Coinfections are increasingly recognized as important drivers of disease dynamics. Consequently, greater emphasis has been placed on integrating principles from community ecology with disease ecology to understand within-host interactions among parasites. Using larval amphibians and two amphibian parasites (ranaviruses and the trematode Echinoparyphium sp.), we examined the influence of coinfection on disease outcomes. Our first objective was to examine how priority effects (the timing and sequence of parasite exposure) influence infection and disease outcomes in the laboratory. We found that interactions between the parasites were asymmetric; prior infection with Echinoparyphium reduced ranaviral loads by 9% but there was no reciprocal effect of prior ranavirus infection on Echinoparyphium load. Additionally, survival rates of hosts (larval gray treefrogs; Hyla versicolor) infected with Echinoparyphium 10 days prior to virus exposure were 25% greater compared to hosts only exposed to virus. Our second objective was to determine whether these patterns were generalizable to multiple amphibian species under more natural conditions. We conducted a semi-natural mesocosm experiment consisting of four larval amphibian hosts [gray treefrogs, American toads (Anaxyrus americanus), leopard frogs (Lithobates pipiens) and spring peepers (Pseudacris crucifer)] to examine how prior Echinoparyphium infection influenced ranavirus transmission within the community, using ranavirus-infected larval wood frogs (Lithobates sylvaticus) as source of ranavirus. Consistent with the laboratory experiment, we found that prior Echinoparyphium infection reduced ranaviral loads by 19 to 28% in three of the four species. Collectively, these results suggest that macroparasite infection can reduce microparasite replication rates across multiple amphibian species, possibly through cross-reactive immunity. Although the immunological mechanisms driving this outcome are in need of further study, trematode infections appear to benefit hosts that are exposed to ranaviruses. Additionally, these results suggest that consideration of priority effects and timing of exposure are vital for understanding parasite interactions within hosts and disease outcomes.

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 trade‐offs including population susceptibility to parasites. We have identified 15 wood frog populations that exist along a gradient from close to agriculture and high, constitutive pesticide tolerance to 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 laboratory 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.

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.

The study of priority effects with respect to coinfections is still in its infancy. Moreover, existing coinfection studies typically focus on infection outcomes associated with exposure to distinct sets of parasite species, despite that functionally and morphologically similar parasite species commonly coexist in nature. Therefore, it is important to understand how interactions between similar parasites influence infection outcomes. Surveys at seven ponds in northwest Pennsylvania found that multiple species of echinostomes commonly co-occur. Using a larval anuran host (Rana pipiens) and the two most commonly identified echinostome species from our field surveys (Echinostoma trivolvis and Echinoparyphium lineage 3), we examined how species composition and timing of exposure affect patterns of infection. When tadpoles were exposed to both parasites simultaneously, infection loads were higher than when exposed to Echinoparyphium alone but similar to being exposed to Echinostoma alone. When tadpoles were sequentially exposed to the parasite species, tadpoles first exposed to Echinoparyphium had 23% lower infection loads than tadpoles first exposed to Echinostoma. These findings demonstrate that exposure timing and order, even with similar parasites, can influence coinfection outcomes, and emphasize the importance of using molecular methods to identify parasites for ecological studies.

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.