Interaction between Insecticide Exposure and Trematode Infection across Four Wood Frog Populations

Amphibian populations are declining worldwide due to a number of stressors including pesticides and parasites. Conservation of these animals can be complicated because populations can differ dramatically in response to the same stressor. When consistently exposed to pesticides, some populations evolve tolerance through the process of natural selection acting across multiple generations. Alternatively, populations that are intermittently exposed to pesticides induce tolerance within a single generation. To date, however, there have been few studies examining the costs associated with these different stress tolerance mechanisms. In this study, we examined how difference in stress tolerance influence susceptibility to parasitic infections. We collected wood frog tadpoles from four different populations: two with evolved tolerance to pesticides and two with the ability to induce pesticide tolerance. We exposed tadpoles from each population to sublethal doses of carbaryl (0 and 5 ppm) for 5 days. Tadpoles were allowed to acclimate in pesticide-free water for 2 days. After this acclimation period, we then exposed tadpoles to 0 or 50 trematode parasites (Echinostoma trivolvis) for 2 days and counted the number of parasites encysted within the body. Exposure to sublethal carbaryl decreased susceptibility to trematodes for tadpole populations with evolved pesticide tolerance. In contrast, exposure to sublethal carbaryl increased susceptibility to trematodes for tadpole populations with induced pesticide tolerance. This suggests that populations with the ability to induce pesticide tolerance incur the cost of increased disease risk. This has important conservation implications for understanding a population’s history and defending against disease

The benefits of coinfection: Trematode infections alter pathogen interactions and disease outcomes in hosts

Emerging infectious diseases are affecting host populations of humans, wildlife, and plants. Historically, epidemiological research has focused on singular hosts and singular pathogens. However, growing evidence suggests that many pathogens co-occur in wild populations, increasing the likelihood of coinfection within hosts. Pathogen interactions within hosts are likely to change disease dynamics, which has important implications for host pathology and pathogen transmission within communities. Incorporating principles from community ecology are important for understanding the consequences of coinfection on disease outcomes within natural systems. Amphibians are a useful study system for exploring the influence of coinfection on both within- and between-host interactions. Two amphibian pathogens that naturally occur with one another are ranaviruses and echinostomes. Here, I examined the influence of pathogen interactions on disease outcomes at both the individual and community level. My objectives were to: 1) examine how the sequence and timing of pathogen exposure influenced infection success and disease outcomes at the individual level, and 2) determine how pathogen interactions influence between-host processes and disease outcomes. In the laboratory experiment using larval gray treefrogs, I found that interactions between ranaviruses and echinostomes were asymmetric. Prior echinostome infection influenced ranavirus infection but there was no effect of ranavirus on echinostomes. The prior infection of echinostomes reduced viral loads across different exposure times by 9%. Additionally, host survival rates in individuals exposed to echinostomes 10 days prior to virus exposure significantly increased by 25% when compared to virus only treatments. These results led me to conduct an experiment to determine if these pathogen interactions could be scaled up to more natural conditions within a community. I found that with the prior infection of echinostomes, viral load decreased by 19%, 27%, and 28% in gray treefrogs, leopard frogs, and spring peepers respectively, with no effect on American toads. These results suggest that macroparasite infection can reduce microparasite replication rates across multiple amphibian species, possibly through cross-reactive immunity. While this is the first experimental study to examine the influence of coinfection on disease dynamics within amphibian communities, my results demonstrate that laboratory patterns of coinfection dynamics within amphibian hosts are broadly applicable to more natural settings and host communities

Amphibian Host and Skin Microbiota Response to a Common Agricultural Antimicrobial and Internal Parasite

Holistic approaches that simultaneously characterize responses of both microbial symbionts and their hosts to environmental shifts are imperative to understanding the role of microbiotas on host health. Using the northern leopard frog (Lithobates pipiens) as our model, we investigated the effects of a common trematode (family Echinostomatidae), a common agricultural antimicrobial (Sulfadimethoxine; SDM), and their interaction on amphibian skin microbiota and amphibian health (growth metrics and susceptibility to parasites). In the trematode-exposed individuals, we noted an increase in alpha diversity and a shift in microbial communities. In the SDM-treated individuals, we found a change in the composition of the skin microbiota similar to those induced by the trematode treatment. Groups treated with SDM, echinostomes, or a combination of SDM and echinostomes, had higher relative abundances of OTUs assigned to Flavobacterium and Acinetobacter. Both of these genera have been associated with infectious disease in amphibians and the production of anti-pathogen metabolites. Similar changes in microbial community composition between SDM and trematode exposed individuals may have resulted from stress-related disruption of host immunity. Despite changes in the microbiota, we found no effect of echinostomes and SDM on host health. Given the current diseaseand pollution-related threats facing amphibians, our study highlights the need to continue to evaluate the influence of natural and anthropogenic stressors on host-associated microbial communities

Amphibian Skin Microbiota Response to Variable Housing Conditions and Experimental Treatment across Space and Time

Host-associated microbial ecology research is becoming a popular tool in conservation. For amphibians, there is recognition that environmental factors and anthropogenic activities can alter the composition and function of skin microbiotas. Despite growing studies on this topic, the environmental conditions and experimental methods that amphibians are exposed to vary among studies, potentially influencing our ability to develop generalizations. Using Northern Leopard Frogs (Lithobates pipiens), we investigated how the movement of individuals between housing conditions affected the amphibian skin microbial communities. In addition, we evaluated whether variation in experimental venue (e.g., culture pools, mesocosms, or laboratory) and time influenced the effect of a common antibacterial (sulfadimethoxine; SDM) on the skin microbiota. We found that the microbial community diversity decreased when tadpoles were transferred from culture pools to mesocosms and subsequently increased between mesocosms and the laboratory. When comparing the effect of SDM exposure on the skin microbiota of tadpoles across experimental venues (mesocosm and lab), there was no effect of SDM on alpha diversity. However, we noted opposing patterns between the control and SDM-treated individuals within the mesocosm and laboratory groups. In the laboratory, there were differences in the abundances of operational taxonomic units (OTU) while in the mesocosm there were differences in OTU turnover. Finally, we found that SDM treatment on amphibian microbial communities was consistent across time in the laboratory. Because researchers are integrating microbial assessments into our understanding of conservation biology, our results underscore the importance of standard housing conditions and taking into consideration that experimental design may yield variable results

Life Stage and Proximity to Roads Shape the Skin Microbiota of Eastern Newts (Notophthalmus viridescens)

Host-associated microbiomes play an essential role in the health of organisms, including immune system activation, metabolism, and energy uptake. It is well established that microbial communities differ depending on the life stage and natural history of the organism. However, the effects of life stage and natural history on microbial communities may also be influenced by human activities. We investigated the effects of amphibian life stage (terrestrial eft vs. aquatic adult) and proximity to roadways on newt skin bacterial communities. We found that the eft and adult life stages differed in bacterial community composition; however, the effects of roads on community composition was more evident in the terrestrial eft stage compared to the aquatic adult stage. Terrestrial efts sampled close to roads possessed richer communities than those living further away from the influence of roads. When accounting for ASVs with predicted antifungal capabilities, in the adult life stage, we observed a decrease in anti-fungal bacteria with distance to roads. In contrast, in the eft stage, we found an increase in anti-fungal bacteria with distance to roads. Our results highlight the need to consider the effects of human activities when evaluating how host-associated microbiomes differ across life stages of wildlife

High landscape-level gene flow in the red-backed salamander (Plethodon cinereus)

Anthropogenic changes are expected to shape the genetic structure of many herpetofaunal populations. Indeed, genetic analyses are often applied in disturbed habitats because they can determine the loss of genetic diversity in isolated populations, identify barriers to dispersal and gene flow, and inform ways in which populations should be managed. As a result, there is a lack of baseline genetic data of organisms in relatively intact landscapes. Plethodontid salamanders are ideal for understanding fine-scale and landscape level genetic structure in continuous landscapes given their high abundance and their sensitivity to ecosystem changes. Given this, our study aimed to assess the landscape-level population structure in red-backed salamanders (Plethodon cinereus) within intact habitat. We investigated genetic variation in 479 individuals from three sites across roughly 26 km of intact hardwood forest. We tested for genetic structure among and within three sites using several metrics, including hierarchical Bayesian analysis, FST analysis, AMOVA, and isolation by genetic distance. The results revealed a single population throughout the study area and genetic structure within sites was not evident. Importantly, the sites in this study are part of a long-term project on the effects of standard silvicultural practices in the American Midwest, making it ideal for future re-evaluation of this red-backed salamander population. We now have baseline information to determine if any future disturbances or fragmentation may alter the genetic structure of this population, as well as how this population may change over time if the study area remains undisturbed

High landscape-level gene flow in the red-backed salamander (Plethodon cinereus)

Anthropogenic changes are expected to shape the genetic structure of many herpetofaunal populations. Indeed, genetic analyses are often applied in disturbed habitats because they can determine the loss of genetic diversity in isolated populations, identify barriers to dispersal and gene flow, and inform ways in which populations should be managed. As a result, there is a lack of baseline genetic data of organisms in relatively intact landscapes. Plethodontid salamanders are ideal for understanding fine-scale and landscape level genetic structure in continuous landscapes given their high abundance and their sensitivity to ecosystem changes. Given this, our study aimed to assess the landscape-level population structure in red-backed salamanders (Plethodon cinereus) within intact habitat. We investigated genetic variation in 479 individuals from three sites across roughly 26 km of intact hardwood forest. We tested for genetic structure among and within three sites using several metrics, including hierarchical Bayesian analysis, FST analysis, AMOVA, and isolation by genetic distance. The results revealed a single population throughout the study area and genetic structure within sites was not evident. Importantly, the sites in this study are part of a long-term project on the effects of standard silvicultural practices in the American Midwest, making it ideal for future re-evaluation of this red-backed salamander population. We now have baseline information to determine if any future disturbances or fragmentation may alter the genetic structure of this population, as well as how this population may change over time if the study area remains undisturbed

DRYAD_JHUA

Data for: Evolved pesticide tolerance influences susceptibility to parasites in amphibian

Data from: Evolved pesticide tolerance influences susceptibility to parasites in amphibians

Because ecosystems throughout the globe are contaminated with pesticides, there is a need to understand how natural populations cope with pesticides and the implications for ecological interactions. From an evolutionary perspective, there is evidence that pesticide tolerance can be achieved via two mechanisms: selection for constitutive tolerance over multiple generations or by inducing tolerance within a single generation via phenotypic plasticity. While both mechanisms can allow organisms to persist in contaminated environments, they might result in different performance tradeoffs including population susceptibility to parasites. We have identified 15 wood frog populations that exist along a gradient from a close to agriculture and high, constitutive pesticide tolerance to a far from agriculture and inducible pesticide tolerance. Using these populations, we investigated the relationship between evolutionary responses to the common insecticide carbaryl and host susceptibility to the trematode Echinoparyphium lineage 3 and ranavirus using lab exposure assays. For Echinoparyphium, we discovered that wood frog populations living closer to agriculture with high, constitutive tolerance experienced lower loads than populations living far from agriculture with inducible pesticide tolerance. For ranavirus, we found no relationship between the mechanism of evolved pesticide tolerance and survival, but populations living closer to agriculture with high, constitutive tolerance experienced higher viral loads than populations far from agriculture with inducible tolerance. Land-use and mechanisms of evolved pesticide tolerance were associated with susceptibility to parasites, but the direction of the relationship is dependent on the type of parasite, underscoring the complexity between land use and disease outcomes. Collectively, our results demonstrate that evolved pesticide tolerance can indirectly influence host-parasite interactions and underscores the importance of including evolutionary processes in ecotoxicological studies