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Microbial diversity in compost, its efficacy against pathogenic fungus Mycena citricolor, and soil erosion at Life Monteverde

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Title:
Microbial diversity in compost, its efficacy against pathogenic fungus Mycena citricolor, and soil erosion at Life Monteverde
Translated Title:
La diversidad microbiana en compostaje, su eficacia contra el hongo patógeno Mycena citricolor, y la erosión del suelo en la Finca Life Monteverde
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Hood, Naomi
Thurmond, Mason
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Coffee--Diseases and pests ( lcsh )
Café-Enfermedades y plagas ( lcsh )
Coffee plantations ( lcsh )
Plantaciones de café ( lcsh )
EAP Spring 2017
EAP Primavera 2017
Costa Rica--Puntarenas--Monteverde Zone
Costa Rica--Puntarenas--Zona de Monteverde
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Reports

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Abstract:
The incidence of Mycena citricolor on coffee plants and soil erosion threaten the productivity of coffee farms in the Monteverde region. Mycena citricolor, commonly known as ojo de gallo, is a pathogenic fungus that causes defoliation and loss of fruit in Coffea arabica. The fungus lives on coffee leaves and in the soil. As soil erodes, fertile material for plant growth is lost and ojo de gallo is given the opportunity to spread across the farm. This study investigated the possibility of novel sustainable practices in maintaining healthy coffee plants at Finca Life Monteverde by applying compost to ojo de gallo. We also compared soil erosion in areas of differing vegetation cover and found that there was not a significant relationship between percent vegetation cover and soil erosion. We compared the inhibition and reduction in number of colonies of Mycena citricolor after application of various composts produced by the farm, a synthetic fungicide called Opus, and the commonly used biocontrol called Trichoderma asperellum (Trichoderma). We found that a compost containing rice and coffee skins, goat and chicken manure, whey, molasses, and carbon produced by the farm significantly inhibited growth of Mycena citricolor the most, and all composts inhibited more so than the synthetic fungicide Opus. Composts have the potential to effectively inhibit the growth of ojo de gallo because their microbial diversity can outcompete the pathogenic fungus. The use of vegetation cover as a means of preventing soil erosion should be further investigated to limit the spread of pathogenic fungi. ( , )
Abstract:
La incidencia de Mycena citricolor en las plantas de café, así como la erosión del suelo, amenazan la productividad de las fincas cafetaleras de la región de Monteverde. Mycena citricolor, comúnmente conocido como ojo de gallo, es un hongo patógeno que causa defoliación y pérdida de fruta en Coffea arabica. El hongo vive en las hojas de café y en el suelo. A medida que el suelo erosiona, el material fértil para el crecimiento de las plantas se pierde y ojo de gallo se expande a lo largo de la finca. El presente estudio investigó el potencial de ciertas prácticas sostenibles para mantener plantas de café saludables en la Finca Life Monteverde, aplicando diferentes compostas al ojo de gallo. Comparamos además la erosión del suelo en áreas de cobertura vegetal diferente, y no encontramos una relación significativa entre el porcentaje de cobertura vegetal y la erosión del suelo. Asimismo, comparamos inhibición y reducción del número de colonias de Mycena ci-tricolor después de la aplicación de diversos compost producidos por la finca, también de un fungicida sintético llamado Opus, y el biocontrol de uso común Trichoderma asperellum (Tricho-derma). El compost que más inhibió el crecimiento del hongo fue el que contenía broza de café, estiércol de cabra, gallinaza, suero de leche, melaza y carbón producido por la finca. Todos los compostas inhibieron más que el fungicida sintético Opus. Los compostas tienen el potencial de inhibir efectivamente el crecimiento de ojo de gallo porque su diversidad microbiana puede competir con el hongo patógeno. El uso de la cobertura vegetal como medio para prevenir la erosión del suelo debe investigarse más a fondo para limitar la propagación de este hongo.
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Student affiliation: Ecology, Behavior, Evolution and Public Health, University of California, San Diego; Environmental Systems, University of California, San Diego.

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Monteverde Institute
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Monteverde Institute
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This item is licensed with the Creative Commons Attribution Non-Commercial No Derivative License. This license allows others to download this work and share them with others as long as they mention the author and link back to the author, but they can’t change them in any way or use them commercially.
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M39-00628 ( USFLDC DOI )
m39.628 ( USFLDC Handle )

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Tropical Ecology Collection [Monteverde Institute]

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Microbial diversity in compost, its efficacy against pathogenic fungus Mycena citricolor and soil erosion at Life Monteverde Naomi Hood Mason Thurmond University of California, San Diego University of California, San Diego Ecology, Behavior and Evolution and Public Health Environmental Systems EAP Tropical Biology and Conservation Program, Spring 2017 9 June 2016 ______________________________________________________________________________ ABSTRACT The incidence of Mycena citricolor on coffee plants and soil erosion threaten the productivity of coffee farms in the Monteverde region. Mycena citricolor commonly known as ojo de gallo, is a pathogenic fungus that causes defoliation and loss of fruit in Coffea arabica The fungus lives on coffee leaves and in the soil. As soil erodes, fertile material for plant growth is lost and ojo de gallo is given the opportunity to spread across the farm. This study investigated the possibility of novel sustainable practices in maintaining healthy coffee plants at Finca Life Monteverde by applying compost to ojo de gallo. We also compared soil erosion in areas of diff ering vegetation cover and found that there was not a significant relationship between percent vegetation cover and soil erosion. We compared the inhibition and reduction in number of colonies of Mycena citricolor after application of various composts prod uced by the farm, a synthetic fungicide called Opus, and the commonly used biocontrol called Trichoderma asperellum (Trichoderma). We found that a compost containing rice and coffee skins, goat and chicken manure, whey, molasses, and carbon produced by the farm significantly inhibited growth of Mycena citricolor the most, and all composts inhibited more so than the synthetic fungicide Opus. Composts have the potential to effectively inhibit the growth of ojo de gallo because their microbial diversity can ou tcompete the pathogenic fungus. The use of vegetation cover as a means of preventing soil erosion should be further investigated to limit the spread of pathogenic fungi. ______________________________________________________________________________ La diversidad microbiana en compost, su eficacia contra el hongo patogno Mycena citricolor y la erosin del suelo en la Finca Life Monteverde RESUMEN La incidencia de Mycena citricolor en las plantas de caf, as como la erosin del suelo, amenazan la productividad de las fincas cafetaleras de la regin de Monteverde. Mycena citricolor comnmente conocido como ojo de gallo, es un hongo patogno que causa defoliacin y prdida de fruta en Coffea arabica El hongo vive en las hojas de caf y en el suelo. A medida que el suelo erosiona, el material frtil para el crecimiento de las plantas se pierde y ojo de gallo se expande a lo largo de la finca. El presente estudio investig el potencia l de ciertas prcticas sostenibles para mantener plantas de caf saludables en la Finca Life Monteverde, aplicando diferentes composts al ojo de gallo. Comparamos adems la erosin del suelo en reas de cobertura vegetal diferente, y no encontramos una rel acin significativa entre el porcentaje de cobertura vegetal y la erosin del suelo. Asimismo, comparamos inhibicin y reduccin del nmero de colonias de Mycena citricolor despus de la aplicacin de diversos compost producidos por la finca, tambin de u n fungicida sinttico llamado Opus, y el biocontrol de uso comn Trichoderma asperellum (Trichoderma). El compost que ms inhibi el crecimiento del hongo fue el que contena broza de caf,

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Ojo de Gallo, Composts, and Soil Erosion Hood, Thurmond 2 estircol de cabra, gallinaza, suero de leche, melaza y carbn pro ducido por la finca. Todos los composts inhibieron ms que el fungicida sinttico Opus. Los composts tienen el potencial de inhibir efectivamente el crecimiento de ojo de gallo porque su diversidad microbiana puede competir con el hongo patgeno. El uso de la cobertura vegetal como medio para prevenir la erosin del suelo debe investigarse ms a fondo para limitar la propagacin de este hongo. ____________________________________________________________________________ Coffee farming is an important part o f Costa economy and culture. At Finca Life Monteverde, ojo de gallo ( Mycena citricolor ) is a common pathogenic fungus that lives on coffee leaves and can infect coffee crops through the soil (Spicer, 2008). This fungus thrives in the same high eleva tion, humid habitats that support the production of world renowned Coffea arabica Plants infected with ojo de gallo typically experience defoliation and fruit loss (Spicer, 2008). This can have serious economic implications for coffee farmers whose harves t is reduced or damaged by the fungus. Presently Life Monteverde uses the synthetic fungicide Opus to combat plants infected with ojo de gallo. Opus is a broad spectrum triazole fungicide containing epoxiconazole for protectant and eradicant activity again st a w ide range of pathogenic fungi Repeated exposure to Opus can have serious health implications for farmers who apply the treatment to their crops. Long term adverse health effects from continuous exposure include organ and reproductive/developmental d amage (Opus, 2014). In an effort to find an alternative to Opus, we investigated adding compost to ojo de gallo as an effective means of prevention by increasing microbial activity. The increase in microbial activity has potential for increasing soil hea lth because soil microorganisms are responsible for central activities associated with soil fertility and plant health (Jousset et al 2014). Thus, microbes are an important driver in the operation of terrestrial ecosystems (Jousset et al 2014). Due to th e overuse of chemical fertilizers, fungicides, herbicides, pesticides and heavy tillage, commercial agriculture has devastated soil microbiota; making it susceptible to soil borne diseases (Bulluck et al 2002). Bacteria and fungi form complex communities of competing and cooperating organisms, hence biodiversity is a major determinant of the growth and activity of microbial communities and affects community health and functioning (Bell et al 2005). Micr obes in healthy soil access, conserve and cycle nutrients and help to regulate the ecosystem. Bacteria and fungi in particular, facilitate fermentation in depleted and sterile soils digesting nutrients and protecting plants against pathogens and other th reats (Bell et al 2005). Mature compost with diverse microbial communities improves the physical and chemical properties of soil; enhancing beneficial bacterial and fungal activity and suppressing infectious pathogenic fungi (Bulluck et al 2002). The co mmonly used biological control for plant pathogenic fungi, Trichoderma was maintained in higher densities in soil treated with organic amendments compared to synthetic fertilizers (Bulluck et al 2002). This microflora community in the compost and its eff ect on soil borne diseases are of particular interest as an alternative to the hazardous chemical Opus. We proposed to test different types of compost used at Life Monteverde to see if there is a correlation between microbial diversity and interactions th at protect against Mycena citricolor Erosion is also a major problem for farmers because it results in the loss of fertile plant material and because the movement of soil can spread soil borne diseases and pathogens, among other things. Thus, soil erosio n is an important factor when considering how to control the distribution and spread of ojo de gallo between coffee crops at Life Monteverde. Heavy precipitation

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Ojo de Gallo, Composts, and Soil Erosion Hood, Thurmond 3 and sloped topography of the region especially contribute to soil erosion in the Monteverde region. We investigated vegetation cover as a proxy for root systems for its potential for holding soil in place. While the relationship between the roots of vegetation and soil erosion has been studied, according to an owner Life Monteverde, there is curren tly not an easy way to control soil erosion nor even measure it (Guillermo Vargas pers. com.) Our study proposed a simple, approachable method to measure the rate of soil erosion and investigate the impact of vegetation coverage upon this rate. Our study poses the questions: How does microbial diversity vary between composts? How effective is compost compared to synthetic fungicide Opus at inhibiting the growth of Mycena citricolor and at killing colonies of ojo de gallo on coffee leaves? What is the relationship between vegetation cover and soil erosion? MATERIALS AND METHODS We conducted our case study at Finca Life Monteverde, Ca itas, Guanacaste, Costa Rica (N10.324414 W 84.84305159999997 from 10 May 2017 through 28 May 2017. Life Monteverde is a coffee farm situated in a tropical pre montane region and landscaped into terraces. The farm contains about 5,000 coffee plants per hectare and 17 hectares total. 2.1 Erosion: We compared soil loss in areas of varying amounts of vegetation at Life Monteverde over a period of 14 days. To measure soil erosion rates, we inserted stakes made of upcycled wood into the soil and recorded vegetation type and percent coverage. The categories varied base d upon the amount of vegetation present: using a densitometer, we grouped each site into vegetation cover 0 24%, 25 49%, 50 74%, or 75 100%. We chose six sites in each of the four categories of vegetation cover throughout the farm. We marked the stake with pencil or Sharpie at level where soil reaches in a line, left the stakes for 14 days, and marked the soil level again. We measured the greatest distance between marks on the stake in centimeters as the soil loss. We used a handheld GPS to pinpoint the loc ation of each stake. 2.2 Microbial Properties of Soil: We compared the effects of different types of compost, Trichoderma biocontrol, and the synthetic fungicide Opus. We conducted laboratory experiments to isolate each treatment from uncontrollable fact ors as much as possible. Our treatments included Compost KOM, Compost KOM L, Compost B, Compost LOM, Compost LOM L, Trichoderma, a synthetic fungicide (Opus), and a control. Compost KOM contains organic matter and waste from the kitchen and KOM L is the li quid that drains from this compost. Compost B is called Bokashi and contains skin from rice and coffee, carbon, soil from the forest, chicken and goat manure, whey, and molasses. Compost LOM is produced by lombricomposting coffee skin (compost with worms), and LOM L is the liquid that drains from this compost. We prepared of each compost (except KOM L and LOM L) by soaking a sample of 2.75g of each compost in containers in a 1:9 ratio (25mL) with Phosphate Buffer solution (Hideaki et al 1 990). Trichoderma and Opus were obtained from Cesar Santamaria at Life Monteverde. The fungicide Opus was prepared in a 250cc:2L ratio as directed on the container. 2.2a Effects of Composts on Infected Leaves:

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Ojo de Gallo, Composts, and Soil Erosion Hood, Thurmond 4 To test the effects of the compost extracts, Trichoderma, and Opus directly on coffee leaves infected with ojo de gallo, we applied six different treatments directly to the leaves, including our control. The treatments for this experiment included Compost KOM L, Compost B, Compost LOM L, Trichoderma, Opus, and a control. We placed three infected leaves in a bucket for each of the six trea tments. We standardized the total number of colonies per bucket to the best of our ability (n=8 or n=9). We applied treatments with a pipette directly to fungal colonies with the leaf horizontal for 30 seconds, then tilted the bucket to directly vertical a gain. We recorded the number of ojo de gallo on leaves colonies and relevant observations about the health of the leaf after three, five, seven, and nine days. We used JMP to run ANOVA tests to determine if there were significant differences between treatm ents on each leaf. 2.2b Measuring antimicrobial properties of compost as an indicator of microflora competition: We compared the antimicrobial properties of our compost extracts compared to Trichoderma and Opus using the agar diffusion test (Bauer et al 1 966). We collected leaves infected with Mycena citricolor and streaked the fungus onto agar plates with scalpels (Bauer et al 1996). We incubated the fungus for approximately seven days at and identified Mycena sp. with the assistance of a biologist at Obregn Biotechnology Lab. We isolated Mycena sp. into a vial with distilled water and swabbed it twice onto 12 plates. We divided each plate in half so that there were four trials in each of six treatments. Immediately upon swabbing the fungus, we appl ied filter paper discs saturated with compost treatments. We observed and measured growth and zones of inhibition daily for three days using calipers. We used JMP Statistical Software to assess the results by running an ANOVA to evaluate variance between a ntimicrobial activity (inhibition zone size) and soil type treatment. 2.2c Surveying biodiversity of composts: We assessed the microbial biodiversity in different composts by swabbing compost treatments and spreading it uniformly over PDA (potato dextrose agar) plates We observed the growth of Com p o st B, LOM, and KOM, as well as a control upon which nothing was swabbed. LOM L and KOM L were not swabbed because we assumed the liquid runoff would have the same microbial composition as its origin compost. We incubated the plates at for one day. We identified fungal morphospecies based upon size, color, texture, and shape of microbe cells using a slide microscope when necessary. We counted the number of different colonies of morphospecies and measured their average sizes with calipers one, two, and four days after initially streaking the compost treatments. We conducted three trials for each treatment and a control. We calculated the Shannon Divers ity Index and evenness, and ran a Special T test to test if there were significant differences between biodiversity o f each compost from each other We tested Compost B vs. KOM, B vs. LOM, B vs. Control, KOM vs. LOM, KOM vs. Control, and LOM vs. Control. RESULTS

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Ojo de Gallo, Composts, and Soil Erosion Hood, Thurmond 5 2.1 Soil Erosion Average soil loss did not differ between percent vegetation cover categories ( Figure 1 ). S oil loss seems to vary depending on vegetation cover, but there is no correlation nor is this assessment statistically supported. Standard deviation between stakes in each category ranged from 30 50%. Figure 1. S oil loss in centimeters over a period of 14 days at 24 different sites. Error bars represent standard deviation. 2.2a Measuring Deleterious Fungal Qualities of Treatments on Leaf A total of five treatments derived from different composts produced at Life Monteverde were compared to the synthetic fungicide Opus for their anti fungal properties. All treatments except Opus resulted in significant inhibition of Mycena sp. in comparison to the control [Figure 2, F (6,57.9) = 20.54 p < 0.0001*]. Compost B and KOM displayed the greatest inhibition, while Opus and control exhibited the smallest inhibition distance on Mycena sp. 0. 0.4 0.8 1.2 1.6 2. 0-24 25-49 50-74 75-100 Soil Loss (cm) % Vegetation Cover

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Ojo de Gallo, Composts, and Soil Erosion Hood, Thurmond 6 F igure 2: Inhibition distance (mm) observed for each treatment applied to fungus cultured from coffee leaves with Mycena sp. spores. Letters represent results from Turkey Kramer HSD pair comparison. Treatments with different letters aresignificsntly different from each other. 2.2b Effects of Composts on Infected Leaves Treatments reduced f ungal colonies more than the control. Compost LOM L appears most effective at reducing the number of living fungal colonies on leaves (Figure 3). All compost treatments were more effective than the Opus treatment at reducing the number of fungal colonies on leaves. However, there was no significant difference in inhibition between any of the treatments after nine days. Figure 3 also shows a decrease in percent of fungal colonies p resent over time, as the control decreases. Figure 3. Percent of fungal colonies remaining on all three leaves, 3, 5, 7 and 9 days after the application of treatments. There were 9 leaves in Control, KOM L, and LOM L buckets and 8 leaves in Compost B, T richoderma, and Opus buckets. 0. 25. 50. 75. 100. Control KOM-L B LOM-L Trichoderma Opus % Fungal Colonies Remaining Treatment Day 3 Day 5 Day 7 Day 9 0 1 2 3 4 5 6 7 Inhibition Distance (mm) Treatmen t KOM KOM L LOM LOM L B Opus Control a b b b b a c d d

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Ojo de Gallo, Composts, and Soil Erosion Hood, Thurmond 7 Figure 4. Change in the number of colonies on each of three leaves per treatment. There is not a statistically significant difference between the treatments and control. 2.2c Sur veying biodiversity of composts Aside from composts KOM and B, there were significant differences between the calculated SWDI of each compost and all other composts (Special T tests, p < 0.05). Composts KOM and B were not significantly different in diversity from one ano ther LOM had the most significantly diverse and even colonies of fungus morphospecies in comparison to all other treatments (Special T Test, p < 0.0001*). Table 1: Diversity analysis of fungal morphospecies cultured from Life Monteverde composts over the course of three days. Compost Type Diversity Index Evenness LOM 0.90 0.51 Comp B 0.30 0.15 KOM 0.24 0.17 Control 0.10 0.15 -2 -1 0 1 2 3 4 5 6 Change in Number of Colonies Treatment Control KOM L B LOM L Trichoderma Opus

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Ojo de Gallo, Composts, and Soil Erosion Hood, Thurmond 8 DISCUSSION Soil Erosion Our results showed that there was not a significant difference in soil loss between th e four selected percent vegetation covers This differs from our expected results; we expected to find vegetation root system would hold soil in place better and percent vegetation cover would account for much of the variance in soil erosion. However, the re are many other factors that influence soil erosion. One field study found that when considering vegetation cover, soil organic matter, leaf litter, and slope gradient, the percent of vegetation cover alone accounted for 76% of the variation in the soil erosion results, while 83% of variation in soil erosion was due to all four aforementioned factors in combination (Meeuwig, 1970). field study concludes that erosion depends heavily upon vegetation cover. In the tropics, steepness of slope and ra infall are factors of particular importance (Presbitero, 2005). Rainfall infiltrates into the soil depending upon various soil properties (organic matter, ratio of sand, silt, and clay) or the water runs off (Presbitero, 2005). Runoff and the flow of water across or down soil also contributes to soil erosion. Another factor to note is the topography of the farm. The terraces of Life Monteverde consist of sloped levels alternating with flat paths. Coffee plants grow on the slopes. We did not standardize the location of the stakes upon the terraces; i.e. the top of the slope or the transition next to the flat path. As soil erodes more in sloped than flat areas (Presbitero, 2005), it is possible that eroded soil could collect in the flat parts of the terraces and actually raise the soil level. This may be why we had greatly variable results (standard deviations of 30 50% in each category). Our results show that percent vegetation cover may have some effect on soil erosion, but did not show a direct relationshi p because there are many factors besides vegetation cover that may have a greater impact upon soil erosion. One source of human error that likely contributed to unexpected results was that our categories of percent vegetation cover were too broad. Perhaps if we had instead examined vegetation cover in 10% increments, our results would have been significant. It is also possible that we had too few trials. Soil erosion is also pertinent to transmission of pathogenic microbes, such as Mycena citricolor Not a ll pathogens are soil borne, but Mycena citricolor is a basidiomycete, which means it can inhabit the soil (Salas, 1972). Prevention of soil erosion is crucial because it maintains fertile material for plant growth, and because it limits the transmission a nd spread of harmful microbes such as ojo de gallo by preventing the movement of infected soil. Vegetation cover may have potential as a means of preventing soil erosion, but our results suggest that many other factors play a role in soil loss. Inhibitio n and Biodiversity Survey Microbial communities associated with fungal inhibition create composite assortments in soil (Kyselkova et al 2009, Mendes et al 2011). Uncovering how the diversity of microbial communities affect interactions with other organisms, namely fungi, may notably improve the understanding of their functioning in protecting plants against soil based pathogens such as Mycena citricolor. R esults of this study support our hypothesis that diversity increases microbial investment in the production of anti fungal compounds and competition. Species richness is a vital driver of the functioning of microbial communities (Bell et al 2005, Langenhe der et al 2010). We showed in vitro that composts with the most diversity resulted in the most significant inhibition of Mycena sp., possibly because of species competition or because some species produced

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Ojo de Gallo, Composts, and Soil Erosion Hood, Thurmond 9 competitive anti fungal compounds. Compost LOM wa s the most diverse in morphospecies colonies harvested over the course of a week on potato dextrose agar (Table 1). Compost LOM inhibited Mycena sp. significantly more than the control and synthetic fungicide Opus (Figure 2). Compost B and diversity indexes did not differ significantly from each other, however were both significantly more diverse than the control (Table 1). Compost B and KOM also displayed the most significant distance of inhibition in comparison to all other treatments in the disc di ffusion portion of our experiment (Figure 2). Antagonistic activity increased with increasing community diversity relative to the control and Opus treatments. This aligns with previous studies that show microorganisms up regulate the production of hostile secondary metabolites in mixed cultures (Jousset et al 2011).The bacteria and fungi in soil form complex communities of competing and cooperating organisms, causing community function to vary with the species present in the soil and their interactions (Jo usset et al 2014). Species may aid each other in resource uptake, but also compete and antagonize each other for limited resources (Jousset et al 2014). The strength of antagonistic interactions between microorganisms may vary with diversity. Pathogens t hat use nutrients to move or grow must compete with the beneficial microflora ( Mehta et al 2014 ). The high population density of fluorescent pseudomonads, actinomycetes and heterotrophic fungi in mature composts has been shown to be responsible for the ge neral suppression of various pathogens (Mehta et al 2014) To further understand this disease suppressive mechanism it would be necessary to look at the regulatory pathways, sensing signals and metabolic activation between competing species of microbes in each type of compost. Determining which specific microbes are responsible for inhibiting pathogenic fungi would als o help reveal why Compost B was more effective at inhibiting Mycena citricolor than other types of microbially diverse composts. There remains the possibility that Opus was ineffective because we used it differently than it is typically used on farms; i.e. we did not reapply the Opus treatment while Life Monteverde reapplies monthly, or perhaps we did not apply the fungicide in sufficient amounts. However, we ultimately attributed the effectiveness to their microbial diversity because all composts were significantly more effective than Opus. Furthermore the results provide an incentive to further investigate other approaches to interpret the consequences of variations in microbial community structure on fungal inhibition. Since soil fungi compri se some of the most infectious plant pathogens, including M. citricolor interactions between bacteria and fungi have direct consequences on plant health, crop yield and ecosystem primary production (Bakker et al 2010, Latz et al 2012). Effects of Composts on Infected Leaves: One in vivo field study found that soils treated with (compost) amendments had increased propagule densities of thermophilic microorganisms and enteric bacteria, but lower propagule densities of pathogenic Phyto pthora and Pythium species of bacteria (Bulluck, 2002). This indicates that higher diversity and density of microorganisms inhibits the survivorship of some other microbes, including pathogenic ones. Our results showed that per leaf, treatments did not sig nificantly vary from control. However, the total number of colonies on all leaves per bucket per treatment did suggest that LOM L was most effective at reducing the number of fungal colonies. The LOM L treatment reduced the number of fungal colonies presen t in the bucket by 100% after 9 days, while the fungal colonies on the control were only reduced by 22.22% after 9 days. This suggests that with more trials or leaves with greater numbers of colo-

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Ojo de Gallo, Composts, and Soil Erosion Hood, Thurmond 10 nies, there may be a difference in the survivorship of fungus upon treatment with composts, especially LOM L. To improve the statistical certainty of the differences in treatments, we might observe leaves with more colonies. Our biodiversity survey of different composts showed that Compost LOM was the most diverse a nd even, and the leaf experiment suggests that Compost LOM L has the greatest effect on reducing the number of colonies. Since Compost LOM L is the liquid that drains from Compost LOM, it is likely that their microbial composition is similar. This supports our hypothesis that Compost LOM L treatment was most effective in the leaf experiment because its morphospecies richness outcompeted Mycena Citricolor and killed the colonies on the leaves. More precise investigation needs to be done in order to fully u nderstand the relationship between vegetation cover and soil erosion, as well as the effect of compost treatments applied directly to infected leaves on the living tree. We developed an accessible and approachable method of measuring soil loss over time us ing upcycled wood stakes, which will help interested farmers measure soil erosion. Our results suggest that compost microbial diversity is likely connected to its effectiveness in inhibiting the growth of Mycena citricolor because composts were significa ntly more effective than Opus. We found that compost treatments may be suitable for decreasing the incidence of ojo de gallo on coffee plants. ACKNOWLEDGEMENTS We would like to sincerely thank Sofa Arce Flores for her guidance throughout our investigati on and writing process, for collecting data for us, procuring equipment and snacks, taking us to our field sites, connecting us with Finca Life, and providing us with infinite happiness in the form of a giggly baby named Abi. We are so appreciative to the owners and workers of Finca Life Monteverde, especially to Guillermo Vargas for his support and to Cesar Santamaria for providing us with composts. Thank you to Luisa for her help in the laboratory and Jorge for helping us construct stakes at the Monteverd e Institute, to all of our EAP instructors for answering questions and encouraging us, and to Andrs Camacho and Becca Sanchez for their suggestions. Thank you to Julian Cassano for making time spent at the farm so enjoyable, and to Hannah Wilson, Annie Gorges, and Marisa Yang for moral support and making us laugh. REFERENCES Bakker, M. G. J. D. Glover, J. G. Mai, and L. L. Kinkel. 2010. Plant community effects on the diversity and pathogen suppressive activity of soil streptomycetes. Applied Soil Ecology 46 : 35 42. BASF Safety Data Sheet. 2014. Opus. Retrieved May 31, 2017, from http:// agro.basf.co.nz/products/fungicides/opus.aspx. Bauer, A. W., Kirby, W. M. M., Sherries, J. C., and Turck, M. 1966. Antibiotic Susceptibility Testing by a Standardized Single Disk Method. TURCK American Journal of Clinical Pathology 45 : 493 496. Bell, T., J. A. Newman, B. W. Silverman, S. L. Turner, and A. K. Lilley. 2005. The contribution of species richness and composition to bacterial services. Nature 436 : 1157 1160. Bulluck, R. L., Brosius, M., Evanylo, G. K., and Ristaino, J. B. 2002. Organ ic and synthetic

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