project (County Sanitation Districts of Los Angeles County) has been surface spreading secondary effluent for infiltration to an underground potable water supply since 1962 The amount of reclaimed water recharged annually averages 16% of the total inflow into the basin The population is estimated to be exposed to 0 to 23 percent of reclaimed water An independent scientific advisory panel to the State of California concluded that the groundwater replenishment project was as safe as the surface water supplies (Asano and Levine 1995, Asano and Levine 1996 Asano 1995 McEwen and Richardson 1996 Pia et al 1996) Water Factory 21 (Orange County Water District) has been blending reclaimed water with deep-well groundwater and using this mixture for deep injection into a heavily used aquifer to prevent salt water intrusion since 1976 (Asano 1995 McEwen and Richardson 1996 Pia et al 1996) The treatment includes lime clarification recarbonation filtration and then half of the flow receives carbon adsorption, the other half of the flow receives reverse osmosis and the entire flow is disinfected Viruses were monitored from 1975 to 1982 in the final effluent and it was demonstrated that the final effluent was essentially free of measurable levels of viruses" and no total coliforms were detected in 179 samples of the effluent tested in 1988 (Crook et al 1989) A planned surface water augmentation project in Georgia is the Clayton County project which utilizes conventional secondary treatment followed by land treatment involving overland flow The treated effluent becomes part of the inflow to a stream that serves as a drinking water source 6

PAGE 19

The Fred Harvey Water Reclamation Plant (EI Paso Public Service Board) recharged reclaimed water to the Hueco Bolson drinking water aquifer since 1985 (Asano 1995 McEwen and Richardson 1996 Pia et al 1996) The water travels to the potable well fields to become part of the potable water supply Treatment of the raw wastewater involves primary treatment act i vated sludge/powdered activated carbon treatment for organic removal nitrification and denitrification lime treatment, recarbonat i on filtration ozonation and granu lar activated carbon adsorption An increase in total dissolved solids content in the aquifer has been observed (McEwen and Richardson 1996). Pilotand Demonstration-Scale Projects Numerous studies have been conducted in an attempt to evaluate the health effects of using reclaimed water in order to supplement existing water supplies or replace the potable supply completely (Crook et al 1989 Asano 1995 McEwen and Richardson 1996 Pia et al 1996 Asano and Levine 1995 Asano and Levine 1996) Health effects refer to the large number of possible effects that can occur after consuming drinking water that is not treated properly Microbiological contaminants such as bacteria, viruses and protozoa can cause acute and chronic health effects (McEwen and Richardson 1996) Currently, toxicological testing is most commonly used to evaluate the effect of complex organic chemical mixtures that may be present in the water 7

PAGE 20

The Pomona Virus Study and the Monterey Wastewater Reclamat i on Study for Agriculture (MWRSA) provided evidence that alternative tertiary treatment systems could effectively remove viruses and potentially pathogen-free effluent could be produced using tertiary treatment and extended disinfection with chlorine (Asano and Levine 1996 Asano and Levine 1995 Asano et al 1992, Yanko 1993 Asano and Mujeriego 1988) The Whittier Narrows Groundwate r Replen i shment Project has been us i ng a m i xture of recla i med water stormwater and surface water to recharge the drinking water aquifer in Los Angeles County since 1962. The amount of recla im ed water in the extracted potable water supply is 0 to 11 percent. A Health Effects Study' begun in 1978 and the major findings published in 1984 stated that both the groundwater and the reclaimed water met Federal Drinking Water Standards ; no viruses were detected in either types of water ; and trace organ i c chem i cals did not exceed the theoretical lifetime risk value (McEwen and Richardson 1996, Asano and Lev i ne 1 996) This landmark study to evaluate the health effects associated with groundwater recharged with recla i med water provided an opportunity to identify the impacts of water reuse on water quality and human health Another objective of the study was to use the data to develop statewide wastewater reclamation criteria for groundwater recharge The study conducted toxicological and chemical stud i es percolat i on studies hydrogeologic studies and epidemiological studies. No measurab l e adverse i mpacts on the groundwater or the health of the populat ion drink i ng the water were found (Asano 8

PAGE 21

and Levine 1995 Asano and Levine 1996 Pia et al 1996 McEwen and Richardson 1996, Nellor 1985) The Potomac Estuary Experimental Water Treatment Plant evaluated the use of the Potomac Estuary as a source of drinking water Approximately 50% of the estuary may be comprised of treated wastewater during drought conditions The treatment plant was operated between 1981 and 1983 Finished water quality was compared to product water from three other treatment plants Microbiological parameters, metals and organics were monitored and two in-vitro toxicological tests were performed. The Ames Salmonella microsome test was used to assess the chemical mutagenesis and mammalian cell transformation assay was used to assess the potential mutagenesis of the reclaimed wastewater. It was determined that the reclaimed water compared favorably with the other product water in terms of the toxicological tests performed (McEwen and Richardson 1996) The City of Denver's Direct Potable Water Reuse Demonstration Project a 5 year project was started in 1985 to determine the feasibility of converting secondary wastewater to drinking water quality and comparing it to the current drinking water supply. Initial research evaluated many treatment processes to determine the optimum treatment sequence This sequence was utilized over a 2 year period to determine if the reclaimed water could meet Federal Drinking Water Standards The reclaimed water and the current drinking water were compared for chron i c toxicity and carcinogenicity in rats and mice A two generation reproductive toxicity study was also conducted No adverse health 9

PAGE 22

effects were reported (Asano 1995 Asano and Levine 1995 McEwen and Richardson 1996 Pia et al 1996 Lauer 1991, Lauer and Rogers 1996 Cond i e et al1994). The San Diego Total Resource Recovery Project began in 1988 to determine if raw sewage could be treated to a quality comparable to the existing Miramar reservoir raw water supply The findings indicated there was no difference in viral concentration in the two waters : the microbiological quality of the reclaimed water was better than the Miramar surface water with lower total coliform counts ; Giardia cysts were not detected in the reclaimed water or the Miramar reservoir ; and the reclaimed water met the microb i ological criteria for recreational waters without disinfection The study concluded that the health risk associated with the use of the reclaimed wa ter was less than or equal to the existing raw water supply (Asano 1995 Asano and Levine 1995 McEwen and Richardson 1996 Gagliardo et al 1996 Danielson et al 1996 de Peyster et al 1993) Tamp a Water Resource Recovery Project is a pilot project that began in 1986 to determine if a reclaimed secondary effluent could be produced to a quality that could be blended with existing surface and groundwaters that are currently used for potable supplies The reclaimed water met all primary and secondary drinking water standards, no organic chemicals were present and it was determined that the reclaimed water was an acceptable raw water supply (McEwen and Richardson 1996 Asano 1995 Final Report CH2M Hill 1993 Hemmer 1994) 10

PAGE 23

Summary The potential for indirect potable reuse is being seriously considered by many communities in the United States and other countries due to the diminishing supply of potable water resources An American Water Works Association (AVWVA) policy on reuse was described by Bergman who notes there are still many obstacles to be overcome, including acceptance of non potable reuse for edible crops and for use inside buildings" (Bergman 1994 ) A review article by Smith (1995) however notes that reuse programs are regulated on a state-by-state basis and more attention is being focused on reclaiming wastewater for potable uses In summarizing various demonstration and pilot projects conducted, six research projects were identified as the most useful for advancing the knowledge of water reclamation and reuse of wastewater. These projects included microbial risk assessment monitoring identification of new indicators of pathogenic microorganisms evaluation of effects of process selection on particle size distribution seasonal storage for reclaimed water nonpotable water management, and evaluation of metals and synthetic organic chemicals in irrigation water (Crook 1994). Van Riper and Geselbracht (1996) stated that the initial success of reclaimed water projects will be sustained if the public perceives the reuse of wastewater is healthy and desirable Federal regulations for water reuse do not exist in the United States However the United States Environmental Protection Agency (EPA) published 11

PAGE 24

Guidel i nes for Water Reuse in 1992 including a rev iew of sta t e standards ( C r oo k and Surampalli 1996 US EPA 1992) Th i s has led to the deve l opment of c riteri a by individual states which has resulted i n a variation of regulations among the d i fferent states The most common parameters monitored i nc l ude b i ochem i ca l oxygen demand (BOD) turbidity or total suspended solids (TSS), total or fecal coliform bacteria nitrogen and chlorine contact time and res i dual. If there is likely to be public contact with the reclaimed water tert i ary treatment i s required to produce a finished water that is virtually pathogen-free ( State of California 1978 Florida Department of Env i ronmental Regulat i on 1990 State of Ari zona 1991 ) Variations among states include the use of the tota l co li form as the indicator organism in California while the fecal coliform is used as the ind i cator organism in Florida Texas and Arizona Florida is the only state that requires mon i toring for TSS ; the other 3 states monitor for turb i dity California and Florida specify treatment processes wh i le Texas and Ar i zona do not. Californ i a and Florida have developed the most comprehensive regula ti ons address i ng t h e many uses of reclaimed water There is still concern regarding the use of highly treated wastewater to supplement water supplies One of the concerns is the emergence of waterborne pathogens 12

PAGE 25

Microorganisms in Wastewater Wastewater contains a wide variety of microbial pathogens, i e bacteria protozoa and viruses (Bitton 1994) The pathogenic protozoa of concern include Cryptosporidium and Giardia. Giardia is a common cause of wate r borne disease in the United States (Craun 1988) and in 1985, Cryptosporidium caused the largest waterborne disease outbreak in Milwaukee which affected over 400 000 people (MacKenzie e t al 1994) Cryptosporidium an enteric coccidian protozoan, has been recognized as a pathogen in humans since 1980 (Madore et al1987, Rose 1988 Lisle and Rose 1995 Marshall et al 1997) Cryptosporidiosis an parasitic infection principally of the intestinal tract, causes profuse watery diarrhea abdominal pain nausea vomiting and fever The disease is se l f-limiting in immunocompetent individuals but can be fatal for immunocompromised ind i viduals The oocyst (the environmental stage of the organism) is extremely resistant to conventional disinfection processes and continues to be implicated in waterborne outbreaks throughout the world (Campbell et al1982, Korich et al1990, Peeters et a l 1989) Cryptosporidium oocysts are ubiquitous in the water environment. Rose et al ( 1988) indicated that 91% of the sewage samples examined contained varying levels of oocysts. The oocysts have been shown to survive in flow-through chambers of river water and tap water after 176 days with die-off rates approximately 95% (Lisle and Rose 1995) The fecal material may protect the oocysts from desiccation thus prolonging the viability of the organism. Sewage 13

PAGE 26

discharge could be a significant source of contamination of oocysts in the environment. Giardia, a flagellated protozoa has been recognized as one of the most common parasites of humans in the United States and the most common cause of waterborne outbreaks (Rose et al 1991 Rose et al 1989, Rodgers et al 1995, Hibler and Hancock 1990) Giardiasis causes diarrhea, abdominal distension flatulence and malaise The Giardia cyst is commonly found in raw sewage in fairly large numbers and has also been shown to be resistant to conventional disinfection processes (LeChevallier et al 1991, Hibler and Hancock 1990) Enteroviruses are a group of human viruses that replicate initially in cells of the intestinal tract These small (22 nanometers) viruses include polio virus, coxsackie virus and echo viruses Another enteric virus of concern is hepatitis A virus The enteric viruses cause illnesses such as diarrhea aseptic meningitis, conjunctivitis myocarditis and hepatitis and are found on a routine basis in untreated wastewater (Gerba and Rose1990). The water industry has become sensitive to protozoan contamination of water supplies. The recent promulgation of the Information Collection Rule (ICR) by the Environmental Protection Agency (EPA) will require drinking water facilities serving over 100,000 population to monitor their finished water for Cryptosporidium Giardia and Enteroviruses (USEPA 1994) This has caused a renewed interest in the enteric protozoa and other microbial contaminants in wastewater which may impact water supplies and their control by advanced treatment processes. Cryptosporidium has been found in raw wastewater at 14

PAGE 27

concentrations of 850-13 700 oocysts/L and Giardia at levels of 3375 cysts/L (Rose et al 1996 Madore et al 1987) Wastewater facilities discharging into watersheds serving as drinking water supplies may need to ensure that the effluent is nearly pathogen-free or that levels of these microorganisms have been reduced to some level of acceptability. Current Indicator Microorganisms for Evaluating Water Quality and Treatment Current regulations in most states for discharg i ng wastewater are based on the concentration of coliform bacteria in the final wastewater effluent (Crook and Surampalli 1996 US EPA 1992) Coliforms are classified as total or fecal coliforms Total coliforms are any aerobic or facultat i ve anaerobic gram negative non-spore fo r ming bacillus that ferment lactose and gas at 37 C after 24 hours (Standard Methods for the Examination of Water and Wastewater 1992) Total col i forms are the presumpt i ve ind i ca tor of fecal contamination and are used in drinking water as a standard for d i s i nfection The fecal coliform a subgroup of the total coliform group, are normal inhabitants of the human and animal intestines. Fecal coliforms are differentiated from total coliforms by incubation at an elevated temperature of 44. 5 C (Standard Methods for the Examination of Water and Wastewater 1992) The presence of fecal coliforms confirms fecal contamination and indicates the increased possibility of water contamination by enteric pathogens The absence of fecal coliforms does not 15

PAGE 28

necessarily guarantee that the pathogens are not present. Coliform bacteria have been the indicators of choice for evaluating disinfection processes during water treatment. Numerous researchers have demonstrated that the coliform standard is not adequate for evaluating the effi cacy of treatment primarily disinfection of viruses and protozoa (Baker and Hegarty 1997 Snowdon and Cliver 1989 Fujioka and Shizumura 1985 Dutka 1973, Funderberg and Sorber 1985 Gerba et al 1979 Metcalf 1978 Cabelli 1977) Viral and protozoan waterborne outbreaks have occurred with drinking water supplies that met current U.S. EPA standards for total coliforms and turbidity (Gerba and Rose 1990 Rose et al 1985 MacKenzie et al 1994 LeChevallier et al 1991 Seligmann and Reitler 1965 Boring et al 1971 Keswick et al 1985, Payment and Armon 1989) Few studies have been done in wastewater in an attempt to correlate indicator coliform bacteria with viruses or protozoa Alternative Indicator Microorganisms Alternative microorganisms such as Enterococci Clostridium perfringens and F-specific coliphage, have been proposed as better indicators of water quality, fecal pollution and public health risks (Cabelli 1977 Fujioka and Shizumura 1985, Armon and Kott 1996 Snowdon and Cliver 1989 Grabow 1990) However little data are available on the use of alternative indicator microorganisms (Enterococci Clostridium perfringens and coliphage) compared 16

PAGE 29

to the conventional indicator microorganisms (total and fecal coliforms) for indicating the presence of pathogens in wastewater effluent. Enterococcus is a subgroup of the fecal streptococci bacterial cocc i of fecal orig i n found i n both an i mals and humans Taxonomically these bacteria possess the group D antigen and conform to the Sherman criteria (C l ausen et al 1977) The enterococcus group includes Streptococcus faecium S faecalis S durans and related biotypes (Clausen et al 1977) Enterococcus generally appears to be more pers i stent than either bacterial pathogens or fecal coliforms (Cohen and Shuval 1973, Dav i es-Galley et al 1994 Sinton et al 1994) C perfringens is an enteric gram posit i ve anaerobic spore-fo r ming pathogenic bacterium found in feces. Although there were considerable controversies about using Clostrid i um as a water quality indicator (Cabelli 1977) more recently a number of scientists (Fujioka and Shizumura 1985 Payment and Franco 1 993) recommend C perfringens as a valuable supplement to other water quality tests due to its spore-forming property particularly in situations where detection of viruses or remote fecal pollution i s desirable Th i s microorganism is consistently present in wastewater at concentrations of 1 03 to 104 colony-forming units (CFU)/1 00 ml, and its resistance to chlorination and other environmental factors is similar to the enter i c viruses and protozoa (Fuj i oka and Shizumura 1985 Payment and Franco 1993) F-specific coliphage is a virus that infects E. coli bacteria and can be found in fecally contaminated water These coliphages contain RNA and one of the host specific characterist i cs is the i r adsorption to long filamentous structures 17

PAGE 30

the F-pili on bacteria (Snowdon and C l iver 1989) Coliphages are acellular and approximately the same size as Enteroviruses so t hey have been suggested as adequate model organ i sms for enteric viruses in water (Have l aar et al 1993 Funderberg and Sorber 1985) Development of the Upper Occoquan Sewage Authority In the late 1960 s increased populat i on growth in the Occoquan Watershed led to the degradation of the Occoquan Reservo i r The V i rginia Water Control Board developed a Policy for Waste Treatment and Water Quality Management in the Occoquan Reservoir' in 1971 because of this degradation This policy mandated the construction of the Upper Occoquan Sewage Authority (UOSA) Water Reclamat i on Plant a state of t he art treatment facility to reclaim all the wastewater generated in the watershed (Robbins 1993) The Upper Occoquan Sewage Authority Water Reclamation Plant has been reclaiming wastewater and discharging to the Occoquan Reservoir s i nce 1978 (Robb i ns 1 985). Th i s reservoir serves as a potable water supply for approximately one million people in Northern Virginia (Asano and Levine 1995, Asano and Levine 1996 Asano 1995 McEwen and Richardson 1996 Pia et al1996). This is one of two planned surface wate r augmentation projects currently in operation in the United States at this time ( Pia e t al1996) Ten to fifteen percent of the reservo ir i s compr i sed of recla i med water on average but during t i mes of drought as much as 90% of the flow into the reservo i r comes from the plant d i scharge (McEwen 18

PAGE 31

and Richardson 1996) Treatment of the water includes primary and secondary treatment along with five advanced wastewater treatment processes including chemical treatment with high lime and recarbonation multimedia filtration granular activated carbon filtration ion exchange and chlorination/dechlorination Treated water is blended with the supply in the reservoir and later treated in a conventional water treatment plant before delivery to customers in that area. Although in operation for many years no studies had been undertaken on the removal of pathogenic microorgan i sms from this reclamation plant. 19

PAGE 32

CHAPTER2 RESEARCH OBJECTIVES The objectives of this project were to evaluate the removal of microorganisms commonly found in wastewater through processes at the UOSA advanced water reclamation facility The specific objectives of this study were to: 1) Examine the removal of bacteria (rout i ne indicators and alternative indicators) protozoa human viruses and coliphage and determ i ne which unit process demonstrated the greatest reduction of microorganisms 2) Determine i f the use of alternative microorgan i sms provided data to better reflect the occurrence of pathogens in wastewater. 3) Evaluate a pilot plant for the removal of Cryptosporidium This study took place at the Upper Occoquan Sewage Authority (UOSA), a 27 million gallons per day (mgd) Water Reclamation Plant located in Northern Virginia. An evaluation of the treatment processes within UOSA was necessary to determine if microbial levels, particularly protozoa and viruses, were reduced in the final effluent. The comparison of the treated effluent with the water quality in the Occoquan reservoir was a major 20

PAGE 33

objective of the study The impact of future regulations such as the Information Collection Rule (ICR), was an important issue in initiating this study 21

PAGE 34

Sampling Sites CHAPTER 3 MATERIALS AND METHODS UOSA treatment system consists of a series of barriers : conventional treatment followed by five advanced treatment processes (Figure 1 ) Samples were collected once per month for one year (from April 1995 through April 1996 with the exception of December 1995) from the eight sites within the UOSA Water Reclamation Plant. A high flow event associated with a large rainfall was sampled in January 1996 Each of eight sites were mon i tored for bacteria, viruses and protozoa. Sampling sites included the headworks untreated wastewater after preliminary screening (001 ) ; secondary effluent (011 ) ; second stage recarbonation effluent (020) ; multimedia filter influent after passage through the open ballast ponds (017); multimedia filter effluent (021 ) ; ion exchange bed effluent (023) ; the dechlorinated final effluent (033) ; and UOSA's final effluent reservoir (060) (Table 1 ) The ion exchange system did not operate in the ammonia-removal mode during this study but did operate as a post GAC treatment filter The data collected were used to assess the levels of microbial contaminants entering the plant the levels after the various treatment processes, and the reduction of microorganisms through the unit processes. The methods used are summarized in Table 2 and described in detail thereafter 22

PAGE 35

N w Figure 1. Schematic of UOSA Process Description ,,.. Conventional Tre a tment Advanced 001 Sertlnlng and Grit Biolog i cal Rttyc le filler Headwork p, ..... Olgut er l Compo ttlng a .......; obil Compost Filter Prttlltt 021 H .. dworke l OeweCtrtd Soli d to Land Dltpo181 or Lendllll .. Exchlngt Flntl Elllutnt RtttiYOir Chlorlnttlon/ Otchlorlnttlon 060 033

PAGE 36

Table 1 Description of Sampling Sites within UOSA Reclamation Fac i lity SITE UNIT PROCESS 001 Headworks untreated wastewater post preliminary screening 011 Secondary effluent 020 Second stage recarbonation post lime treatment (high pH 11. 3) 017 Filter influent post carbonation post passage through open ballast pond 021 Multi-media filtered effluent 023 Granular activated carbon (GAC) contactor effluent 033 Final plant effluent post dechlorination 060 Final effluent reservoir Table 2 Summary of Microorganism and Method ORGANISM Enterococci Total col i forms Fecal coliforms Clostridium METHOD Membrane filtration mE agar/EIA agar Membrane filtration mEndo agar Membrane filtration mFC agar Membrane filtration mCP agar REFERENCE Standard Methods for the Examination of Water and Wastewater 9230C Standard Methods for the Examination of Water and Wastewater 92228 Standard Methods for the Examination of Water and Wastewater 92220 Armon and Payment 1988 ................ Enterovirus 1 MDS filter US EPA 1994 Adsorption elution BGM cells -c/YiJiaspandlum .. ---cart"rra9e-ti"itraiiO n-------Ros eaTaf 1 9 9 f--IFA Giardia Cartridge filtration Rose et al 1991 IFA 24

PAGE 37

Microbiological Sampling Bacteria For each sampling event a single one-liter grab sample was collected in a sterile 1 liter sample bottle from each site for bacterial and d i rect coliphage analyses Samples were analyzed upon collection for the bacteria at the UOSA laboratory (Table 2). The membrane filtration technique detailed in Standard Methods for the Examination of Water and Wastewater was used to determine Enterococci densities (Standard Methods for the Examination of Water and Wastewater 1992 Method 9230C). One-hundred ml aliquots of each sample were assayed in duplicate and up to 200 ml volumes were sampled for disinfected water Dilutions of the samples were used as necessary for s i tes 001, 011 and 017 The membrane filters were p l aced on mE agar (Difco Detroit Ml) and incubated for 48 hours at 41C The membranes were then transferred to EIA agar (Difco, Detroit Ml) and incubated at 41 C for 20 minutes. The pink to red colonies were evaluated for a black or reddish-brown precipitate on the of the filter and identified as enterococci. A membrane filtration technique was used to determine Clostridium perfringens densities (Payment and Franco 1993 Cabelli 1977) One-hundred ml aliquots of each sample were assayed in duplicate as previously described The membrane filters were placed on mCP agar (Acumedia Baltimore MD) 25

PAGE 38

and incubated at 45C in an anaerobic gas-pak jar for 18-24 hours. The plates were placed in a ziploc bag containing a petri dish filled with ammonium hydroxide for 1 minute to identify C perfringens colonies Colonies that turned a bright pink were identified as C perfringens The membrane f i ltration technique detailed in Standard Methods 92228 and 9222D (Standard Methods for the Examination of Water and Wastewater 1992) was used to determine total coliform and fecal coliform densities respectively One-hundred ml aliquots of each sample were assayed in duplicate as previously described The membrane filters were placed on M Ende medium (Difco Detroit Ml) for total coliforms and incubated at 35 C for 24 hours Red colonies that produced a metallic green sheen were identified as total coliforms The membrane filters were placed on M -FC medium (Difco Detroit Ml) for fecal coliforms and incubated in a water bath at 44. 5C for 24 hours Blue colonies were identified as fecal coliforms Protozoa A slightly modified version of the ICR protozoan protocol was utilized for the collection and detection of Cryptosporidium oocysts and Giardia cysts (Federal RegisterNol. 59, No 28/February 10, 1994 Appendix C to Subpart M Proposed ICR Protozoan Method for Detecting Giardia cysts and Cryptosporidium oocysts in Water by Fluorescent Antibody Technique) Protozoan samples were collected by filtration through a 1.0 um nominal 26

PAGE 39

porosity 10-inch yarn wound cartr i dge filter (Microwynd AMF Cuno Balt i more MD) The volume of water filtered was monitored by attached flow meters After collect i on the filters were put in ziploc baggies and placed on ice for transport to the University of South Florida Upon receipt at the University of South Florida the filter was cut and washed to recove r the accumulated debris and density gradients were used to separate the oocysts and cysts from the sediments. The final concentrate was filtered onto 0.22 urn cellulose acetate membrane filters in duplicate and stained with monoclonal antibodies. These monoclonal antibodies are tagged with a f l uorescent label fluorescein isothiocyanate (FITC), which specifically binds to the oocyst and cyst wall and when examined using epifluorescen t microscopy the oocyst or cyst fluoresce green. Equ i valent volumes from the concentrates which were examined under the microscope were calcu l ated and the concentration of cysts and oocysts per 100 L were determined Human Viruses All samples were analyzed for enterov i ruses by Dr Sam Farrah at the University of Florida The Proposed Virus Monitoring Protocol was utilized for the collection and detection of human enterovi r uses (Federal RegisterNol. 59 No 28/February 10, 1994 Appendix D to Subpart M Proposed Virus Monitoring Protocol) Samples were collected by filtration through a 1 0-inch pos i tively charged pleated cartridge filter designed to capture viral particles (1 MDS 27

PAGE 40

George Edwards Company Pelham AL) The volume of water filtered was monitored by attached flow meters After collection the filters we r e secured in ziploc baggies and placed on ice for transport to the Univers i ty of F l orida. The human enteric viruses were eluted from the filters usi n g beef extract and concentrated by a flocculation method. The v i ruses were grown on BGM cell culture i n 75 cm2 flasks The flasks were evaluated daily for cell destruction caused by viruses Positive and negat i ve cells were passaged once into new cells for confirmation An MPN method was used to enumerate the concentration of enteroviruses Coliphage An aliquot of water f rom the 1 liter grab samp l e was ana l yzed using an aga r overlay technique described by Adams 1959 Escherichia coli (E. coli American Type Culture Collection ( ATCC) #15597 Rockville MD ) was used as the bacterial host and was grown to stationary phase 24 hours before each coliphage assay. A one to two -ml aliquo t of the sample was added to a tube containing 3 ml of tryptic soy broth (Difco Detroit Ml) containing 1 5% agar (Difco Detro i t Ml) kept liquid at 48 C then 0 1 ml of host bacteria was added mixed and poured onto a tryptic soy agar petri plate. Replicate plates were set up in order to assay 8 0 ml from most sites. Dilutions ( made with sterile phosphate buffer solution ) of the samples were necessary for untreated water The plates were incubated at 37 C for 24 hours After 24 hours the petri plates 28

PAGE 41

.were removed from the incubator and examined for the presence of plaques (clearings in the bacterial lawn) Plates containing less than 300 plaque-forming units (PFU) were counted Pilot Studies A pilot facility representing the chemical treatment process was built at UOSA and two separate challenges were conducted (Figure 2) Initially the pilot facility was evaluated for detention time using a fluorogenic dye It was determined that the flow rate of 3 gpm simulated the surface loading rate and the flow rate of 7 gpm simulated the hydraulic detention time of the full-scale system. The actual and theoretical detention times were 130 and 150 minutes respectively for the 3 gpm flow rate and 54 and 64 minutes respectively for the 7 gpm flow rate. The pilot plant influent water was secondary effluent drawn from the line feeding the full-scale facility This was seeded during the first trial with fluorescent beads (3 um in diameter as a surrogate for Cryptosporidium oocysts) and two bacterial viruses, MS2 coliphage and PRD1 bacteriophage Fifty (50) ml each of MS2 stock and PRD1 stock (1011/ml) and one ml of approximately 109 fluorescent beads were added to 1 L of secondary effluent in a 3 L carboy. The influent was injected over a 6-minute time period with a peristaltic pump An influent sampling port was set up and samples were collected every 30 seconds to 1 minute Effluent samples were collected in 500-ml bottles every 30 minutes 29

PAGE 42

w 0 Isolation Valve From Rapid Mix In Figure 2. Schematic of Pilot Plant Flowmeter Roll Two 90 ELS Sample Wlthdrawl Teo w/flex, tygon & pinch clamp Polymer InJection Filling Sludge Drain w/Valva Sampling Tee w/ Valve to Plant Waste

PAGE 43

for 2 to 3 hours. The effluent samples were neutralized with 1 N sulfuric acid immediately upon collection The second pilot study was seeded with MS2 coliphage and PRD1 bacteriophage and formalinized Cryptospor i dium parvum oocysts. One hundred(100) ml each of MS2 stock and PRD1 stock (1 011/ml) were added to 1 L of secondary effluent in a 3 L carboy. Five ml of 1 08/ml formalinized Cryptosporidium parvum oocysts was added approximately 1 minute after the injection of the phage to prevent the inactivation of MS2 and PRD1 due to the formalin present in the oocyst stock solution The samples were collected as previously described MS2 coliphage (ATCC catalog number 15597-81) were propagated for use in the pilot studies by inoculating a 1 L flask containing 200 ml of tryptone yeast extract (TYE) with 2 0 ml of the host bacteria. Escherichia coli (E. coli) (ATCC catalog number 15597) The culture flask was _placed in a shaking incubator maintained at 37C When the bacterial density reached approximately 1 08/ml colony forming units/ml -(CFU/ml) (wh.ich had been previously determined) an aliquot of the virus stock (approximately 1 012/ml PFU/ml), was added to provide a multiplicity of infection (MOl) of 0 1 The culture was shaken continuously unt i l the host cells lysed Then 02 g. of lysozyme and 6 0 ml of sterile 0 2M ethylenediaminetetraacetic acid (EDTA) were added to the culture to lyse the host cells and release tbe virus (bacteriophage) and the sample was incubated for an additional 30 minutes i n a shaking incubator at 37 C. The propagated virus and cellular debris were then centrifuged for 20 minutes at 3600 31

PAGE 44

rpm and filtersterilized using a 0.45 urn sterile membrane filter. The resulting stock was titered by the agar overlay technique and refrigerated at 4C until needed PRD1 bacteriophage was propagated in the same manner with the host bacteria Salmonella Pilot influent samples were diluted and effluent samples were assayed directly for the bacteriophage using an agar overlay technique with each of the respective hosts as previously described (Adams 1959). Plaques were enumerated after 24 hours incubation at 3JOC. Samples containing oocysts were filtered and stained with monoclonal antibodies The inf l uent samples were assayed directly (0.1 to 1.0 ml) for the oocysts by filtering the sample onto 0 .22 urn cellulose acetate membrane filters (Sartorius) using a Hoefer manifold and stained with monoclonal antibodies (Crypt-a-Gio, Waterborne Inc. New Orleans LA). These monoclonal antibodies are tagged directly with a fluorescent label, fluorescein isothiocyanate (FITC) which specifically binds to the oocyst wall and when examined using epifluorescent microscopy the oocysts fluoresce green. The membrane filters were mounted onto a glass slide using 2% 1 ,4-Diazabicyclo[2 2 2) octane (DABCO) and a coverslip and the entire membrane filter was counted under 400X epifluorescent microscopy. A similar procedure was used for the fluorescent bead samples except the monoclonal antibodies were not necessary The sample was filtered onto a membrane filter mounted onto a glass slide with DABCO and examined microscopically The effluent samples were concentrated by centrifugation at 1050 x g if necessary or 1 0 to 20 ml of sample were filtered 32

PAGE 45

onto 0 22 um cellulose acetate membrane filters in duplicate and then stained with monoclonal antibodies The effluent samples for the fluorescent beads wer e processes similarly with the exception of staining with monoclonal antibodies Fluorescent beads and oocysts/ml were calculated in the influent and the effluent. 33

PAGE 46

UOSA Treatment Processes CHAPTER4 RESULTS The UOSA treatment system consists of a series of processes : conventional and secondary activated sJudge treatment followed by five advanced treatment processes (Robbins 1993) Primary treatment consists of screening grit removaJ, and primary clarificat i on. Secondary treatment is provided by a complete-mix activated sludge system comprised of biological treatment clarification and return activated sludge pumping The act iv ated sludge system includes pre-aeration basins (selectors) that promote a more efficient microbial culture. In the biological reactors microorganisms (activated sludge) decompose organic pollutants and form a biological floc which is separated from the water by settling in the secondary clarifie r s A portion of the settled floc is returned to the selectors to activate the biological reactions The remainder is combined with primary sludge and transferred to the anaerobic digestion system The digested sludge is currently dewatered and composted. The conventional system is followed by high pH chemical treatment, filtration, granular activated carbon (GAC) adsorption ion exchange 34

PAGE 47

and disinfection using sodium hypochlorite followed by dechlorination using sulfur dioxide. The high pH chemical treatment system includes rap i d mix basins flocculation basins chemical clarification first stage recarbonation clarification, and second stage recarbonation. Calcium hydroxide is the principal coagulant and carbon dioxide is used for recarbonation Mechanical mixers in the rapid mix basins completely mix the coagulant with the wastewater The pH increases to 11. 3 thereby enhancing inactivation of some microorganisms and precipitating phosphorus heavy metals and suspended solids (Grabow et al 1978) The small coagulated particles (floc) are then slowly stirred in the flocculation basins to promote aggregation into larger particles, which settle more readily in the chemical clarifiers Following chemical clarification the pH is restored to neutral by two stage recarbonation with intermediate clarification In first stage recarbonation, the pH is adjusted to 9 7 which causes precipitation of carbonates and other materials. The carbonates form a small dense floc which settle readily in the recarbonation clarifiers After recarbonation clarification more carbon dioxide is added to adjust the water to pH 7 The chemical treatment effluent flows into ballast ponds for flow equalization and is pumped at a uniform rate through the remaining advanced treatment processes Multimedia pressure filters are used to remove suspended sol i ds and turbidity A backwash equalization tank prevents hydraulic surges Granular activated carbon (GAC) adsorption performed in an upflow expanded bed removes a wide range of synthetic organic compounds GAC loses its adsorptive 35

PAGE 48

capacity over time and is periodically regenerated in an on-site multiple-hearth furnace The ion exchange columns remove ammonia and activated carbon fines UOSA S unoxid i zed nitrogen standard (1. 0 mg/L) is normally met by nitrification in the secondary treatment systems ; but during drought conditions nitrogen is removed in ammonia form by the ion exchange system The ion exchange system does not operate in the ammonia-removal mode very often but can operate as a post GAC treatment filter Disinfection is accomplished by chlorination Sodium hypochlorite is applied to the ion exchange effluent at the chlorinat i on mix station The mixing station is capable of breakpoint chlorinating ammonia remaining after nitrification or ion exchange treatment. The chlorinated water then flows through the chlorine contact chamber The contact chamber usually contains the effluent for 30 minutes of contact time and the average total residual chlorine (TRC) after contact time is 1 5 mg/L with a standard deviation of 0.3 mg/L and the free residual chlorine (FRC) average is about 70% of the TRC. The water is then dechlor i nated with sulfur dioxide prior to discharge into UOSA s final effluent reservoir. Bacteria Clostridium perfringens were detected at an average of 3. 7 x 104 and 4.4 x 1 03 CFU/1 00 ml in the untreated and secondary wastewater respectively The 36

PAGE 49

second stage recarbonation had an average of 5.1 CFU/1 00 ml, and the multi media influent averaged 16 5 CFU/100 ml. The multi-med i a effluent and the carbon adsorption effluent both averaged 1 8 and 2.2 CFU/1 00 ml ( Table 3 ). Clostridium perfringens was detected in the final effluent post dechlorination at an average concentration of 3 5 CFU/1 00 ml in 1 0% of the samples and at an average concentration of 4 9 CFU/1 00 ml in 78% of the final effluent reservoir samples (Figures 3 and 4) Clostridium was reduced by the high pH chemica l treatment by approximately 3 log10 (99. 9%) and the overall reduction for Clostridium was 5 log10 (99 999%) (Figure 5) The enterococci were detected i n the untreated wastewater at an average of 5 0 X 105 CFU/100 ml, an"d lhe secondary effluent averaged 2 2 X 103 CFU/1 00 ml. Enterococci were reduced to average levels of 23. 5 1 06.4 12 and 10. 8 CFU/1 00 ml in the second stage recarbonation multi-media influ e nt and effluent and carbon adsorption effluent respectively (Table 4) Ente r ococci were detected in the final effluent post dechlorination at an average concentration of 2 7 CFU/1 00 ml in 17% of the samples and at an average concentration of 10 1 CFU/1 00 ml in 73% of the final effluent reservoir samples (Figures 3 and 4). The overall reduction for Enterococci was 6 log10 (99 9999%) (Figure 5) Total coliforms were detected in untreated wastewater and secondary effluent at averages of 2.4 x 107 and 1 7 x 105 CFU/1 00 ml, respectively. Levels were reduced from 121 CFU/100 ml after second stage recarbonation to an average of 1.6 CFU/1 00 ml in the final effluent postdechlorination (Table 5) 37

PAGE 50

Four of 11 samples were positive (36%) for total coliforms in the final effluent postdechlorination and average levels increased in the final effluent reservoir to an average of 180 3 CFU/1 00 mL in 100% of the samples (Figures 3 and 4) The overall reduction for total coliforms was 7 log10 (99 99999%) (Figure 5) Fecal coliforms were detected in the untreated wastewater and secondary effluent at averages of 9 0 x 105 and 7 8 x 103 CFU/1 00 mL respectively (Table 6) Levels were reduced from 23 CFU/1 00 mL to <0 5 CFU/1 00 mL in the final effluent dechlorination Levels increased to 568 CFU/1 00 mL prior to the multimedia sand filter (post passage through the open ballast pond) and also increased in the final effluent reservoir to an average of 22 9 CFU/1 00 mL in 100% of the samples (Figures 3 and 4) Conventional treatment and chemical treatment achieved a total 4 6 log10 reduction of fecal coliforms The overall reduction for fecal coliforms was 6.2 log10 (>99 9999%) (Figure 5) 38

PAGE 51

w \0 Table 3 Clostridium perfringens (CFU/100 mL) Sample site ----> 001 011 020 017 021 023 033 060 # of samples 9 9 10 10 10 10 10 9 # positive 9 9 9 9 7 5 1 7 % samples positive 100% 100% 90% 90% 70% 50% 10% 78% Sensitivity Limits < 0 5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 Minimum CFU/100 mL 15,000 311 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 Maximum CFU/100 mL 57,000 28,000 12 95 6.0 6.0 3. 5 12.5 Mean (arithmetic) : Positives only 36,667 4,452 5.1 16.5 1.8 2.2 3.5 4.9 All samples 36,667 4,452 4.6 14.9 1.3 1.1 0.35 3.8 REMOVAL EFFICIENCY UNIT PROCESS % 87.85 99.9 -224 91.28 15.4 68.18 N / A CUMULATIVE % 87.85 99.99 99.96 99.996 99.997 99.999 N/A Clostridium perfringens; a gram positive, spore forming, capsulated, gas producing, nonmotile, anaerobic bacillus and is a normal inhabitant of the intestinal tract of man and animals. Food poisoning is caused by c perfringens, in rare cases intestinal gas gangrene. The data show the greatest absolute removal of C. perfringens was in conventional treatment, but the high pH chemical treatment provided the greatest log reduction. Removal efficiencies for processes following chemical treatment were perhaps underestimated to some extent by very low influent counts. ,. ,. L

PAGE 52

0 Table 4 Enterococci (CFU/100 mL) Sample site ----> 001 011 020 017 021 023 033 060 # of samples 12 12 11 12 12 12 12 11 # positive 12 12 10 10 8 10 2 8 % samples positive 100% 100% 91% 83% 83% 83% 17% 73% Sensitivity Limits < 0.5 < 0.5 < 0.5 < 0.5 < 0 5 < 0.5 < 0.5 < 0.5 I I Minimum CFU/100 mL 50,000 233 < 0.5 < 0.5 < 0.5 < 0 5 < 0.5 < 0.5 Maximum CFU/100 mL 870,000 11,000 80 370 45.5 59 4.5 61 Mean (arithmetic) : Positives only 500,000 2,183 23.5 106.4 12 10.8 2 7 10. 1 All samples 500,000 2,183 21.4 88.7 10 9 0.45 7.4 REMOVAL EFFICIENCY UNIT PROCESS % 99.56 99.0 -314 88.73 10.0 95. 0 N/A CUMULATIVE % 99.56 99.996 99.98 99.998 99.998 99.9999 N/A Enterococci is a name commonly used for some bacteria within the genus Streptococcus. These organisms are generally found in human and animal feces. Enterococcus appears to be more persistent in the environment than either bacterial pathogens or fecal coliform. The data show conventional treatment ( 011) and chemical treatment (020) had about equal removal efficiency for enterococci. Together these two unit processes achieved a 4.4 log1 0 reduction of enterococci. On average, the UOSA plant achieved a 6 log1 0 reduction of enterococci during this study. N/A -Not applicable

PAGE 53

..... Table 5 Total coliform (CFU/100 mL) Sample site ----> 001 011 020 017 021 023 033 060 I # of samples 10 10 11 10 11 11 11 6 # positive 10 10 11 10 11 11 4 6 % samples positive 100% 100% 100% 100% 100% 100% 36% 100 Sensitivity Limits < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 <0.5 Minimum CFU/100 mL 2.4*106 24,000 0.5 20 5.5 2.0 < 0.5 17 Maximum CFU/100 mL 42*106 610,000 800 >16 1 000 >1, 6oo >1,600a 4.0 600 Mean (arithmetic) : Positives only 24*106 170,000 121 1,822 178 166 1.6 180 All samples 24*106 170,000 121 1,822 178 166 0.58 180 REMOVAL EFFICIENCY UNIT PROCESS % 99.29 99.93 -1,701 90.23 6.74 99.65 CUMULATIVE % 99.29 99.999 99.992 99.999 99.9993 99.9999 99 Total coliform are any aerobic or facultative anaerobic, gram negative, non spore forming, bacilli that ferment lactose to acid and gas at 37 C after 24 hours. Most of the total coliform species are widespread in the environment. Some total coliform species reside mainly in the intestines of human and animals, and are short lived in the environment. Total coliform are the presumptive indicator of fecal contamination and are used in drinking water as the standard for disinfection. Conventional treatment and high pH chemical treatment had similar removal efficiency for total coliform. These two processes achieved a total 5 log10 reduction of total coliform. The total plant achieved a 7.6 log10 reduction of total coliform on average during this study. Samples were too numerous to count at dilutions used; analyzed by MPN method

PAGE 54

"-' Table 6 Fecal col i f orm (C FU/100 mL) -----Sample site ----> 001 011 020 017 021 023 033 060 # of samples 11 11 10 11 11 10 11 8 # positive 11 11 9 11 10 9 0 8 % samples positive 100% 100% 90% 100% 91% 90% 0% 100 Sensitivit y Limits < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 < 0.5 Minim u m CFU/100 m L 90,000 1,200 0.5 1.0 < 0.5 0.5 < 0.5 3 0 I Maximum CFU/100 mL 5.7*106 41,000 130 5,000 240 130 < 0.5 71 I Mean (arithmetic) : Positives only 900,000 7,764 23 568 37 21 < 0 5 22.9 All samples 900,000 7,764 20.7 568 33.6 18.9 < 0.5 22.9 REMOVAL E FFICIENCY I UNIT PROCESS % 99.14 99.73 -2,744 94.08 43.75 > 97.6 N/A CUMULATIVE % 99.14 99.998 99.937 99.996 99.9979 >99.99999 N/A Fecal coliform, a sub group of total coliform, are normal (usually non-pathogenic) inhabitants of human and animal intestine. Fecal coliform are differentiated from total coliform by incubation a t an elevated temperature of 44.5 C and fermentation of lactose. Fecal coliform is the confirmation indicator for fecal contamination, and indicates a good chance the water may be contaminated by enteric pathogens. Absence of fecal coliforms does not guarantee absence of viruses and protozoa. Conventional treatment and chemical treatment achieved a total 4 6 log10 reduction of fecal coliform. Fecal coliform were not detected in the dechlorinated effluent. N/A -Not applicable -'

PAGE 55

+:w _J E 0 0 ::> lL 0 Figure 3. Average Levels for Positive Samples through the Treatment Train for Bacteria 1 .000E+081 .000E+07 1,000,000 1 0 000 1 ,000. 100-10 1 001 011 020 .... 017 '* 021 023 Sampling Sites 033 ..... Enterococci ->K C lo s t ridium +Tota l Co liform s +Fec al Coliform s 12 samples No fecal coliforms detected in final effluent 060

PAGE 56

""'" ""'" 120-100 Q) > :E 80 (/) 0 0... 60 1 ...... c Q) 0 Q> 40 -0... 20 0 Figure 4. Percentage of Samples Positive through the Treatment Train for Bacteria 001 011 020 017 021 023 033 Sampling Sites """ Enterococci *Clostridium +Total Coliforms Fecal Colifo rms 12 samples Clostridium 1 0% positive in the final effluent Fecal Coliform not detected in final effluent 060

PAGE 57

.!> V1 Figure 5 Log 10 Removal of Sites Com pared to Untreated Wastewater Log10 Removal 10r-------------------------------------. 8 6 4 2 0 011 020 017 021 023 033 S a mpling Sites C l ostrid ium Q h otal Coliforms 0 Fecal Coliforms

PAGE 58

Protozoa Cryptosporidium oocysts were detected in the u n treated wastewater at an average of 1 ,484/1 00 L. No oocysts were detected in the secon d ary effluent but two of 12 samples in the second stage recarbonation were pos i tive at an average concentrat i on of 3.45 oocysts/100 L (Table 7). Cryptosporidium oocysts were detected in one of the 12 samples (8%) in the final effluent post dechlorination a t a concentrat i on of 0.44 oocysts/1 00 L. One of 11 samples were positive for Cryptosporidium at a co n centrat i on of 5. 7 oocysts/1 00 L in the final effluent reservoir (Fi gures 6 and 7) Convent i onal and h i gh pH chem i cal treatment reduced the oocysts by 99.96% Overall reductions for Cryptosporidium were at least 41og10(99.99%) (Figure 8). Giardia cysts were detected in the untreated wastewater and secondary effluent at averages of 4 9 x 104 and 2 3 x 103 CFU/1 00 L, respectively (Table 8). After second stage recarbonation one of 12 samples (8%) in the mult i media effluent was positive for Giardia cysts at a concentration of 61. 3 cysts/1 00 L. Two of 12 samples (17%) were pos i tive for Giardia cysts in the final effluent post dechlorination at an average concentration of 6 6 cysts/1 00 L Levels increased in the final effluent reservo i r to an average of 42 9 cysts/1 00 L i n 18% of the samples (Figures 6 and 7) Conventional treatment and high pH chemical t r eatment achieved a 2.7 log10 reduction and the overall plant achieved a 4.6 log10 reduction (Figure 8) 46

PAGE 59

Human Viruses Enteroviruses were detected in untreated wastewater and secondary effluent at averages of 1 085 and 23.6 PFU/1 00 L. Levels were reduced to 0.2, 0.3 0 116 and 0 123 PFU/1 00 L in the second stage recarbonation, multi-media i nfluent and effluent and GAC effluent respectively (Table 9) Enteroviruses were not detected after chlorination or in the final effluent reservoir (Figures 6 and 7). Overall plant reduction of enterovirus was greater than 41og1D(99 99%) (Figure 8). Coliphage Coliphage were detected in the untreated wastewater at an average of 3 8 x 105 PFU/1 00 ml and 1 8 x 103 PFU/1 00 ml in the secondary effluent. Coliphage were reduced to an average of 28, 142 33, and 25 PFU/1 00 ml through the second stage recarbonation multi-media influent and effluent and carbon adsorption effluent respectively (Table 1 0) Coliphage were not detected in the final effluent postdechlorination but levels increased to 20. 6 PFU/1 00 ml in 45% of the samples in the final effluent reservoir (Figures 6 and 7) Overall reductions across the plant were 4.4 log10 (Figure 8) 47

PAGE 60

.1:' Table 7 Cryptosporidium (Oocysts/100 L) Sample site ----> 001 011 020 017 021 023 033 060 # of samples 12 12 12 12 12 12 12 11 # positive 2 0 2 2 1 2 1 1 % samples positive 100% 0% 17% 17% 8.3% 17% 8.3% 9.1% Sensitivity Limits < 15.0 <12.4 < 1.0 < 1.0 < 0.42 < 0.9 < 1.2 < 1.95 Min oocysts/100 L 277.8 <12.4 2.1 6.4 < 0.42 < 0.9 < 1.2 < 1.95 Max oocysts/100 L 2,690 <2500 4.8 56.3 0.9 2.82 0.44 5.7 Mean (arithmetic) : Positives only 1,484 N / A 3.45 31.3 0.9 2.7 0.44 5.7 All samples 1,484 <312.2 0.575 5.217 0.075 0.45 0.037 0.52 REMOVAL EFFICIENCY UNIT PROCESS % 78.96 99 .816 -807 98.56 -300 0.8370 N / A CUMULATIVE % 78.96 99.961 99.65 99.995 -99.97 99.998 N/A Cryptosporidium is the name assigned to a fairly large variety of very small protozoan oocysts. The oocysts do not normally occur in high concentrations in the natural environment Each oocyst has the potential to release a maximum of four viable Cryptosporidium sporozoites which can initiate infection. Thus, the potential for infection from ingesting one oocyst is quite high. Some oocysts are pathogenic to man, causing chronic diarrhea which in severe cases in the immunocompromised can b e fatal. The data indicate the combination of conventional and high pH chemical treatment reduced the plant influent oocyst population by 99.96%. On average the total plant achieved a 99.998% (4.6 log1 0 ) reduction. N / A -Not applicable

PAGE 61

1.0 T able 8 Giardia (C ysts/100 L) Sample site ----> 0 0 1 0 1 1 020 017 021 023 033 060 # of samples 12 12 1 2 12 12 12 12 11 # positive 12 8 6 2 1 0 2 2 % samples positive 100% 67% 50% 17% 8.3% 0% 17% 18.2% Sensitivity L imits < 15.0 < 10. 0 < 3.0 < 4.6 < 1. 0 < 1.2 < 1.2 < 5.7 M in. cysts/100 L 2,246 24. 5 < 3 0 < 4.6 < 1.0 < 1.2 < 1.2 < 5 7 Max cysts/100 L 142,631 17,671 728 326.8 61.3 < 1.2 12. 8 7.5 Mean (arithmetic): 48,691 2,297 170 220 61.3 < 1.2 6 6 42. 9 Positives onl y 48,691 1,531 8 5 3 6.7 5.1 < 1.2 1.1 7 8 All samples REMOVAL EFF I CIENCY UNIT PROCESS % 96.856 94.45 56.82 86.10 76. 4 > 8.3 N/A CUMULATIV E % 96.856 99.825 99.925 99.989 99.9975 99.998 N / A Giardia is the name for a group of single-celled, flagellated, pathogeni c protozoa found in a variety of vertebrates, including, mammals, b irds, and reptiles. These organisms exist as trophozoites (active or feeding stage form) inside the host intestinal tract or as cysts excreted in the feces, depending on the stage of their life cycle. Giardia lamblia (causes diarrhea) is the clas sical exampl e of the grou p associated with humans a n d its cysts are founc in wastewater in fairly large numbers. Conventional treatment and high pH chemical treatment a chieved similar removal efficiencies for Giardia lamblia. Combined, these t wo processes achieved a 2 7 log10 reduction. On average t h e total p lant achieved a 4.6 log1 0 reduction. N/A -N ot ap p licab l e

PAGE 62

1..11 0 Table 9 Enteroviruses (MPN/100 L} Sample site ----> 001 011 020 017 021 023 033 060 # of samples 1 2 12 12 12 12 12 12 11 # positive 12 1 2 1 3 3 3 0 0 % samples positive 100% 100% 8.3% 25% 25% 25% 0% 0 % Sensitivity Limits < 1.0 < 1. 0 < 0.012 < 0.03 5 < 0 .043 < 0.056 < 0.05 < 0.14 Minimum CFU/100 mL 150 1.4 < 0.012 < 0.035 < 0.043 < 0.056 < 0.05 < 0 .14 Maximum CFU/100 mL 4,600 120 0.22 0.7 0.27 0.18 < 0 .13 < 0.91 Mean (arithmetic} : Positives only 1,085 23.6 0.22 0.3 0.116 0.123 < 0.085 < 0.28 All samples 1,085 23.6 0.018 0.1548 0.09 0.097 < 0.085 < 0 .28 REMOVAL EFFICIENCY UNIT PROCESS % 97.8 99.92 -760 41.86 -7. 8 > 12.4 N/A CUMULATIVE % 97.8 99.998 99.99 99.992 99.991 > 99.992 N/A .. Enteroviruses are a group of viruses that replicate initially in cells of the intestinal tract. These small (22 nm.} viruses include polio virus, coxsackie and echo viruses, and Hepatitis A virus. They are ribonucleic acid (RNA} v iruses. I n this study a cumulative 99.998% rem o val (3. 38 log reduction} of enteroviruses was achieved by the combination of conventional and c h emical treatment. Enteroviruses were not detected in dechlorinated final effluent. The plant overall removal efficiency was 99.999% or a 4.3 log10 reduction. N/A -Not applicable

PAGE 63

VI ._. Table 10 Coliphage (PFU/100 mL) Sample site ----> 001 011 020 017 021 023 033 060 # of samples 12 12 12 12 12 12 12 11 # positive 12 12 4 8 3 1 0 5 % samples positive 100% 100% 33% 67% 25% 8.3% 0% 45% Sensitivity Limits < 12.5 < 12.5 < 12.5 < 12.5 < 12.5 < 12. 5 < 12.5 < 12.5 Minimum CFU/100 mL 60,000 200 < 12.5 < 12.5 < 12.5 < 12. 5 < 12.5 < 12.5 Maximum CFU/100 mL 860,000 5,500 75 790 75 25 < 12.5 50 I Mean (arithmetic) : Positives only 380,000 1,821 28 142 33 25 < 12.5 20. 6 All samples 380,000 1,821 9.3 94.6 8.3 2.1 < 12.5 9.4 REMOVAL EFFICIENCY UNIT PROCESS % 99.52 99.49 -1017 91.226 74.7 50 N / A CUMULATIVE % 99.52 99.998 99.975 99.998 99.999 99.999 N / A Coliphage are viruses that use Escherichia coli bacteria as their host. The direct method detected large numbers of coliphage in the plant influent. The method did not detect Coliphage in the dechlorinated effluent, thus the reported mean values ( < 12. 5 PFU/100 milliliter) are a function of the method sensitivity. The data seem to suggest direct method assay may be more appropriate for highly contaminated water than for clean water. The p lant achieved a 4.4 log10 reduction of coliphage as measured by the direct assay method. N/A -Not applicable

PAGE 64

lJl N Figure 6 Average Levels for Positive Samples for Enteroviruses, Protozoa and Coliphage* 1 000 000 _J 100,000 0 0 T" 10, 000 (/) ... (/) >. 0 0 1 000 0 100 (/) (/) >. 0 10 lL 0... 1 -. x x. .... 0 1 001 011 020 017 021 023 Sampling Si tes *Phage *Ente r ovi ru s X Cryptosporidium Giardia Coliphage results PFU/1 00 ml No enteroviruses detected in final effluent or reservoir No coliphage d ete c t ed in final e ffluent ........ x-033 ... ... 060

PAGE 65

1.11 w Q) ;!::: en 0 a.. ..... c Q) 0 .... Q) a.. Figure 7. Percentage of Samples Posit i ve for Enteroviruses, Protozoa and Coliphage 120 100. \ \ \ 80 \ \ \ \ ..... 60 40 20 --.:_ 0 ,, 001 011 020 017 021 023 033 Sampling Sites *Coliphage Enterovirus XCryptosporidium Giardia 12 samples 060

PAGE 66

Vl Figure 8. Log1 0 Removal of Sites Compared to Untreated Wastewater Log1 0 Removal 6.-------------------------------------, 5 4 3 2 1 0 011 020 017 021 023 033 Sampling Sites .Phage [8) Ent e r ovirus Q Cryptosporidium !ZIGiardia

PAGE 67

Pilot Studies Tables 11 and 12 show the results of the fluorescent bead and bacteriophage experiments at 7 gpm and at 3 gpm. Tables 13 and 14 show the results for the oocyst and bacteriophage experiment at the same two flow rates The influent total numbers were calculated by multiplying the average influent levels (#s/ml) with the total time Figures 9 1 0, 11, and 12 diagram the removal of fluorescent beads Cryptosporidium oocysts MS2 and PRD1 bacteriophages respectively by chemical lime treatment at 7 gpm and at 3 gpm. The influent was injected over 4 minutes and monitoring showed levels of 1 x 1 04 fluorescent beads/ml after 1 minute (Figure 9) The retention time in the pilot faciliity was 2 hours Beads were first detected in the effluent after 30 minutes and ranged in levels between 1 and 36/ml. After 3 hours the level dropped off to zero. Cryptosporidium oocysts were detected in the influent at concentrations of 6 x 1 02/ml after 3 minutes (Figure 1 0) Oocysts were also first detected after 30 minutes and ranged from 0 02 to 0 .22/ml. Similar results were observed for MS2 and PRD1 bacteriophage with influent concentrations of 1 x 1 08 PFU/ml after 1 minute (Figures 11 and 12) PRD1 bacteriophage was detected first in the effluent after 5 minutes and at higher concentrations than MS2 The bacteriophage were inactivated by the high pH which was maintained at 11. 2 throughout the experiments MS2 coliphage was more sensitive to inactivation with 99.9999% reduction as compared to PRD1. A large difference was observed between the two sets of experiments for PRD1. PRD1 was inactivated 55

PAGE 68

99. 999% in the first experiment ; only 94 to 95% inactivation was observed i n the second experiment. This may have been due to better attention to the neutralization of the high pH immediately upon collect i on of the sample in the second experiment. The removal of the formalinized oocysts ranged from 99% to 99.65% while the beads at either flow rate were removed by approximately 98% 56

PAGE 69

Table 11 Pilot Scale Study on the Removal of Beads and Phage by Chemical Lime Treatment at a Flow Rate of 7 gpm Fluorescent MS2 Phage PRD1 Phage Beads Average Influent 4 25 X 10;j 9 3 X 101 5 88 X 101 levels (#s/mL) Time (minutes) 5.17 5 17 5 17 Influent Total 2 197 X 104 4 .82 X 108 3 04 X 108 Numbers Average Effluent 6 67 0 0 1 15 X 10 levels (#s/mL) Time (minutes) 65 65 30 Effluent Total 433 150 3.45 X 10 Numbers Percent 98.02 > 99 99996 98 86 Reduct i ons Table 12 Pilot Scale Study on the Removal of Beads and Phage by Chemical Lime Treatment at a Flow Rate of 3 gpm Fluorescent MS2 Phage PRD1 Phage Beads Average Influent 1 13 X 104 2 3 X 10 3.47 X 10 levels (#s/mL) Time (minutes) 5 75 5 75 5 .75 Influent Total 6 .5x104 1 33 X 109 1.997 X 10 Numbers Average Effluent 14.4 85.4 10 levels (#s/mL) Time (minutes) 210 210 210 Effluent Total 3024 1 79 X 104 2100 Numbers Percent 95 35 99.998 99.9989 Reductions 57

PAGE 70

Table 13 Pilot Scale Study on the Removal of Cryptosporidium and Phage by Chemical Lime Treatment at a Flow Rate of 7 gpm Cryptosporidium MS2 Phage PRD1 Phage Average Influent 296 1 59 X 105 1 03 x 101 levels (#s/ml) Time (minutes) 7 3.5 3 5 Influent Total 2072 5.55 X 105 3.6 X 107 Numbers Average Effluent 0.12 9 56 2 .74x104 levels (#s/ml) Time (minutes) 60 60 60 Effluent Total 7 2 573.75 1 .64 X 10 Numbers Percent 99 65 99 99989 95.4 Reductions Table 14 Pilot Scale Study on the Removal of Cryptosporidium and Phage by Chemical Lime Treatment at a Flow Rate of 3 gpm Cryptosporidium MS2 Phage PRD1 Phage Average Influent 513.2 1.9x 105 7.95 x 101 levels (#s/ml) Time (minutes) 6 5 3.5 3 5 Influent Total 3335 8 6 65 X 105 2 78 x 105 Numbers Average Effluent 0 22 1 0 1.15x10 levels (#s/ml) Time (minutes) 150 150 150 Effluent Total 33 67 150 1.73 X 101 Numbers Percent 99 .01 99.99998 93.8 Reductions 58

PAGE 71

V1 \0 _J E en "0 Cd Q) Ill Figure 9 Fluorescent Bead Removal by Chemical Lime Treatment 100,000.-----------------------, 1,000 100 10 \ ,.,.-.,<. 0 .Q. '\.. ',"o.o-o, \ a. 1 0 1 ('I') ...l() ('I') C\J lO lO f'-. ('I') ('I') l() .ql() l() C\J l() 0 0 lO ll> ll> ll> l() 0 oo C\JW ...-...-C\J C\J ('I') l() ...-...-...-C\J Time (Minutes) *Influent 3gpm 0 Effluent 3gpm .s:;;.l nfluent 7gpm *Effluent 7gpm

PAGE 72

"' 0 _J E -en ..... en >-0 0 0 Figure 1 0 Cryptosporidium Removal by Chemical Lime Treatment 1 ,000. 100 10 1 0 1 0 .01 0.001 I I I }J I ., I 0 -o--o ...... 0 -...... o ... ., .,
PAGE 73

0\ ,_. .....J E :J u. a.. 1 .000E+09 1. 000E+08 1 0 0 0 E + 0 7 1,000,000 100,000 10,000 1 ,0001 1 oo I 101 1 F i gure 11. MS2 Remova l by Chem i cal Lime T r eatme n t I I 0 0 I ' o o o-o 0 1 C') -yo-10 C') (\J 10 10 ,...._ C') C') 10 .q-10 1.{) (\J 10 0 0 10 1.0 I{) I{) 10 0 00 NOO .... .., -.-.-.-(\J -.--.-(\J (\J C') 1.{) Time (Minutes) Infl uent 3gpm 0 Effluent 3gpm -Q-I n fluent 7gpm ... Eff lu en t 7gpm

PAGE 74

0\ N _J E ::J LL a.. Figure 12 PRD1 Bacteriophage Removal by Chemical Lime Treatment 1 .000E+09 1 .000E+08 1 .000E+07 1 ,000,000 100,000 1 0 ,000 1 ,000 100 10 1 <) I I I ... 0o o o o ... Time (Minutes) *Influent 3gpm ()-Effluent 3gpm 7gpm .... Effluent 7gpm

PAGE 75

CHAPTERS DISCUSSION Five of the unit processes that removed the m i croorganisms included acti vated sludge treatment chemical lime t reatment filtration GAC and chlorina t ion Microbial removal was at least 99. 9% (3 log10) through chemical lime treatment and further reducti on occurred after chlor i nation The coliform bacteria as expected, were less resistant to the un i t processes than the pathogens (Figure 13) While the fecal coliform is the indicator used for discharge requ i rements the data clearly show that Giardia cysts were not removed to the same level as the fecal coliforms through the unit processes. Alternative indicators such as Clostridium and Enterococci demons t rated increased res i stance to the unit processes and showed sim i lar removals to the protozoa (Figures 14 and 15) Clostridium appears to be the best conservative indicator because it can still be found after reduction through the unit processes Possibly the spore-forming properties of this microorganism are similar to the oocyst or cyst stage which allows for increased resistance against phys i cal and chemical treatment processes Coliphage have been suggested as a superior indicator for human viruses particularly for d i sinfection efficacy The percent of samples positive for 63

PAGE 76

0\ Figure 13. Comparison of Fecal Coliform Removal to Pathogen Removal throughout the Treatment Plant Log1 0 Removal 6 5 4 3 2 1 0 Act. Sludge Chemical Lime Filtration Un i t Processes GAC Chlori nation &.1 Fecal Colif o rms Cryptosporidium ID Giardia

PAGE 77

(j\ V1 Figure 14. Comparison of Clostridium Removal to Pathogen Removal throughout the Treatment Plant Log1 0 Removal 5 4 3 2 1 0 Act. Sludge Chemical Lime Filtration Unit Processes GAG Chlorination Cryptosporidium OJ Giardia l3J Enterovirus

PAGE 78

0\ 0\ Figure 15. Comparison of Enterococci Removal to Pathogen Removal throughout the Treatment Plant Log1 0 Removal 6 5 4 3 2 1 0 Act. Sludge Chemical Lime Filtration Unit Processes GAC Chlorination Cryptosporidium []Giardia I8J Enterovirus

PAGE 79

Enteroviruses and coliphage concentrations throughout the unit processes were similar It is interesting to note how closely coliphage indicates the presence of viruses in wastewater (Figure 16) Coliphage concentrations were higher than enteroviruses in both unit processes that demonstrated the greatest removal of microorganisms (high pH chemical treatment and disinfection). Clostridium concentrations were also higher than the Enteroviruses (Figure 17) The ability of coliphage to indicate the presence of pathogens in the reclaimed water provides evidence of its potential for serving as an additional alternative indicator microorganism. Clostridium showed increased resistance to the unit processes as compared to both total coliforms and fecal coliforms This supports research that has demonstrated the use of coliphage and possibly Clostridium for assessing the reliability of treatment processes (Rose et al 1996, Grabowet al1978, Baker and Hegarty 1997 Armon and Kott 1996 Fujioka and Shizumura 1985, Cabelli 1977) Analysis of the entire data set by Pearson s Correlation Coefficient was performed and the results were somewhat misleading, due to the high correlations suggested (Table 15) This may be due to the fact that the concentrations of microorganisms were high at the primary and secondary sampling sites. Analysis of the data after chemical lime treatment by Pearson s Correlation Coefficient showed no correlations with any of the indicator microorganisms to Cryptosporidium The strongest correlations for Enteroviruses were shown with Enterococci Clostridium and coliphage with values of 0.5456, 0 5295 and 0 3523, respectively The strongest correlations for Giardia were 67

PAGE 80

0\ 00 120100 -Q) 80 "(j) 0 a.. 60 c Q) 0 .... Q) 40-a.. 20 o 001 12 samples Figure 16 Comparison of Enteroviruses and Coliphage in Positive Samples 011 020 017 021 023 033 Sampling Sites *Coliphage *Entero viru s 060

PAGE 81

0\ \0 120-100 Q) 80 (/) 0 a.. 60 -c Q) 0 L. Q) 40 a.. 20 o 001 12 samples Figure 17. Comparison of Enteroviruses and Clostridium in Positive Samples ** "* "* 011 0 2 0 017 021 0 2 3 033 Sampli n g S i tes >IE C l os t rid i um Enter o v i rus 060

PAGE 82

demonstrated with Clostridium and coliphage with values of 0.3819 and 0 3538 respectively (Table 16) Table 15 Correlation Matrix of Ind i cator and Pathogenic Microorganisms for Entire Treatment Process Organism Total Fecal Entero Clostri Crypto Giardia Coliphage Virus Fecal 0.9498 1 .000 0 9378 0.8359 0 1142 0.8006 0 8929 0 8762 coliforms Enterococci 0.9052 0 9378 1 .0000 0.8362 0 1366 0 8404 0 8834 0 8982 Clostridium 0.8689 0 8359 0.8362 1 0000 0.1531 0 .7455 0.8330 0 .7935 Crypto 0 1223 0 1442 0 1366 0 1531 1.0000 0.1894 0 1565 0 1721 Giardia 0 .7572 0 8006 0 8404 0 7455 0.1894 1.0000 0.7941 0.8705 Coliphage 0 8593 0.8929 0 8834 0 8330 0 1565 0 .7941 1 .0000 0 .8473 Enterovirus 0.8454 0 8762 0 8982 0.7935 0 1721 0 8705 0 8473 1 0000 Table 16 Correlation Matrix of Indicator and Pathogenic Microorganisms after Lime Treatment Organism Total Fecal Entero Clostri Crypto Giardia Coliphage Virus Fecal 0 2448 1.000 0 6915 0.4324 0.1603 0 5084 0 5269 coliforms 0.0606 Enterococci 0.3063 0 .6915 1 0000 0 5461 0.0204 0.2474 0 3948 0.5456 Clostridium 0 4786 0 4324 0.5461 1.0000 0.0438 0.3819 0 4473 0 .5295 Crypto 0.0426 -0 0606 0 0204 0 0438 1.0000 0.2177 0 0830 0.0202 Giardia 0 2446 0.1603 0 2474 0 3819 0 2177 1.0000 0 3538 0 .3799 Coliphage 0.2970 0.5084 0 3948 0 4473 0 0830 0.3538 1 0000 0.3523 Enterovirus 0.2679 0.5269 0 5456 0.5295 0 .0202 0.3799 0.3523 1 0000 Pilot scale studies of the chemical lime treatment process demonstrated removals of Cryptosporidium oocysts by 99% which was also observed during the full-scale process For the coliphage the indigenous monitoring showed a 99% reduction and represents the assay of a heterogenous population of 70

PAGE 83

coliphage some of which may be very sens it ive to the high pH (such as MS 2) and some of which may be more resistant to the high pH (such as PRD1 ). More information should be gathered regarding the resistance of PRD1, which may be a more valid surrogate for the high pH chemical treatment for the human enteroviruses which are known to be more resistant to h i gh pH than the coliphage Table 17 compares the removals of the pilot studies and the removals estimated by the i ndigenous monitoring program Table 17 Removal and Inactivation of Cryptosporidium Beads and Phage by Chemical Treatment by P il ot Studies Compared to Monitoring Data Condition Oocyst Bead MS2 PRD1 Indigenous Evaluated Removal Removal Removal/ Removal/ Coliphage (%) Inactivation Inactivation Removal/ Inactivation Pilot Scale 98.99 99.53 99. 9999 93. 8 NA 3gpm 99. 9999+ 99 999+ Pilot Scale 99.65 98. 02 99. 9999 95.4 NA 7gpm 99 99999+ 99 999+ Full-Scale 99.8 NA NA NA 99.5 Monitoring +Experiment 1, perhaps problems with the neutralization of the sample Removal efficiencies and log reductions were greatest through secondary treatment and high pH chemical treatment. This was similar to results reported by Grabow and Isaacson (1978) that excellent reductions were observed for microorganisms at an operational pH of 11. 2 After secondary and chem i cal treatment system detection of indicator bacteria coliphage enteroviruses and 71

PAGE 84

protozoa were sporadic and the levels were near the limits of detection of the various methods The results in this study were similar to those for enterovirus removal through the reclamation plant as described by Yanko (1993) and Rose et al (1996). Analysis of 10 years of enteric virus monitoring from 6 tertiary treatment water reclamation plants i n California found only 1 sample out of 590 samples was positive for enteric viruses in the final effluent (Yanko 1993). Removals of viruses through secondary treatment was at least 99. 8%. The primary barrier in these plants was disinfection In the California studies chlorine contact times averaged approximately 90 minutes with final total residuals of 4-5 mg/L. The virus monitoring data from this 1 0 year period confirmed the results of the Pomona Virus Study and suggest that the viral risks associated with the use of reclaimed water are within acceptable levels These studies also found that male specific coliphage were more resistant to standard disinfection practices A study conducted at a water reclamation plant by Rose et al (1996) in St. Petersburg, Florida evaluated filtration and disinfection at 4 mg/L for a minimum of 30 minutes. This study demonstrated a 5 log10 removal for enteroviruses but 8% of the samples were positive for enteroviruses in the storage tank at low levels (approximately 1/500 L) Asano and Mujeriego state that for efficient virus removal and inactivation two major operating criteria must be met: (1) the effluent must be low in suspended solids and turbidity prior to disinfection to reduce shielding of viruses and chlorine demand and (2) sufficient disinfectant dose and contact time provided for the wastewater (Asano and Mujeriego 1988) 72

PAGE 85

Giardia cysts and Cryptosporidium oocysts were reduced by 4.1 and 3 3 log10, respectively but 25% and 16% of the samples were positive for Giardia cysts and Cryptosporidium oocysts in the storage tanks (Rose et al1996). UOSA has two major barriers for viruses : chemical lime treatment and disinfection. This is different than other studies and probably is superior since no human viruses were detected in the final effluent. The true removal (only >) cannot be calculated Enteroviruses were not detected in any of the samples from the post dechlorination effluent at an average total residual chlorine of 1 5 mg/L or in the final effluent reservoir but levels of bacteria and coliphage were higher unless the residual was kept above 1 9 mg/L. Total coliforms enterococci, Clostridium, Cryptosporidium and Giardia were all detected in the final effluent. UOSA treatment processes achieved approximately 4 log10 reduction with 17% and 8% of the samples posit i ve for Giardia cysts and Cryptosporidium oocysts respectively, in the final effluent postchlorination. Levels of the microorganisms increased slightly (with the exception of enteroviruses and Giardia) after passage of the effluent through the open ballast pond prior to multi-media filtration which was not suprising with the wild geese population at the plant. The percentage of samples positive and the concentrations were compared for the treated reclaimed water and the final effluent reservoir water. In every case the treated final effluent was of better quality than the water in the UOSA effluent reservoir (Table 18) The concentration and percent of samples positive increased for the indicator microorganisms and the pathogens in the final 73

PAGE 86

effluent reservoir The naturally occurr ing indicator bacteria and protozoa detected in the reservoir were most likely contr i buted from animals No human viruses were detected in the effluent reservoir during th i s study 74

PAGE 87

-....j V1 Table 18 Comparison of the UOSA Final Effluent to the Final Effluent Reservoir Water Quality Microorganism Percentage of Average I Samples Concentration for Positive Positive Samples Effluent Final Effluent Effluent Final Effluent Reservoir Reservoir Clostridium1 10 78 3 5 4.9 Total coliforms 1 36 100 1 6 180.3 Fecal coliforms 1 0 100 < 0.5 22 9 Enterococci1 17 73 2.7 10.1 Coliphage;l 0 45 < 12.5 20.6 Enterovirus;, 0 0 < 0.085 < 0 .28 Cryptosporidium4 8.3 9.1 0 .44 5.7 Giardia5 17 18.2 6 6 42.9 CFU/1 00 ml, ;lPFU/mL, "PFU/1 00 L, qOocysts/1 00 L, Cysts/1 00 L

PAGE 88

CHAPTER 6 CONCLUSION This is the first major study to examine comprehensively bacteria protozoa, alternative indicators and viruses through a full-scale advanced wastewater treatment facility. One year of monitoring for a total of 96 samples supported the following conclusions: Chemical lime treatment with second stage recarbonation is the most efficient barrier to the passage of microorganisms. Pilot scale studies of the chemical lime treatment process demonstrated removals of Cryptosporidium oocysts by 99% which was also observed during the full-scale process The removal mechanism for protozoa after the chemical treatment system appears to be physical removal since other experiments conducted but not presented in this research showed no decrease in viability of oocysts after exposure to high pH or high pH and disinfection combination Of all the indicators, Clostridium and coliphage best reflected the removal of enteroviruses for the chemical treatment system and the disinfection process The percentage of samples positive and the concentrations for the bacteria coliphage and protozoa were compared for the treated reclaimed water and the final effluent reservoir water receiving the effluent. In every case the 76

PAGE 89

treated water was of better quality that the ambient water in the UOSA final effluent reservoir. This study has developed an extens i ve database for the concentrations of bacteria protozoa and viruses present in the raw wastewater and the removals of these microorganisms through the various unit processes in an advanced water reclamation fac i lity The data that have been collected on microbial removals from this study provides i nformation necessary for determining the impact of water quality and potential health risks for surface water augmentation or groundwater recharge of drinking water supplies through planned indirect potable reuse This study also provides in i t i a l data for the use of alternative indicator microorganisms in addition to the conventional fecal coliform to indicate the presence of pathogens i n water. Thi s may have greater reliabil i ty in the future for ensuring the protection of drinking water supplies 77

PAGE 90

REFERENCES 1. Adams, M.H. 1959 Bacteriophages lnterscience Publishers New York 1959 2. Armon and Kott. 1996. Bacteriophages as Indicators of Pollution Critical Reviews i n Environmental Science and Technology 26(4) : 299-335. 3. Armon, R., and Payment, P. 1988 A modified m-CP medium for enumerating Clostridium perfringens from water samples Can J Microbial. 34:78-79 4 Asano, T 1995 Drinking Repurified Wastewater. Journal of Environmental Engineering 121: 548 5 Asano, T. and Mujeriego, R. 1988 Pretreatment for Wastewater Reclamation and Reuse In Pretreatment in Chemical Water and Wastewater Treatment (ed H H Hahn and R Klute) Springer-Verlag Berlin Heidelberg : 347356 6. Asano, T., Leong, L Y.C. Rigby, M.G., and Sakaji, R.H. 1992 Evaluation of the California Wastewater Reclamation Criter i a using Enteric Virus Monitoring Data Wat. Sci. Tech 26 : 1513 1524 7 Asano, T. and Levine A.D 1995 Wastewater reuse : a valuable link in water resources management. Water Quality International 4 : 20-24 8. Asano, T. and Levine, A.D. 1996. Waste-water Reclamation Recycling and ReusePast Present and Future. Water Science and Technology 33 : 1-14 9 Baker, Katherine H. and Hegarty, J.P. 1997 Detection and occurrence of i ndicator organisms and pathogens Water Environment Research 69 : 403-415. 10. Bitton, G. 1994 Wastewater Microbiology Wiley-Liss New York, 77 100. 11. Boring, J.R. Ill, Martin, W.T and Elliot, L.M. 1971 Isolat i on of Salmonella typhimurium from municipal water Riverside Calif 1965 American Journal of Epidemiology 93 : 49-54 78

PAGE 91

12 Cabelli, V.J. 1977. Clostridium perfringens as a water quality indicator In Bacteria/Indicators/Health Hazards Associated With Water ASTM STP 635 A.W. Hoadley and B J Dutka (Ed ) American Society for Testing and Materials 1977 : 247-264 13. Clausen, E.M., Green, B.L., and Litsky, W. 1977 Fecal streptococci : indicator of pollution In Bacteria/Indicators/Health Hazards Associated With Water ASTM STP 635 A.W Hoadley and B J Dutka (Ed.) American Society for Testing and Materials, 1977:247-264 14. Cohen, J., and Shuval, H.l. 1973 Water Air and Soil Pollution 2:85-95. 15 Condie, L.W., Lauer, W.C., Wolfe, G.W., Czeh, E.T., and Bums, J.M. 1994 Denver Potable Water Reuse Demonstration Project. Food and Chemical Toxicology 32 : 1021-1030 16 Craun, G. F. 1988. Surface water supplies and health. Journal American Water Works Association 80:40-52 17 Crook, J. 1994 Assessment of Water Reclamation and Reuse Research Needs Proc 1994 AWWAIWEF Water Reuse Symp Dallas Texas :371. 18 Crook, James, Asano, Takashi, and Nellor, Margaret. 1990 Groundwater Recharge with Reclaimed Water in California. Water Environment and Technology 2:42-49. 19. Crook, James and Surampalli, Rao Y. 1996 Water Reclamation and Reuse Criteria in the U S Wat. Sci. Tech 33:451-462. 20 d' Angelo, Salvatore 1996 A\!INVA and WEF Prepares Guidelines For Using Reclaimed Water to Augment Potable Water Resources. Proc 1996 AWWAIWEF Water Reuse Symp., San Diego, California : 55-58 21. Danielson, R.E., Pettegrew, L.A., Soller, J.A., Olivieri, A W., Eisenberg, D.M., and Cooper, RC. 1996 A Microbiological Comparison of a Drinking Water Supply and Recla i med Wastewater for Direct Potable Reuse Proc. 1996 AWWAIWEF Water Reuse Symp San Diego Califomia : 727-734. 22 Davies-Colley, R.J., Bell, R.J., and Donnison, A.M. 1994. Sunlight inactivation of enterococci and fecal coliforms in sewage effluent diluted in seawater Applied Environmental Microbiology 60:2049-2058 23 de Peyster, Ann, Froines, John R., Olivieri, Adam W., and Eisenberg, Don M. 1993 Aquatic Biomonitoring of Recla i med Water for Potable Use : The San Diego Health Effects Study Journal of Toxicolc>gy and Environmental Health 39:121-141 79

PAGE 92

24. Dutka, B.J. 1973. Col i forms are an inadequate index of water quality Journal of Environmental Health 36 : 39 25. Florida Department of Environmental Regulation. 1990. Reuse of Reclaimed Water and Land Application Chapter 17-610. Florida Administrative Code, Florida Department of Environmental Regulation, Tallahassee Florida 26. Fujioka, R.S. and Shizumura, L.K. 1985 Clostridium perfringens a reliable indicator of stream water quality Journal Water Pollution Control Federation 57 : 986-992. 27. Funderberg, S.W. and Sorber, C.A. 1985 Coliphages as indicators of enteric viruses in activated sludge Water Research 19:547. 28. Gagliardo, P.F., Findley, P., Richardson, T.G., and Weinberg, K. 1996 Optimization of Reclamation and Repurification at San Diego North City In Proc AWWAM/EF Water Reuse Conference San Diego California 29. Gerba, C.P., Goyal, S M., LaBelle, R.L. et al. 1979 Fa i lure of indicator bacteria to reflect the occurrence of Enteroviruses inmarine waters American Journal of Public Health 69 : 1116-1119 30. Gerba, C.P. and Rose, J.B. 1990 Viruses in source and dri nking water In Drinking Water Microbiology, (ed G A McFeters) Springer-Verlag, New York: 380-396. 31. Grabow, W O.K. 1990 Microbiology of Drinking Water Treatment: Reclaimed Wastewater In Drinking Water Microbiology (ed G A McFeters), Springer-Verlag, NewYork: 185-203 32 Grabow, W.O.K., and Isaacson, Margaretha. 1978. Microbiological Quality and Epidemiological Aspects of Reclaimed Water. Prog. Wat. Tech 10: 329-335 33. Grabow, W.O.K., Middendorff, lrmela G and Sasson, Nerine C. 1978 Role of Lime Treatment in the Removal of Bacteria Enteric Viruses and Coliphages in a Wastewater Reclamation Plant. Applied and Environmental Microbiology 35 : 663-669. 34. Grabow, W.O.K., Burger, J.S., and Nupen, E.M. 1980. Evaluation of Acid Fast Bacteria, Candida albicans Enteric Viruses and Conventiona l Indicators for Monitoring Wastewater Reclamation Systems Prog Wat. Tech. 12: 803-817 35. Grabow W.O.K., Bateman, B.W., and Burger, J.S. 1978. Microbiological Quality Indicators for Routine Monitoring of Wastewater Reclamation Systems. Prog Wat. Tech. 10:317-327. 80

PAGE 93

36 Hattingh, W.H.J., and Bourne, D.E. 1989 Research on the health implications of the use of recycled water in South Africa SAMJ 76 : 7-10. 37. Havelaar, A.H., Olphen, M.V., and Drost, Y.C. 1993 F-specific RNA bacteriophages are adequate model organisms for enteric viruses in fresh water Applied Environmental Microbiology 59 : 2956 2962 38 Hemmer, J. et al. 1994. Tampa Water Resource Recovery Project. Proc. 1994 AWWAIWEF Water Reuse Symp Dallas Texas : 557. 39. Hibler, C. and Hancock, C Waterborne Giardiasis 40. Hrudey, Steve E., Hrudey, Elizabeth J., and Shaw, Nola J. 1991 Health Effects associated with waste treatment, disposal and reuse. Research Journal WPCF 63: 437-444 41. Lauer, William C. 1991. Water 9 Quality for Potable Reuse. Water Science and Technology 23 : 2171-2180 42. Lauer, W., and Rogers, S.E. 1996 The Demonstration of Direct Potable Water Reuse : Denver's Pioneer P r oject. Proc AWWANVEF Water Reuse Conference San Diego Ca l ifornia. 43 LeChevallier, M.W., Norton, W.D., and Lee, R.G 1991 Giardia and Cryptosporidium spp in surface water supplies Applied Environmental Microbiology 57 :261 0-2616 44. LeChevallier, M.W., Norton, W.O. and Lee, R.G 1991. Giardia and Cryptosporidium spp in Filtered Drinking Water Supplies Applied Environmental Microbiology 57 : 2617-2621 45. Lisle, J.T. and Rose, J B 1995 Cryptosporidium contamination of water in the USA and UK: a mini-review. J. Water SRTAqua 44 : 103-117. 46. MacKenzie, William R., Hoxie, Neil J Proctor, Mary E., Gradus, M. Stephen, Blair, Kathleen A., Peterson, Dan E., Kazmierczak, James J Addiss, David G., Fox, Kim R., Rose, Joan B., and Davis, Jeffrey P 1994 A massive outbreak in Milwaukee of Cryptosporidium infection transmitted through the public water supply The New England Journal of Medicine 331(3) : 161-167 47 Madore, M.S., Rose, J.B., Gerba, C.P., Arrowood, M J and Sterling, C.R 1987 Occurrence of Cryptosporidium oocysts in sewage effluents and selected surface waters Journal of Paras i tology 73: 702-705. 81

PAGE 94

48. Marshall, Marilyn M., Naumovitz Donna, Ortega, Ynes, and Sterling, Charles R. 1997 Waterborne Protozoan Pathogens Clin Microbial. Rev. 10: 68-85 49. McEwen, Brock and Richardson, Tom. 1996. Indirect Potable Reuse : Committee Report Proc. 1996 AWWAM/EF Water Reuse Symp., San Diego California : 485-503. 50. Metcalf, T.G. 1978. Indicators for viruses in natural waters In Water Pollution Microbiology 2 (ed. R. Mitchell), Wiley-lnterscience New York : 301-325 51. Neller, Margaret H., Baird, Rodger B., and Smyth, John R. 1985. Health Effects of Indirect Potable Water Reuse Journal AWWA : 88-96 52 Olivieri, A.W., Eisenberg, D.M., Cooper, R.C., Tchobanoglous, G., Gagliardo, P. 1996 Recycled Water a Source of Potable Water : City of San Diego Health Effects Study Water Science and Technology 33 : 285-296 53. Payment, P. and Franco, E. 1993. Clostridium perfringens and somatic coli phages as indicators of the efficiency of dri nking water treatment for viruses and protozoan cysts. Applied Environmental Microbiology 59:2418-2424. 54. Payment, Pierre, and Armon, Robert. 1989. Virus Removal by Drinking Water Treatment Processes Critical Reviews in Environmental Contro l19:15-31. 55 Pia, Michelle M., Grebbien, Virginia, and Gaston, John M. 1996 Potable Reuse and the Emerging Conflicts with Drinking Water Regulations 1996 AWWANVEF Water Reuse Symp., San Diego Califomia : 715-721. 56. Robbins, M.H.1993 Supplementing a surface water supply with recla i med water. In Proceedings of the AVVWA Annual Conference and Exposition June 610, San Antonio TX, AWWA, Denver CO. 57. Rodgers, Mark R., Flanigan, Debbie J. and Jakubowski, Walter. 1995 Identification of Algae Which Interfere with the Detection of Giardia Cysts and Cryptosporidium Oocysts and a Method for Alleviating This Interference Applied and Environmental Microbiology 61: 3759-3763 58 Rose, J.B. 1988 Occurrence and Significance of Cryptosporidium in water Journal Amer i can Water Works Association 80 : 53-58. 59. Rose Joan B., Landeen, LeeK., Riley Kelley R., and Gerba, Charles P. 1989 Evaluation of Immunofluorescence Techniques for Detection of Cryptosporidium Oocysts and Giardia Cysts from Environmental Samples Applied and Environmental Microbiology 55:3189-3196. 82

PAGE 95

60 Rose, J.B., Gerba, C.P., and Jakubowski, Walter. 1991. Survey of Potable Water Supplies for Cryptosporidium and Giardia. Environmental Science and Technology 25 : 1393-1400 61. Rose, Joan B., Robbins, Millard, Friedman, Debra, Riley, Kelley, Farrah, Samuel R., and Hamann, Carl L. 1996 Evaluation of Microbiological Ban iers at the Upper Occoquan Sewage Authority 1996 AWWA/WEF Water Reuse Symp., San Diego, California : 291-305. 62. Rose, Joan B., Dickson, Linda J Farrah, Samuel R., and Carnahan, Robert P. 1996. Removal of Pathogenic and Indicator Microorganisms by a Full Scale Water Reclamation Facility. Water Research 30 : 2785-2797 63. Seligmann, R. and Reitler, R. 1965 Enteropathogens in water with low Escherichia coli titer. Applied Environmental Microbiology 57 : 1572 157 4. 64. Sinton, L.W., Davies-Colley, R.J., and Bell, R.G. 1994 Inactivation of enterococci and fecal coliforms from sewage and meatworks effluents in seawater chambers. Applied Environmental Microbiology 60:2040-2048 65. Smith, Robert G. 1995. Water reclamation and reuse Water Environment Research 67:488-495 66. Snowdon, Jill A. and Cliver, Dean 0. 1989 Coliphages as Indicators of Human Enteric Viruses in Groundwater. Critical Reviews in Environmental Control 19: 231-249 67. Standard Methods for the Examination of Water and Wastewater. 1992 APHA AWWA, WEF, 18th Edition, Washington, D C 68. State of Arizona. 1991. Regulations for the Reuse of Wastewater. Arizona Administrative Code Chapter 9, Article 7, Arizona Department of Environmental Quality, Phoenix, Arizona. 69. State of California. 1978. Wastewater Reclamation Criteria. California Administrative Code, Title 22, Division 4 California Department of Health Services, Sanitary Engineering Section Berkeley, Ca l ifornia 70. Stander, G.J., and Clayton, A.J. 1977 Planning and construction of wastewater reclamation schemes as an integral part of water supply. In Water; Wastes and Health in Hot Climates, (ed. R Feachem M. McGarry and D Mara) Wiley, London : 383-391. 71. U.S. EPA 1992. Guidelines for Water Reuse U S Environmental Protection Agency, Center for Environmental Research Information, Cincinnati Ohio. 83

PAGE 96

72. USEPA, Monitoring Requirements for Public Drinking Water Supplies ; Proposed Rule, Federal Register, 59(28) : 6332-6429 1994 56. USEPA, Monitoring Requirements for Public Drinking Water Supplies ; Proposed Rule Federal Register 59(28) : 6430-6440 1994 73. van Leeuwen, J. (Hans). 1996 Recla i med water-an untapped resource Desalination 106:233-240 7 4 Van Riper, C. and Geselbracht, J. 1996 Water reclamation and reuse Water Environment Research 68 : 516-520 75. Yanko, William A. 1993 Analysis of 10 years of virus monitoring data from Los Angeles County treatment plants meeting California wastewater reclamation criteria. Water Environ. Res 65 : 221-226 84

PAGE 97

APPENDICES 85

PAGE 98

APPENDIX A 86

PAGE 99

Table Al 4/28/95 SAMPLE ID SITE DESCRIPTION 001 I UNTREATED WASTEWATER I 011 SECONDARY EFFLUENT 020 S ECOND STAGE RECARB. 017 MULTIMEDIA. INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT oJj FINAL EFF POSTDECIIL. 060 FINAL EFFL. RESERVOIR NA -Sample was not analyzed. co --...! ENTEROCOCCI 6. 5 X 10' 1 0 X 10 3 0 < 10.0 < 10 0 1.0 < 0 5 < 1.0 TOTAL/FECAL PHAGE COLIFORMS 11/100 ll/ 100mL m t. NA 6.5 X 10' NA. 200 NA. < 12.5 NA. < 12 5 NA. < 12 5 NA. < 12 5 NA < 12 5 NA < 14 ) ENTEROVIRUS CLOSTRIDIUM CRYPTOSPORIDIUM GIARDIA MPN/100 L ll/ 100mL II/100L II/100L 690 0 NA < 1123 22 4 6 14 0 NA. < 33.9 67 9 < 0 .37 NA. 2 1 4 22 < 0 .10 NA < 0 .74 < 0.74 < 0 .069 NA < 0.42 < 0 42 < 0.077 NA 2.82 < 0 94 < 0 061 NA 0 44 0 44 < 0 .14 NA < 1.95 1. 9 5

PAGE 100

Table A2 5/24/95 SAMPLE 10 SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF POSTDECHL. 060 FINAL EFFL. RESERVOIR Plate too numerous to count (a)Problem with analysis 00 00 ENTEROCOCCI ll/ 100mL 5 0 X 104 400 1.5 5.5 0.5 < 0.5 < 0 5 < 0 5 TOTAL/FECAL PHAGE COLI FORMS lll100mL 111100 mL I 3. 2 X 105 1 8 X 10' I 1 2 X 101 l 5 X 101 15 0 I 3 0 < 12 5 92 0 I 57.0 25 0 52 0 I 11.0 < 1 2 5 2.0 I (a) < 12 5 4 .01<1.0 < 12.5 < 1 0 I 19.0 < 12.5 ENTEROVIRUS CLOSTRIDIUM MPN/100 L lll 100mL 2 4 X 10' 3 1 < 0 12 2 0 < 0 067 2 5 < 0 045 0 5 < 0 11 0.5 < 0 11 < 0.5 < 0 21 < 0 5 CRYPTOSPORIDIUM III100L 277.8 < 1 9 8 < 3 9 6.4 0.9 < 0 9 < 2 1 < 3.7 GIARDIA II/100L 22500 59 5 < 3 9 < 6 4 < 0.9 < 0 9 < 2 1 7 5 'd (1) :::1 p.. t-' X ,...... (") 0 :::1 rt t-' :::1 (1) p.. ..._,

PAGE 101

Table A3 6/27/95 SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 C? \.() MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF POSTDECHL. 060 FINAL EFFL RESERVOIR Problems with analysis. ENTEROCOCCI 11/lOOmL 7 1 X 10' 4 5 X 101 88 5 15. 5 3 0 < 0.5 16.0 TOTAL/FECAL PHAGE COLIFORMS 11/100 l#/100mL mL 2. 4 X 10' I 8.6 X 105 2. 5 X 101 1.5 X 10'/ 1.6 X 101 3 8 X 101 7 o I lal < 12.5 114.0 I 118.0 < 12.5 41. o I 29 o < 12 5 37.o I 9.o < 12.5 1.0/cl.O < 12.5 I < 12.5 -ENTEROVIRUS CLOSTRIDIUM MPN/100L 11/lOOmL 1 0 X 101 3 0 X 104 1.2 X 101 4 5 X 101 < 0.012 < 0 5 < 0.035 4 0 0 .037 < 0 5 0.120 < 0 5 < 0.069 < 0.5 < 0.14 < o s CRYPTOSPORIOIUM II/100L < 1. 15 X 101 < 24.5 < 2.6 < 5 0 < 10.3 < 4 7 < 4.6 5.7 GIARDIA II/100L 1.22 X 10' 24.5 2 6 < 5.0 < 10.3 < 4 7 < 4 6 < 5.7 > 't:l 't:l (I) :::1 r:L ..... :< > ....... () 0 :::1 rt ..... g (I) 0.. -......;

PAGE 102

Table A4 7/24/95 I SAMPLE ID I SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECHL. 060 FINAL EFFL. RESERVOIR ENTEROCOCCI ll/100mL 7 5 X 10' 600. 0 6.0 92 0 6 0 0 5 4 5 0 5 o17 Echovirus 11 ; 023 -Echovirus 11 b Too numerous to count \.0 0 TOTAL/FECAL PHAGE COLIFORMS 11/100 ll/ 100mL mL 2. 2 X 10' I 5.5 X 101 3 6 X 10 2 4 X 10'/ 5.5 X 10' l 2 X 1 01 6.0 I 2 0 < 12 5 b I 202 0 < 16 7 17 0 I 8 o < 12 5 8 0 I 8 0 < 25 0 <1.0 I <1.0 < 12.5 b I 3 o < 12. 5 ENTEROVIRUS CLOSTRIDIUM MPN/100 L ll/100mL 260 0 4 5 x 1 o 4.32 1.4 X 10' < 0.097 1.0 07 2.5 0.043 2.5 0 068. 6 0 < 0.047 3 5 < 0.23 10.0 CRYPTOSPORIDIUM II/100L < 603.8 < 12 4 < 5.1 < 5 0 < 4 9 < 4 6 < 1.2 < 11 3 GIARDI A II/ 1 00 L 5 1 X 10' 37.2 5 1 < 5. 0 < 4.9 < 4 6 < 1.2 < 11.3 '"0 (I) l:l 0.. > ,....._ (') 0 l:l rt ..... g (I) 0.. .......-

PAGE 103

Table AS 8/30/95 SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECHL. 060 FINAL EFFL RESERVOIR Too numerous to count "Not determined <.0 ...... ENTEROCOCCI lll100mL 8 2 X 10' 2.7 X 10' 11.5 130.0 15.5 5.0 1.0 4.0 TOTAL/FECAL PHAGE CO!, I F ORMS llllOOOIL 111100 mL 4 5 X 10' I 6 3 X 10' 4 1 X 104 1 5 X 10'1 2 8 X 10' 1 0 X 1 04 10. 5 I 3.5 < 12.5 44o.o I 275.0 12.5 6o. o I 3o.o 12.5 55. 0 I 17. 5 < 12. 5 <0.5 I < 0 5 < 12. 5 I 4 5 < 12.5 ENTEROVIRUS C LOSTRIDIUM MPNI100 L llllOOmL 1. 3 X 101 b 47. 0 < 0.103 < 0 .081 b < 0 .043 < 0 .056 < 0 .055 < 0.39 CRYPTOSPORIDIUM III100L <2959 < 375.8 < 11. 0 < 4.6 < 1.0 < 4.6 < 2 0 < 42.4 GIARDIA III100L 44386 375. 8 < 11. 0 < 4 6 < 1 0 < 4 6 < 2.0 <42. 4 .g-'d ro ::1 p.. ,..-... (") 0 :::J rt t::l s:: ro p.. ..._,

PAGE 104

Table A6 9/26/95 SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND SThGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECHL. 060 FINAL EFFL. RESERVOIR 0 20 Coxsackie 85 <..0 N ENTEROCOCCI ll/100mL 8 7 X lO s 850. 0 4 5 5 370. 0 17 0 16 5 < 0.5 6.5 TOTAL/FECAL PHAGE COLIFORMS 11/100 11/lOOmL mL 3 3 X 101 / 5 9 X lOs 2 9 X 10' 3 7 X 10'/ 2 8 X 10' 7 6 X 10' 160. 0 I 10.0 12 5 825 0 I 480.0 75 0 10. 0 I 27 5 < 12 5 46 5 I 11 0 < 12 5 <0 5 I <0.5 < 12.5 245. 0 I 16.5 12 5 ENTEROVIRUS CLOSTRIDIUM MPN/100L 11/lOOmL 1 2 X 10' 5, 7 X 104 57. 0 1 1 X 10' 0 22 7.0 0.18 5 0 < 0 12 0 5 < 0 09 < 0 5 < 0 09 < 0.5 < 0.23 2.5 CRYPTOSPORIDIUM II/100L 2690 < 79 1 4.8 < 12.5 < 1.0 < 0.9 < 4 9 < 10 6 GIARDIA II/100L 3 9 X 104 79.1 4 8 < 12.5 < l. 0 < 0 9 < 4.9 < 10.6 'd ::1 0. X > ......... (') 0 ::s rt g 0. ........

PAGE 105

\.() w Table A7 10/24/95 SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER I 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFI.UENT 033 FINAL EFF. POSTDECHL. I 060 FINAL EFFL. RESERVOIR ENTEROCOCCI ll/100mL 8 7 X lOs 1 4 X 101 17 5 23.5 1.0 3 0 < 0.5 61.0 TOTAL/FECAL PHAGE COLIFORMS 11/100 ll/100m L mL 2. 8 X 10' I 5.5 X lOs 5 7 X 10' 2 4 X lOS I 1.3 X 101 4 1 X 10' 155 I 57. 5 < 12 5 335 I 49 5 12 5 20.0 I 9.5 < 12 5 17 5 I 4 0 < 12 5 o 5 I < o 5 < 12 5 6oo. o I 55.0 J7. 5 ENTEROVIRUS CLOSTRIDIUM MPN/100 L 11/lOOmL 150 0 1 6 X 10' 1.4 833 0 < 0 14 0 5 < 0 .086 < 0.5 < 0 .051 < 0 5 < 0.066 < 0 5 < 0.061 < 0.5 < 0.15 0 5 CRYPTOSPORIDIUM II/100L < 1. 4 X 101 < 66.2 < 1. 1 < 20 2 < 1. 1 < 0.9 < 2 3 < 22 5 GIARDIA II/100L 9 95 X 10' < 66.2 < 1 1 < 20.2 < 1 1 < 0 9 < 2 3 < 22.5 .6" 'd (IJ ::l 0.. ...... X > -(') 0 ::l rt ...... ::;1 (:::: 11> 0..

PAGE 106

Table AS 11/28/95 1..0 .:-' SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECHL. 060 FINAL EFFL. RESERVOIR ENTEROCOCCI ll/100mL 2 6 X 10' 233.j 4 0 14.5 < 0 5 < 0.5 < 0 5 < 0.5 TOTAL/FECAL PHAGE COLIFORMS 11/100 ll/100mL mL 3. 5 X 10' I 1 4 X 1il5 3 5 X 10' 6 3 x 1 o 1 2. 0 X 101 1.6 X 101 58. 5 I 4 0 75. 0 3oo.o I 52.5 175. 0 43. 0 I 10.5 75.0 25. 0 I 1 0 25. 0 1.0 I < o 5 < 12. 5 4o.o I 11.0 50. 0 ENTEROVIRUS CLOSTRIDIUM MPN/100 L ll/100mL 320.0 3. 2 X 10' 2.5 311.0 < 0.14 3 5 < 0 .069 11. 0 < 0.063 < 0.5 < 0 .062 < 0.5 < 0.049 < 0.5 < 0 .24 4 5 CRYPTOSPORIDIUM II/100L < 4 5 X 101 < 63.3 < 10.9 < 9 6 < 10.1 < 27. 5 < 5.7 < 120. 0 GIARDIA II/100L 6. 3 X 10' < 63.3 < 10.9 < 9.6 < 10.1 < 27. 5 < 5.7 < 120. 0 .fl '0 ro ::l p.. 1-' !I> .--.. n 0 ::l rt 1-' ::l ro p.. '-'

PAGE 107

Table A9 1/23/96 SAMPLE ID SITE DESCRIPTION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF POSTDECHL. OVERFLOW 'MPN method u sed t..<.) V1 ENTEROCOCCI ll/100mL 3.1 X 10' 2. 7 X 101 80.0 325 0 45. 5 59. 0 < 0 5 350.0 TOTAL/FECAL PHAGE COLIFORMS 11/100 ll/100mL mL' > 1 6 X 10' I 6. 4 X 10' 9.0 X 10' 9 0 X 10' I 2 1 X 101 5 0 X 101 BOO I 130 12 5 > 1 6 X 10' I 790.0 5. 0 X 10' >1.6xlO'I 12 5 240 > 1 6 X 10' I < 12. 5 130 <1.11<1.1 < 12.5 eo.o I 9.0 25 0 ENTEROVIRUS CLOSTRIDIUM MPN/100 L ll/100mL 187.0 1. 5 X 10' 1.5 888. 9 < 0.39 9 0 0.66 95.0 0 .27 6 0 0 .18 2.5 < 0.13 < 0 5 0 39 27.0 CRYPTOSPORIDIUM II/100L <1.1x101 < 354 7 < 31.6 < 10.9 < 5 3 < 25.5 < 29.9 < 10. 4 GIARDIA II/100L 6. 5 X 10' < 354. 7 727. 8 326.9 < 5 3 < 25 5 < 28.9 239.2 "d 10 1:1 p.. r' :>< > r-.. (") 0 ::I rt r' g 10 p.. '-'

PAGE 108

Table AlO 2/6/96 I \\) 0\ SAMPLE ID SIT E DESCRIPTION 001 UNTRE A T E D WAS T EWATER 011 SECONDARY EFFLUENT 0 20 SECOND STAGE RECARB. 0 1 7 MULT I M E DIA INFLUENT 0 2 1 MULTI-MEDIA EFFLUENT 02 3 GAC E FFLU ENT 033 FINAL E FF. P O S TOECHL. 060 F INAL EFFL RESERVOIR ENTEROCOCCI ll/100mL 1 6 X 1 05 288. 9 < 0 5 3 0 < 0.5 0 5 < 0 5 1 5 TOTAL/FECAL PHAGE COL I FORM S 11/ 100 #/100mL m L 3.2x10'/ 6.4 X 10' 1 3 X 10 5 8 X 10'1 4 00 1.3 X 10' o 5 I < 0 5 < 12.5 9 6 5 I 10 0 12 5 38.0 I 2 5 < 25.0 20.5 I 2 0 < 25 0 < o 5 I< o 5 < 1 2 5 15 o o I n.o 2 5 0 --E NTEROV IRUS CLOSTRIDIUM MPN/1 00 L ll/100mL 4 6 X 1 03 2 1 X 10' 6 9 338 9 < 0 15 1.5 < 0 14 5 0 < 0 1 5 0 5 < 0.12 < 0 5 < 0.13 < 0 5 < 0 9 1 9 0 -------C R Y PTOSPORIOIUM GIARDI A #/100L #/100L < 2 4 X 103 9 6 X 10' < 155. 3 < 15 5 3 < 25.1 275 7 56 3 112 7 <' 1 0 0 < 10. 0 < 29.6 < 2 9 6 < 25 1 < 2 5 1 < 9 5 5 < 9 5 5 .6" "d (1) :::1 A. > ........ (") 0 ::s rt .... ::s c (1) p.. ._,

PAGE 109

Table All 3/5/96 I I \...0 -.....! SAMPLE ID SITE DESCRIP TION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECARB 01 7 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECIIL. 060 FINAL EFFL RESERVOIR ENTEROCOCCI ll/100mL 5.9 X 10' l.1X 10' 54 5 62.0 26 0 19.0 < 0 5 14 0 TOTAL/FECAL PHAGE COLIFORMS 11/100 11/lOOmL mL 4.2x10'/ 7.2 X 104 1.2 X 1 01 6 1 X 10'/ 1. 3 X 10' 9.1 X 10' 11 5 I o.5 12.5 20.0 I 1.0 37. 5 11 0 I 2.0 < 12 5 6.0 I o.5 < 12.5 < o 5 I < o s < 1 2 5 11.0 I 6 5 25.0 ENTEROVIRUS CLOSTRIDIUM CRYPTOSPORIDIUM MPN/100 L ll/100mL II/100L 490.0 2.6 X 104 < 9508 7 21.0 2. 8 X 10' < 2524.4 < 0.20 12 0 < 11. 0 < 0.24 1 2 5 < 38.4 < 0 09 1.5 < 10.2 < 0 14 1.5 2 6 < 0 14 < 0 5 < 12 8 < 0 25 12.5 < 92 2 GIARDIA II/100L 14263 1 17671 < 11.0 < 38. 4 61.3 < l. 3 12 8 < 92 2 'd 0. > ,....._ (') 0 ::I rt t-' g (l) 0. '-"'

PAGE 110

Table A12 4/9/96 \,!) 00 SAMPLE ID SITE DESCRIPT ION 001 UNTREATED WASTEWATER 011 SECONDARY EFFLUENT 020 SECOND STAGE RECAAB. 017 MULTI-MEDIA INFLUENT 021 MULTI-MEDIA EFFLUENT 023 GAC EFFLUENT 033 FINAL EFF. POSTDECHL. 060 FINAL EFFL RESERVOIR ----------ENTEROCOCCI TOTAL/FECAL PHAGE ll/100mL COLIFORMS 111100 lll 100mL mL 2 8 X lOs 1 6 x10'/ 6.0 X 10' 1.8 X 10' 972.2 2 8 X 650. 0 3 3 X 101 8 0 14.5/1.5 < 12 5 10 0 3o.o I 5.o < 12 5 < 0 5 5 5 I < < 12 5 0 5 0 5 8 0 I < 0 5 < 12.5 < 0 5 < 0 5 I < 0.5 < 12.5 5 5 30 o I 9 5 < 12 5 ENTEROVIRUS CLOSTRIDIUM CRYPTOSPORIDIUM MPN/100 L ll/100mL II/100L 4 4 X 10' 5 3 X 10' < 1 3 X 101 4 9 2 7 X 10' < 61.3 < 0 .17 9 5 < 22 3 < 0.13 11 0 < 65.2 < 0 066 1 0 < 11.2 < 0.074 0 5 < 9.2 < 0 082 < 0 5 < 12 8 < 0 19 4 5 < 61 7 GIARDIA II/100L 1 3 X 10' 61 3 < 22 3 < 65.2 < 11.2 < 9 2 < 12 8 < 61 7 I i .&" '1:l II) ::l p. t-' > ,....... (') 0 ::l t-' ::l c II) p. '-"'

PAGE 111

APPENDIX B 99

PAGE 112

Experiment 1 9/26/95 3gpm TIME (sec) i SAMPLE : Beads/mL i MS2 phage/mL 60 I 1-1 4 60E+03 I 1.10E+08 I 120 j 1-2 ; 3.98E+04 I 5.70E+08 150 1-3 I 2 .11E+04 I 7 80E+08 180 : 1-4 ; 3 78E+04 I 6.30E+08 210 i 1-5 : 8.20E+03 I 1.50E+08 235 i 1-6 : 4 00E+02 I 5 90E+07 1 270 i 1-7 4 40E+02 8.80E+06 295 1-8 ; 1 12E+02 1 80E+06 320 : l-9 5.50E+01 8 60E+05 345 1-10 5 20E+01 I 4.80E+05 Omin E-1 < 1 0 3 10E+01 30 min E-2 3 60E+01 I 1 00E+01 60min E-3 : 4 30E+01 I 5 40E+02 95 min : E-4 9.00E+OO i 1.10E+01 125 min I E-5 1 .20E+01 1.10E+01 155 min E-6 1 20E+01 I 1 1 0E+01 185 min E-7 2 .00E+OO l 1 40E+01 210 min : E-8 1.00E+OO I 8 60E+01 Background I < 1 0 1 < 1 0 I Seed ; 1 44E+04 I 3 00E+09 PRD1 phage/ml 3 00E+06 5 .00E+ 07 9 90E+07 1 60E+08 2 90E+07 4 50E+06 1.20E+06 3 30E+05 2.30E+05 1 10E+05 1 50E+03 > 10. 0 > 10. 0 : > 10. 0 > 10. 0 > 10. 0 > 10 0 > 10 0 < 1 0 4 40E+08 100

PAGE 113

Appendix B (Continued) TIME (sec) 30 50 75 110 135 160 200 235 270 310 Omin 30min 65 min 95min 125 min 155 min Background : Seed Experiment 2-9/26/95 7gpm SAMPLE : Beads/ml : MS2 phage/mL 1-1 2 70E+03 1.50E+07 1 2 1.08E+04 : 2 50E+08 ; 1-3 6 40E+03 i 2 50E+08 ; 1-4 1 80E+03 l 2.40E+07 1-5 : 6 40E+03 : 1 50E+08 1-6 1 00E+04 I 1.80E+08 1-7 4.40E+03 : 5.70E+07 1-8 1.82E+02 ; 4.70E+06 1-9 3 20E+01 6.20E+05 1-10 2 .00E+OO : 1 90E+05 E-1 1.00E+OO ; < 1 0 E-2 1 70E+01 < 1 0 I E-3 2 .00E+OO I < 1 0 E-4 < 1 0 < 1 0 E-5 < 1 0 < 1.0 I I E-6 < 1 0 < 1.0 i < 1.0 1 20E+01 I I 2.41E+04 : 2 90E+09 i I PRD1 phage/mL 3 00E+08 7.10E+06 1.20E+08 5 90E+07 3 40E+06 8 40E+07 2.00E+07 1 10E+06 1.30E+05 4 20E+04 1 10E+02 7 90E+01 < 1 0 < 1 0 < 1 0 < 1.0 < 1 0 4 40E+08 101

PAGE 114

Appendix B (Continued) Experiment 1 3/5/96 7gpm TIME (sec) SAMPLE i Oocysts/ml. MS2 phage/ml ; 0 1-1 : I 1.00E+06 30 ; 1-2 I I 2 80E+08 60 I 1 3 ; I 2 90E+08 i 90 i 1-4 i I 3 .20E+08 i I 120 1-5 i 2 90E+08 I I 150 i 1-6 I 6.50E+07 I 180 j 1-7 i I 4 .00E+06 I 210 I 1-8 i j 1 .90E+07 I 270 ; 1-10 6.55E+02 I I 300 : 1-11 3 .60E+02 I I 330 i 1-12 4 .45E+02 i i 390 i 1-13 : 1.90E+01 I 420 i 1-14 1.00E+OO i i 1 min I E-1 < 0.01 < 1.0 I i Smin E-2 i < 0 .01 I 1.00E+OO I 10 min i E-3 ; < 0.01 < 1.0 15 min E-4 i < 0 .01 I 1.00E+OO 20 min E-5 < 0 .01 I 2 20E+01 I 25 min I E-6 0 .02 5.00E-01 I : 30min : E-7 I 1.20E-01 < 1 0 i 60 min I E-8 : I 0.22 i 2.50E+OO I Background-! 1 3.50E+OO I I Background-E : I 3.00E+OO I Seed I i 5.40E+09 PRD1 phage/ml 1.00E+06 2 30E+07 1 80E+07 2 20E+07 1 40E+07 2 80E+06 2 20E+05 1 30E+06 2 00E+01 1 20E+01 1.40E+01 2.30E+02 4 80E+03 3 10E+04 1 00E+05 8 30E+04 < 1.0 < 1 0 2 60E+08 1.02

PAGE 115

Appendix B (Continued) Experiment 2 3/5/96 3gpm TIME (se c) SAMPLE : Oocysts/mL MS2 phage/mL 0 i 1-1 : 1 00E+06 30 I 1-2 ; i 3 50E+08 60 I 1-3 I 3.80E+08 90 1-4 I i 3 70E+08 120 1-5 I I 2.70E+08 150 1-6 I I 1.60E+08 180 1-7 : I 5.30E+06 I 210 1-8 I I 3.20E+06 240 I 1-9 5 85E+02 : 270 1-10 8 60E+02 300 !-11 i 8 20E+02 330 1-12 6.05E+02 ; 360 i 1-13 2 06E+02 390 I 1-14 3 .00E+OO o min E-1 l < 0 .01 I S.OOE-01 10 min ; E-2 : < 0 .01 ; 1.00E+OO 20 min E-3 i 1 00E-02 ; 2.00E+OO 30 min E-4 1.20E-01 I 1.00E+OO 40 min : E-5 0.41 I 1 .50E+OO I 50 min E-6 ; S .OOE-01 1 .50E+OO 60 min E-7 I 0 45 < 1 0 90 min E-8 i 0 .27 l S.OOE-01 120 min E-9 0 15 1.00E+OO 150 min E-10 0 .11 I 1 .00E+OO Background I I i 7 30E+01 Background-E : 2.50E+OO Seed I 4 10E+03 2 20E+08 PRD1 phage/mL 1.00E+06 1.40E+08 1.30E+08 1 50E+08 1 .50E+08 : 6 .00E+07 3 .30E+06 1 .50E+06 : : 5 .50E+OO : : 1 00E+01 9.30E+03 > 100. 0 2 .20E+05 2 .60E+05 i 2.30E+05 1 .80E+05 1 .50E+05 1 .10E+05 3 50E+01 1 70E+01 1.20E+08 103

PAGE 116

APPENDIX C 104

PAGE 117

..... 0 VI Bacter i a and Protozoa H it s afte r Second Stage Recar b o n at i on ( 020 ) 1 .00E+ 02 ..----------------. -l 1 .00E+01 E 0 0 T::::> u. 0 1 .00E+00 I / \ / \ / .... \ / ... I I r \. / \ I :' : v . . . .. I f ..... ) 1 ,000 1 00 10 1 1 .00E-01' It H I It Ji H H j.j *0.1 A P R M A Y JUN JUL AUG SEP OCT NOV JAN FEB MAR APR 1995 1 1996 _J 0 0 TC/) ..... C/) >-(.) 0 0 "0 c: ro C/) ...... C/) >-(.) ... E ntero c o c ci ... C perfring en s "*" Crypt ospo r id ium ,. Giardia

PAGE 118

,_. 0 (J'\ Bacteria and Viral Hits after Second Stage Recarbonation (020) _J 1 .00E+01 E 0 0 or:::::> LL 0 1.00E+00 \ \ \ 1 I tl / I I 'I I I 10 1 .00E-01' i i APR MAY JUN JUL AUG SEP OCT NOV JAN FEB MAR APR 1995 1 1996 _J 0 0 or:::::> LL a. Enterococci ... C perfringens _,._Coliphage ..... Enterovirus :x> "0 "0 (I) ::I p. 1-' :< 0 r-.. n 0 ::I rt 1-' ::I c (I) p. ......-

PAGE 119

..... 0 -...J Bacteria and Protozoa Hits in the F i nal Effluent (033) 1.00E+01 100 __J 0 l 0 10 __J I\ en E +"" en 0 I \ >. 0 (.) 1.00E+00 I 0 0 ::J I \ lL '0 0 I \ c: 1 I \ en +"" I en >. I (.) I I 1.00E-01' I I o. 1 APR MAY JUN JUL AUG SEP OCT NOV JAN FEB MA R APR 1995 1 1996 -Enterococci C. perfringens Cryptosporidium ...... Giardia !l> '0 '0 (l) p.. ..... >: (") ----n 0 rt f-' c (l) p..