Tag Archives: Benchwork

1) Quantifying and characterizing the Chagas disease parasite burden in kissing bug vectors across land use change gradients

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Rhodnius pallescens, principal vector of Chagas disease in Panama

Mentors: Dr. Nicole Gottdenker and Juliana Hoyos
Abstract: Anthropogenic environmental change has been associated with the emergence of zoonotic vector borne pathogens. In this project, the REU scholar will investigate how parasite burden with the Chagas disease agent Trypanosoma cruzi varies in kissing bug vectors collected across different gradients of deforestation and reforestation in rural areas of central Panama. The student will learn laboratory methods in molecular detection of trypanosomes in kissing bugs and will also apply statistical methods to investigate how land cover, bug abundance, microclimate, stage, and bug population characteristics relate to parasite burden in the bugs. 
Is the project computational, empirical, or both? Both.

How Various Feeding Rates Affect Pupation Rates in Anopheles stephensi Larvae

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Jacob Glover, a student at Franklin College, worked in the lab of Dr. Ash Pathak

Abstract Anopheles Stephensi are a dangerous vector for countless diseases without cures currently. If mosquitos could be contained or controlled, then this could eliminate these diseases they carry without having to find a vaccine or other types of cures for them. One of the best stages to control mosquitos is within their larval stage of life. In this stage, they are so fragile that minor changes in food or environment can kill them or stunt their growth. If mosquito larvae could be purposefully stunted or killed off, then there would be one less major vector for disease. Our experiment tests this hypothesis to see if manipulating the rate at which mosquito larvae are fed if that impacts their pupation rates.Within this experiment, we test to see if various feeding rates affect their pupation rates, ultimately testing if their growth stays the same under different feeding conditions. This was done by giving them different amounts of TetraFin pellets on Tuesdays and Wednesdays to see if there was a difference in growth rates. Overall, the results show that no significant changes occur in the pupation rates of mosquito larvae when feeding rates are altered.

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Sensing Your Friends Getting Eaten is Stressful, Having a Parasite Makes it Worse

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Helen Gloege, a student in Mount Holyoke College, worked in the lab of Dr. Andy Davis

Abstract Daphnia and other animals face a multitude of different stressors in their daily lives. Parasites can cause various physiological changes in animals, yet few prior studies have looked at the combination of parasitism and stress in animal models. This study used Daphnia which are microscopic plankton that may have ectoparasite-like organisms attached to their surface. Vorticella, a single-celled ectoparasite, is one of these organisms that can be found attached to Daphnia. During this experiment, when Daphnia ambigua were exposed to a “stressor” the vorticella parasite led to an increased heart rate in the Daphnia. The “stressor” was made of macerated Daphnia and aimed to simulate a predation event. Daphnia without vorticella present appeared to have no discernable reaction to the stressor. While Daphnia with vorticella increase the heart rate and physiological stress reaction in Daphnia. When over twenty vorticella were present on the daphnia the heart rate continued to increase during the study period.

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Tricks Not Treats: Wolbachia’s Manipulation of Sex in Infected D. subquinaria Offspring

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Madeline Sheppard, a student at Eckerd College, worked in the lab of Dr. Kelly Dyer

Abstract Wolbachia is a bacteria that is found in up to 60% of all insects, which is transmitted exclusively from mother to offspring through the egg. In many host species Wolbachia infection does not Wolbachia are gram-negative maternally transmitted bacterial endosymbionts that are found in upwards of 60% of arthropods. Here, we aimed to determine the phenotype expressed in D. subquinaria when they are infected with a strain of Wolbachia originally found in a closely related species, D. recens. Wolbachia infection is expressed as one of two phenotypes: Cytoplasmic incompatibility (CI), which causes the deaths of most infected offspring, and Male Killing (MK), which causes the death of the sons of infected males. Additionally, we tested how Wolbachia interacts with the varied genetic backgrounds of D. subquinaria populations, as well as how that would influence which phenotype is displayed in infected offspring. We tested a total of 15 lines of D. subquinaria by taking 2 to 3 infected females and crossing them with 1 uninfected male. The F1 offspring from those crosses were collected, and the sex distribution per line’s offspring was determined. Ten lines produced a significant number of flies (n>10). We tested for Wolbachia infection by using PCR, and determined the F1 generation was positive for Wolbachia. All lines expressed evidence of the MK phenotype, and four lines showed nearly complete MK (98-100% female). The remaining six expressed partial MK (60-84% female). In the future, we aim to determine whether there is a genetic suppressor of the MK phenotype in D. subquinaria, as well as to generalize the results of this project to further understand the population dynamics of Wolbachia-infected flies.

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Temperature fluctuation on disease transmission in multi-host communities

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Jenavier Tejada, a student at Denison University, worked in the lab of Dr. Alex Strauss

Abstract The dilution effect seeks to explain disease transmission in environments with multiple species. Essentially, the dilution effect predicts an increase in diversity will lead to a decrease in disease transmission. In zooplankton communities, the resistant diluter, Ceriodaphnia dubia can lessen disease in the host Daphnia dentifera caused by the parasite Metschnikowia bicuspidata. However, dilution is only effective when diluters and hosts co-exist; because when they compete, competitive exclusion can occur. Fitness of both D. dentifera, and C. dubia depend on temperature. Specifically, C.dubia benefits in warmer temperatures and D. denifera in cooler temperatures. Therefore, in environments where temperatures fluctuate,  this may lead to co-existence, greater abundances of the diluter, and less disease transmission. We are testing whether the dilution effect reduces infection prevalence when a diluter is present, and how dilution effects differ at a constant 20˚C versus a fluctuating temperature around the same mean. We designed a multi-generational mesocosm experiment with communities that contained the host and parasite, and communities that contained the hosts, parasites, and diluters at both constant and fluctuating temperatures. We hypothesize that the changing environmental conditions caused by fluctuating temperature will lead to more diluters, causing a greater dilution effect via co-existence of the host and diluter. This project will help us learn more about the possible effects of climate change – especially variable temperature – on disease dynamics in communities with multiple species.

Tejada_REU-Poster

Approximating abundance of Daphnia dentifera using environmental DNA (eDNA) samples

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Emily Landolt, a student in St. Norbert College, worked in the lab of Dr. Alex Strauss

Abstract Freshwater zooplankton, such as Daphnia dentifera, are helpful model organisms for studying infectious disease dynamics and are ecologically important because of their role in food webs. They are consumers of primary producers like algae and are prey for other organisms such as fish. However, field sampling of freshwater zooplankton can be costly in terms of time and effort. The use of eDNA to estimate species abundances rather than an assessment of presence/absence in aquatic ecosystems is an exciting concept because it allows for more efficient and potentially more accurate field sampling. eDNA sampling methods for this study system can also be important for monitoring populations and epidemics in the field to ensure ecosystem health. This methodology has been studied in fish but is not yet well understood for invertebrates. We explored this idea by generating artificial mesocosm populations of Daphnia. Sampling for species abundance was done in two ways: using a traditional observational method of taking a subsample and counting the number of Daphnia, and a novel molecular method to quantify the amount of eDNA using a qPCR assay. Our results are a correlation between the observed species abundance and the molecular quantification of eDNA. Preliminary results show insignificant relationships indicating complexity that needs to be explored. Possible factors that affected the amount of eDNA in the water include water chemistry (i.e. pH), age structure, time since introduction, and water replenishment from evaporation/sampling. In the future, more research will be needed to explore the factors of eDNA persistence and how it can be used to better approximate species abundances of aquatic invertebrates like Daphnia.

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Roadkill as sentinels for parasite detection in a wild squirrel population

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Jonah Giermann, a student at College of St. Scholastica, worked in the lab of Dr. Sonia Altizer

Abstract Disease surveillance of wild populations is difficult, as capture and release techniques can be time-iRoadkill is an excellent display of human-wildlife conflict. Carcasses can inform ecologists about population trends, species distribution, and behavior. Carcasses can be inspected for parasitic infection, but data obtained is subject to bias from a variety of factors. We chose rock squirrels as our study system because their behavior makes them host to a variety of parasitic species. I asked questions regarding the factors influencing measures of infection (richness, abundance, diversity) in roadkill, and how those measures compare in live-trapped samples.Samples were collected in Zion National Park from 2020-2022. Roadkill intestines were dissected, and whole worms were counted. Fecal samples were obtained from live trapped squirrels. Fecal flotation using sodium solution was conducted and any resulting eggs were counted. We found that squirrels collected in autumn host more parasites than the other two seasons. Male squirrels are collected as roadkill more frequently than females. Males display exploratory and dispersal behavior, likely causing more vehicle collisions. Parasite richness was higher in the roadkill population than it was in the livetrap population.  We used the Shannon diversity index to get a parasite diversity value for each sample type. The diversity index factors in both richness and abundance to analyze diversity. Roadkill had higher diversity, but livetrap had a similar value. Microparasites were found in live-trapped samples but could not be seen using our roadkill sampling method. Based on these results, roadkill seems to be an effective indicator of parasitic infection in wild populations, provided biases are accounted for.

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Spatial Variation in Oyster Macroparasites Across the Georgia Coast

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Sofia Markiewicz, a student at Scripps College, worked in the lab of Dr. Jeb Byers

Abstract Oysters are a key coastal foundation species that have declined drastically across the US coasts due to the combined effects of overharvesting, pollution, and disease. With climate change, there The eastern oyster (Crassostrea virginica) is a keystone species and ecosystem engineer that stabilizes sediments, cycles nutrients, improves water quality, and provides habitat for fish and crustaceans. Oysters are prone to several macroparasites, including pea crabs (Zaops ostreum), mud blister worms (Polydora websteri), and boring sponge (Cliona spp.), all of which can damage their gill tissue and shells. Although oyster populations have been widely studied in other areas of the eastern United States, the geographic and environmental factors that influence macroparasite infection in Georgia’s oyster population are still largely unknown. In this study, we sampled oysters from 24 reefs across eight distinct sites along the Georgia coastline and examined them for macroparasite infection. The relationships between macroparasite prevalence and geographic location and environmental conditions (specifically reef complexity, reef shell density, dissolved oxygen, water temperature, and salinity) were examined. We found no correlation between location and macroparasite prevalence for any of the macroparasites examined. However, increased prevalence of blister worms was correlated with low salinity and low reef complexity. Prior research has also shown that shellfish infected with blister worms exhibit decreased shell strength, and are therefore more vulnerable to damage and predation. Understanding what conditions affect blister worm prevalence and how they may be altered by by climate change (e.g. changing salinity) is important for evaluating locations where oyster reefs are likely to have low macroparasite infection and be less prone to damage, in order to better maintain high-quality reef habitat. This is especially crucial in a relatively understudied environment such as Georgia, where the effects of these conditions are less well known. 

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Seeing What Floats: Comparing Fecal Diagnostic Techniques for the Detection of Zoonotic Cestode Eggs

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Sierra Felty, a student at Radford University, worked in the lab of Dr. Christopher Cleveland

Abstract Coyotes (Canis latrans) are an anthropogenically abundant and increasingly widespread species, Members of the genus Echinococcus are parasitic cestodes that pose a zoonotic threat to wildlife, livestock, domestic animals, and humans. They utilize wild canids, such as coyotes and foxes, as their definitive hosts. Two species of interest are E. multilocularis and E. granulosus given their impacts on native wildlife, agriculture, and human health. E. multilocularis uses rodents as intermediate hosts, and E. granulosus utilizes cervids as intermediate hosts. As the rate of urbanization continues to rise, humans and domestic animals are at greater risk of infection through more frequent interactions with wild canid hosts. Given the increased risk of infection, especially in non-endemic regions, it is important to have reliable detection methods in place. Our study sought to test the sensitivity and detection limits of three different fecal flotation methods (centrifugal, passive, and Mini-FLOTAC) in recovering Echinococcus spp. eggs. For each method, fecal samples were spiked with a known concentration of eggs (25, 40, and 60 eggs per gram of feces) and zinc sulfate was used as the flotation solution. Our findings indicated that the centrifugal flotation and Mini-FLOTAC were the most sensitive for detecting Echinococcus spp. eggs and the Mini-FLOTAC had the highest egg recovery. Therefore, the Mini-FLOTAC appears to be the most reliable fecal flotation method in detecting Echinococcus spp. eggs.

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Predictors for SARS-CoV-2 Seropositivity in Owned and Feral Cats in North Georgia

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Sarah Blankespoor, a student at California Polytechnic University, worked in the lab of Dr. Mark Tompkins

Abstract Little is known about the epidemiology of SARS-CoV-2 in animal populations. Cats are a host for the virus, with cat-to-cat transmission demonstrated in lab settings. Both feral and owned cats interact with many species and could drive interspecies transmission. This project investigates dynamics of SARS-CoV-2 by evaluating seropositivity predictors in cats. Serum samples were taken from owned cats brought to the University of Georgia Veterinary Teaching Hospital from 08/2021-06/2022 and feral cats captured locally from 01/2022-06/2022. Samples were tested for anti-SARS-CoV-2 antibodies using indirect ELISAs. For feral cats with n=33, none of the samples were positive. For owned cats with n=193, 10 of the samples were positive (5.2%). There is preliminary evidence for lasting antibodies with two repeat positive cats, with samples taken up to 3 months apart. Binary logistic regression models for the owned cats were determined in R through multi-model inference. Two terms were present in the 3 equivalent best models: cumulative human COVID-19 cases by county, with a positive coefficient; and days since the pandemic started, with a negative coefficient. These results suggest that cats acquire SARS-CoV-2 infections from humans rather than other cats or wildlife. The negative coefficient for time in the models can be explained by the delta and omicron surges at the beginning of the study period. SARS-CoV-2 surges in humans have a ripple effect into the larger ecosystem, particularly for cats owned by humans with COVID-19. Future research should continue to investigate this impact over a larger time scale and expand feral cat sample size to confirm observed trends.

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Infection and Spore Yield of Daphnia Microsporidian

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Hannah O’Grady, a student at Mount Holyoke College, worked in the lab of Dr. Alex Strauss.

Abstract An important part of understanding how diseases spread and impact a community is understanding the tradeoffs that occur when a parasite generalizes. While sampling ponds in Whitehall Forest we discovered a potentially novel microsporidian that was able to infect at least five zooplankton in the Cladocera superorder at relatively high infection prevalence. We designed an experiment to investigate the potential costs of its generalism. We exposed isoclonal lines of two different species of Daphnia, each from two different lakes, to spores gathered from the dominant host in each lake to test whether the microsporidian was 1) more successful at infecting Daphnia of one species over the other or 2) whether it was more successful at infecting Daphnia from the same lake or 3) the same species from which the parasite spores were gathered. We also counted spore yield (a metric of parasite fitness) from the six infected species gathered from the field to test for differences across species, lakes or time. None of the Daphnia exposed to the spores in the lab became infected, leading us to hypothesize that there is an intermediate host for this parasite. Spore yields from field-collected hosts did differ significantly among host species, with higher spore yields in D. laevis (mean=446,415.55±412,977.53) and Diaphanosoma (mean=253,888.83±78,085.34) than in D. ambigua (mean=40729.12±41,396.68), D. parvula(mean=46250.06±49,432.78), and Simocephalus (mean=17083.17±13,025.14). There were no significant differences in spore yield across lakes or days. More research will be needed to find the intermediate host for the microsporidian as well as to determine its exact genus and species.

OGrady

Royally Split: Morphological divergence of parasites in milkweed butterflies

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Katie Yan, a student from Skidmore College, worked in Dr. Sonia Altizer‘s lab.

Abstract Ophryocystis elektroscirrha (OE) is a protozoan parasite found in Monarchs (Danaus plexippus). Same or similar OE-like parasites have been found in Queens (D. gilippus) and other Danaus butterflies. Experimental cross infection provided evidence of parasite specialization on natal host species via low infection rates on novel hosts, motivating in-depth analysis of parasite morphometric and genetic variation across host species from different locations. To examine OE and OE-like spore morphology across Danaus species, we looked at five host species from previously curated museum samples including: Jamaican Monarch (D. cleophile), Plain Tiger (D. chrysippus), Queen (D. gilippus), Common Monarch (D. plexippus), and Lesser Wanderer (D. petilia). Analysis of wild-collected museum spores showed that on average, Jamaican Monarchs and Common Monarchs had larger spores than other species. This relationship is consistent with the phylogenetic relationships of host similarity within the Danaus genus and further supports the hypothesis of parasite specialization on hosts. We then examined the influences of host and environmental factors on parasite morphology by analyzing spores from a cross-infection experiment involving Monarchs and Queens. Hosts in this experiment were fed either tropical milkweed (Asclepias curassavica) or swamp milkweed (A. incarnata). Monarchs and Queens were infected at a high rate by their natal parasites, some Monarchs were infected by Queen parasites, and no Queens were infected by Monarch parasites. Morphometric analysis of spores showed that Monarch parasites from Monarch hosts were largest, and that Queen parasites from Queen hosts were smallest; Queen parasites from Monarch hosts were intermediate in spore size. Other spore traits (shape, hue, density) were similar across treatments. Additionally, we found that spore size positively correlated with host wing area, suggesting that larger spores are found on larger butterflies. Milkweed species, sex, and final spore load did not predict variation in spore morphology. In summary, we found strong evidence for parasite specialization on different host species based on differences in spore size; further work should ask whether molecular genetic divergence of OE parasites across host groups matches differences in spore morphology and host phylogeny.

Yan

Phylogenetics of Dracunculus Nematodes in North America

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Madeline Giner, from the University of Texas at San Antonio, studied the phylogenetics of Dracunculus in the lab of Dr. Christopher Cleveland.

The Dracunculus genus contains parasitic nematodes that infect a variety of hosts, including reptiles and mammals. Dracunculus medinensis, the Guinea worm, has gained much attention due to its history of infecting humans. Less studied are other dracunculids, including Dracunculus insignis and D. lutrae which are native to North America. D. insignis can infect a variety of mammalian hosts, whereas D. lutrae specifically infects North American river otters (Lontra canadensis). The goal of this project was to investigate the genetic diversity of Dracunculus in wildlife species from the Eastern USA and investigate spatial and host patterns of infection. Phylogenetic relationships were examined using the cytochrome c oxidase I (COI) gene targets. We hypothesized otters would predominantly have D. lutrae, and D. insignis would be identified from other hosts. Our experimental methods included DNA extraction, gene-specific amplification (PCR), Sanger sequencing, and phylogenetic analysis using the software Geneious. Our results indicate that a majority of worms from otters (19/65), raccoons (Procyon lotor, 22/22), and Virginia opossums (Didelphis virginiana, 2/2) were D. insignis. However, a worm from an otter from Florida had 100% identity to a novel Dracunculus sp. previously detected in Georgia, and a Georgia otter worm is closely related to another novel dracunculid species from Florida. In conclusion, D. insignis was present in most locations and hosts, D. lutrae is absent, and an additional host is now known for two novel Dracunculus species. These data provide new information about Dracunculus diversity in US wildlife, but additional investigation is required.

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