Canopy coverage, light, and moisture affect thermoregulatory trade-offs in an amphibian breeding habitat.

When amphibians thermoregulate, they face a fundamental trade-off between the ability to maintain activity and an increased rate of dehydration at higher temperatures. Canopy coverage affects both the thermal and hydric conditions of the environment and can therefore influence amphibian thermoregulation. Frogs require proper conditions to thermoregulate to successfully grow, survive, and reproduce. But while we know how canopy and environmental variables typically affect operative temperature, less is known about effects on amphibian water loss rates. In this study, we measure the effect of canopy coverage on the conditions available for thermoregulation at a breeding pond of the California red-legged frog, Rana draytonii. We use agar frog models to estimate the thermal and hydric capacities frogs would experience in locations with different canopy coverage and microhabitats. At each site, we deployed models under four microhabitat treatments: wet/sun, wet/shade, dry/sun, and dry/shade. We modeled how environmental variables affected operative temperature and evaporative water loss from agar frogs. We found positive effects of air temperature, the sun treatment, and reduced canopy cover on operative temperature, and negative direct or indirect effects of these variables on evaporative water loss, consistent with the hypothesized trade-off between thermoregulatory behavior to increase temperature and the increased desiccation risk due to higher water loss. Additionally, our results indicate that the availability of wet microhabitats can allow frogs to reduce water loss, potentially mitigating the risk of desiccation when thermoregulating to achieve higher operative temperatures. Our findings suggest, that with access to proper microhabitats, amphibians can mitigate the fundamental trade-off and receive benefits of thermoregulating at high temperatures.

Glove decontamination procedures to prevent pathogen and DNA cross-contamination among frogs.

Working with aquatic organisms often requires handling multiple individuals in a single session, potentially resulting in cross-contamination by live pathogens or DNA. Most researchers address this problem by disposing of gloves between animals. However, this generates excessive waste and may be impractical for processing very slippery animals that might be easier to handle with cotton gloves. We tested methods to decontaminate cotton or nitrile gloves after contamination with cultured Batrachochytrium dendrobatidis (Bd) or after handling heavily Bd-infected Xenopus laevis with layered cotton and nitrile gloves. Bleach eliminated detectable Bd DNA from culture-contaminated nitrile gloves, but gloves retained detectable Bd DNA following ethanol disinfection. After handling a Bd-infected frog, Bd DNA contamination was greatly reduced by removal of the outer cotton glove, after which either bleach decontamination or ethanol decontamination followed by drying hands with a paper towel lowered Bd DNA below the detection threshold of our assay. These results provide new options to prevent pathogen or DNA cross-contamination, especially when handling slippery aquatic organisms. However, tradeoffs should be considered when selecting an animal handling procedure, such as the potential for cotton gloves to abrade amphibian skin or disrupt skin mucus. Disposing of gloves between animals should remain the gold standard for maintaining biosecurity in sensitive situations.

Large-scale spatial drivers of avian schistosomes in Northern Michigan inland lakes.

Avian schistosomes are snail-borne trematode parasites (Trichobilharzia spp.) that can cause a nasty skin rash in humans when their cercariae mistake us for their normal bird hosts. We sought to investigate drivers of the spatial distribution of Trichobilharzia cercaria abundance throughout Northern Michigan lakes. For 38 sites on 16 lakes, we assessed several dozen potential environmental predictors that we hypothesized might have direct or indirect effects on overall cercaria abundance, based on known relationships between abiotic and biotic factors in wetland ecosystems. We included variables quantifying local densities of intermediate hosts, temperature, periphyton growth rates, human land use and hydrology. We also measured daily abundance of schistosome cercariae in the water over a 5-week period, supported by community scientists who collected and preserved filtered water samples for qPCR. The strongest predictor of cercaria abundance was Lymnaea host snail density. Lymnaea density was higher in deeper lakes and at sites with more deciduous tree cover, consistent with their association with cool temperature habitats. Contrary to past studies of human schistosomes, we also found a significant negative relationship between cercaria abundance and submerged aquatic vegetation, possibly due to vegetation blocking cercaria movement from offshore snail beds. If future work shows that these effects are indeed causal, then these results suggest possible new approaches to managing swimmer's itch risk in northern MI lakes, such as modifying tree cover and shallow-water vegetation at local sites.

Using Metabolic Theory to Describe Temperature and Thermal Acclimation Effects on Parasitic Infection.

AbstractPredicting temperature effects on species interactions can be challenging, especially for parasitism, where it is difficult to experimentally separate host and parasite thermal performance curves. Prior authors proposed a possible solution based on the metabolic theory of ecology (MTE), using MTE-based equations to describe the thermal mismatch between host and parasite performance curves and account for thermal acclimation responses. Here, we use published infection data, supplemented with experiments measuring metabolic responses to temperature in each species, to show that this modeling framework can successfully describe thermal acclimation effects on two different stages of infection in a tadpole-trematode system. All thermal acclimation effects on host performance manifested as changes in one key model parameter (activation energy), with measurements of host respiration generating similar MTE parameter estimates and acclimation effects compared with measurements of the host's ability to clear encysted parasites. This result suggests that metabolic parameter estimates for whole-body metabolism can sometimes be used to estimate temperature effects on host and parasite performance curves. However, we found different thermal patterns for measurements of host prevention of initial parasite encystment emphasizing potential challenges when applying MTE-based models to complex parasite-host systems with multiple distinct stages of infection.

Pesticides alter ecosystem respiration via phytoplankton abundance and community structure: Effects on the carbon cycle?

Freshwater systems are critical to life on earth, yet they are threatened by the increasing rate of synthetic chemical pollution. Current predictions of the effects of synthetic chemicals on freshwater ecosystems are hampered by the sheer number of chemical contaminants entering aquatic systems, the diversity of organisms inhabiting these systems, the myriad possible direct and indirect effects resulting from these combinations, and uncertainties concerning how contaminants might alter ecosystem metabolism via changes in biodiversity. To address these knowledge gaps, we conducted a mesocosm experiment that elucidated the responses of ponds composed of phytoplankton and zooplankton to standardized concentrations of 12 pesticides, nested within four pesticide classes, and two pesticide types. We show that the effects of the pesticides on algae were consistent within herbicides and insecticides and that responses of over 70 phytoplankton species and genera were consistent within broad taxonomic groups. Insecticides generated top-down effects on phytoplankton community composition and abundance, which were associated with persistent increases in ecosystem respiration. Insecticides had direct toxic effects on cladocerans, which led to competitive release of copepods. These changes in the zooplankton community led to a decrease in green algae and a modest increase in diatoms. Herbicides did not change phytoplankton composition but reduced total phytoplankton abundance. This reduction in phytoplankton led to short-term decreases in ecosystem respiration. Given that ponds release atmospheric carbon and that worldwide pesticide pollution continues to increase exponentially, scientists and policy makers should pay more attention to the ways pesticides alter the carbon cycle in ponds via changes in communities, as demonstrated by our results. Our results show that these predictions can be simplified by grouping pesticides into types and species into functional groups. Adopting this approach provides an opportunity to improve the efficiency of risk assessment and mitigation responses to global change.

Hot and sour: parasite adaptations to honeybee body temperature and pH.

Host temperature and gut chemistry can shape resistance to parasite infection. Heat and acidity can limit trypanosomatid infection in warm-blooded hosts and could shape infection resistance in insects as well. The colony-level endothermy and acidic guts of social bees provide unique opportunities to study how temperature and acidity shape insect-parasite associations. We compared temperature and pH tolerance between three trypanosomatid parasites from social bees and a related trypanosomatid from poikilothermic mosquitoes, which have alkaline guts. Relative to the mosquito parasites, all three bee parasites had higher heat tolerance that reflected body temperatures of hosts. Heat tolerance of the honeybee parasite Crithidia mellificae was exceptional for its genus, implicating honeybee endothermy as a plausible filter of parasite establishment. The lesser heat tolerance of the emerging Lotmaria passim suggests possible spillover from a less endothermic host. Whereas both honeybee parasites tolerated the acidic pH found in bee intestines, mosquito parasites tolerated the alkaline conditions found in mosquito midguts, suggesting that both gut pH and temperature could structure host-parasite specificity. Elucidating how host temperature and gut pH affect infection-and corresponding parasite adaptations to these factors-could help explain trypanosomatids' distribution among insects and invasion of mammals.

Temperature and mass scaling affect cutaneous and pulmonary respiratory performance in a diving frog.

Global climate change is altering patterns of temperature variation, with unpredictable consequences for species and ecosystems. The Metabolic Theory of Ecology (MTE) provides a powerful framework for predicting climate change impacts on ectotherm metabolic performance. MTE postulates that physiological and ecological processes are limited by organism metabolic rates, which scale predictably with body mass and temperature. The purpose of this study was to determine if different metabolic proxies generate different empirical estimates of key MTE model parameters for the aquatic frog Xenopus laevis when allowed to exhibit normal diving behavior. We used a novel methodological approach in combining a flow-through respirometry setup with the open-source Arduino platform to measure mass and temperature effects on 4 different proxies for whole-body metabolism (total O2 consumption, cutaneous O2 consumption, pulmonary O2 consumption, and ventilation frequency), following thermal acclimation to one of 3 temperatures (8°C, 17°C, or 26°C). Different metabolic proxies generated different mass-scaling exponents (b) and activation energy (EA ) estimates, highlighting the importance of metabolic proxy selection when parameterizing MTE-derived models. Animals acclimated to 17°C had higher O2 consumption across all temperatures, but thermal acclimation did not influence estimates of key MTE parameters EA and b. Cutaneous respiration generated lower MTE parameters than pulmonary respiration, consistent with temperature and mass constraints on dissolved oxygen availability, SA:V ratios, and diffusion distances across skin. Our results show that the choice of metabolic proxy can have a big impact on empirical estimates for key MTE model parameters.

Consistent effects of pesticides on community structure and ecosystem function in freshwater systems.

Predicting ecological effects of contaminants remains challenging because of the sheer number of chemicals and their ambiguous role in biodiversity-ecosystem function relationships. We evaluate responses of experimental pond ecosystems to standardized concentrations of 12 pesticides, nested in four pesticide classes and two pesticide types. We show consistent effects of herbicides and insecticides on ecosystem function, and slightly less consistent effects on community composition. Effects of pesticides on ecosystem function are mediated by alterations in the abundance and community composition of functional groups. Through bottom-up effects, herbicides reduce respiration and primary productivity by decreasing the abundance of phytoplankton. The effects of insecticides on respiration and primary productivity of phytoplankton are driven by top-down effects on zooplankton composition and abundance, but not richness. By demonstrating consistent effects of pesticides on communities and ecosystem functions and linking pesticide-induced changes in functional groups of organisms to ecosystem functions, the study suggests that ecological risk assessment of registered chemicals could be simplified to synthetic chemical classes or types and groups of organisms with similar functions and chemical toxicities.

Caldicellulosiruptor bescii Adheres to Polysaccharides via a Type IV Pilin-Dependent Mechanism.

Biological hydrolysis of cellulose above 70°C involves microorganisms that secrete free enzymes and deploy separate protein systems to adhere to their substrate. Strongly cellulolytic Caldicellulosiruptor bescii is one such extreme thermophile, which deploys modular, multifunctional carbohydrate-acting enzymes to deconstruct plant biomass. Additionally, C. bescii also encodes noncatalytic carbohydrate binding proteins, which likely evolved as a mechanism to compete against other heterotrophs in carbon-limited biotopes that these bacteria inhabit. Analysis of the Caldicellulosiruptor pangenome identified a type IV pilus (T4P) locus encoded upstream of the tapirins, that is encoded by all Caldicellulosiruptor species. In this study, we sought to determine if the C. bescii T4P plays a role in attachment to plant polysaccharides. The major C. bescii pilin (CbPilA) was identified by the presence of pilin-like protein domains, paired with transcriptomics and proteomics data. Using immuno-dot blots, we determined that the plant polysaccharide xylan induced production of CbPilA 10- to 14-fold higher than glucomannan or xylose. Furthermore, we are able to demonstrate that recombinant CbPilA directly interacts with xylan and cellulose at elevated temperatures. Localization of CbPilA at the cell surface was confirmed by immunofluorescence microscopy. Lastly, a direct role for CbPilA in cell adhesion was demonstrated using recombinant CbPilA or anti-CbPilA antibodies to reduce C. bescii cell adhesion to xylan and crystalline cellulose up to 4.5- and 2-fold, respectively. Based on these observations, we propose that CbPilA and, by extension, the T4P play a role in Caldicellulosiruptor cell attachment to plant biomass.IMPORTANCE Most microorganisms are capable of attaching to surfaces in order to persist in their environment. Type IV (T4) pili produced by certain mesophilic Firmicutes promote adherence; however, a role for T4 pili encoded by thermophilic members of this phylum has yet to be demonstrated. Prior comparative genomics analyses identified a T4 pilus locus possessed by an extremely thermophilic genus within the Firmicutes Here, we demonstrate that attachment to plant biomass-related carbohydrates by strongly cellulolytic Caldicellulosiruptor bescii is mediated by T4 pilins. Surprisingly, xylan but not cellulose induced expression of the major T4 pilin. Regardless, the C. bescii T4 pilin interacts with both polysaccharides at high temperatures and is located to the cell surface, where it is directly involved in C. bescii attachment. Adherence to polysaccharides is likely key to survival in environments where carbon sources are limiting, allowing C. bescii to compete against other plant-degrading microorganisms.

Direct onshore wind predicts daily swimmer's itch (avian schistosome) incidence at a Michigan beach.

Swimmer's itch (SI) is a painful rash caused by skin penetration by free-swimming infectious cercariae of avian schistosomes, snail-borne helminth parasites related to the causative agents of human schistosomiasis. The goal of this study was to determine if commonly collected environmental data could be used to predict daily fluctuations in SI incidence at an inland beach in northwestern Michigan. Lifeguards collected daily data over four summers, including the number of self-reported SI cases, total swimmers, water temperature, wind speed and wind direction. Mixed-effects binomial regression revealed that wind direction, wind speed and time of day were the best predictors of daily SI risk. Swimmers entering the water in the morning or on days with direct onshore wind perpendicular to the shoreline had the greatest SI risk. However, there was a negative effect of wind speed after accounting for direction, where SI risk was greatest on days with a gentle breeze originating directly offshore. These results suggest that at this beach, direct onshore winds generate a surface-water current that causes SI cercariae to aggregate in the shallow waters used by swimmers. Data are needed from additional sites to confirm whether the onshore wind is a generally important driver of SI incidence.

Temperature dependence of parasitic infection and gut bacterial communities in bumble bees.

High temperatures (e.g., fever) and gut microbiota can both influence host resistance to infection. However, effects of temperature-driven changes in gut microbiota on resistance to parasites remain unexplored. We examined the temperature dependence of infection and gut bacterial communities in bumble bees infected with the trypanosomatid parasite Crithidia bombi. Infection intensity decreased by over 80% between 21 and 37°C. Temperatures of peak infection were lower than predicted based on parasite growth in vitro, consistent with mismatches in thermal performance curves of hosts, parasites and gut symbionts. Gut bacterial community size and composition exhibited slight but significant, non-linear, and taxon-specific responses to temperature. Abundance of total gut bacteria and of Orbaceae, both negatively correlated with infection in previous studies, were positively correlated with infection here. Prevalence of the bee pathogen-containing family Enterobacteriaceae declined with temperature, suggesting that high temperature may confer protection against diverse gut pathogens. Our results indicate that resistance to infection reflects not only the temperature dependence of host and parasite performance, but also temperature-dependent activity of gut bacteria. The thermal ecology of gut parasite-symbiont interactions may be broadly relevant to infectious disease, both in ectothermic organisms that inhabit changing climates, and in endotherms that exhibit fever-based immunity.

Shifts in temperature influence how Batrachochytrium dendrobatidis infects amphibian larvae.

Many climate change models predict increases in frequency and magnitude of temperature fluctuations that might impact how ectotherms are affected by disease. Shifts in temperature might especially affect amphibians, a group with populations that have been challenged by several pathogens. Because amphibian hosts invest more in immunity at warmer than cooler temperatures and parasites might acclimate to temperature shifts faster than hosts (creating lags in optimal host immunity), researchers have hypothesized that a temperature shift from cold-to-warm might result in increased amphibian sensitivity to pathogens, whereas a shift from warm-to-cold might result in decreased sensitivity. Support for components of this climate-variability based hypothesis have been provided by prior studies of the fungus Batrachochytrium dendrobatidis (Bd) that causes the disease chytridiomycosis in amphibians. We experimentally tested whether temperature shifts before exposure to Batrachochytrium dendrobatidis (Bd) alters susceptibility to the disease chytridiomycosis in the larval stage of two amphibian species-western toads (Anaxyrus boreas) and northern red legged frogs (Rana aurora). Both host species harbored elevated Bd infection intensities under constant cold (15° C) temperature in comparison to constant warm (20° C) temperature. Additionally, both species experienced an increase in Bd infection abundance after shifted from 15° C to 20° C, compared to a constant 20° C but they experienced a decrease in Bd after shifted from 20° C to 15° C, compared to a constant 15° C. These results are in contrast to prior studies of adult amphibians highlighting the potential for species and stage differences in the temperature-dependence of chytridiomycosis.

Thermal acclimation has little effect on tadpole resistance to Batrachochytrium dendrobatidis.

Given that climate change is predicted to alter patterns of temperature variability, it is important to understand how shifting temperatures might influence species interactions, including parasitism. Predicting thermal effects on species interactions is complicated, however, because the temperature-dependence of the interaction depends on the thermal responses of both interacting organisms, which can also be influenced by thermal acclimation, a process by which organisms adjust their physiologies in response to a temperature change. We tested for thermal acclimation effects on Lithobates clamitans tadpole susceptibility to the fungus Batrachochytrium dendrobatidis (Bd) by acclimating tadpoles to 1 of 3 temperatures, moving them to 1 of 5 performance temperatures at which we exposed them to Bd, and measuring Bd loads on tadpoles post-exposure. We predicted that (1) tadpole Bd load would peak at a lower temperature than the temperature for peak Bd growth in culture, and (2) tadpoles acclimated to intermediate temperatures would have overall lower Bd loads across performance temperatures than cold- or warm-acclimated tadpoles, similar to a previously published pattern describing tadpole resistance to trematode metacercariae. Consistent with our first prediction, Bd load on tadpoles decreased with increasing performance temperature. However, we found only weak support for our second prediction, as acclimation temperature had little effect on tadpole Bd load. Our results contribute to a growing body of work investigating thermal responses of hosts and parasites, which will aid in developing methods to predict the temperature-dependence of disease.

Effects of pesticides on exposure and susceptibility to parasites can be generalised to pesticide class and type in aquatic communities.

Pesticide pollution can alter parasite transmission, but scientists are unaware if effects of pesticides on parasite exposure and host susceptibility (i.e. infection risk given exposure) can be generalised within a community context. Using replicated temperate pond communities, we evaluate effects of 12 pesticides, nested in four pesticide classes (chloroacetanilides, triazines, carbamates organophosphates) and two pesticide types (herbicides, insecticides) applied at standardised environmental concentrations on larval amphibian exposure and susceptibility to trematode parasites. Most of the variation in exposure and susceptibility occurred at the level of pesticide class and type, not individual compounds. The organophosphate class of insecticides increased snail abundance (first intermediate host) and thus trematode exposure by increasing mortality of snail predators (top-down mechanism). While a similar pattern in snail abundance and trematode exposure was observed with triazine herbicides, this effect was driven by increases in snail resources (periphytic algae, bottom-up mechanism). Additionally, herbicides indirectly increased host susceptibility and trematode infections by (1) increasing time spent in susceptible early developmental stages and (2) suppressing tadpole immunity. Understanding generalisable effects associated with contaminant class and type on transmission is critical in reducing complexities in predicting disease dynamics in at-risk host populations.

pH-mediated inhibition of a bumble bee parasite by an intestinal symbiont.

Gut symbionts can augment resistance to pathogens by stimulating host-immune responses, competing for space and nutrients, or producing antimicrobial metabolites. Gut microbiota of social bees, which pollinate many crops and wildflowers, protect hosts against diverse infections and might counteract pathogen-related bee declines. Bumble bee gut microbiota, and specifically abundance of Lactobacillus 'Firm-5' bacteria, can enhance resistance to the trypanosomatid parasite Crithidia bombi. However, the mechanism underlying this effect remains unknown. We hypothesized that the Firm-5 bacterium Lactobacillus bombicola, which produces lactic acid, inhibits C. bombi via pH-mediated effects. Consistent with our hypothesis, L. bombicola spent medium inhibited C. bombi growth via reduction in pH that was both necessary and sufficient for inhibition. Inhibition of all parasite strains occurred within the pH range documented in honey bees, though sensitivity to acidity varied among strains. Spent medium was slightly more potent than HCl, d- and l-lactic acids for a given pH, suggesting that other metabolites also contribute to inhibition. Results implicate symbiont-mediated reduction in gut pH as a key determinant of trypanosomatid infection in bees. Future investigation into in vivo effects of gut microbiota on pH and infection intensity would test the relevance of these findings for bees threatened by trypanosomatids.

Temperature-mediated inhibition of a bumblebee parasite by an intestinal symbiont.

Competition between organisms is often mediated by environmental factors, including temperature. In animal intestines, nonpathogenic symbionts compete physically and chemically against pathogens, with consequences for host infection. We used metabolic theory-based models to characterize differential responses to temperature of a bacterial symbiont and a co-occurring trypanosomatid parasite of bumblebees, which regulate body temperature during flight and incubation. We hypothesized that inhibition of parasites by bacterial symbionts would increase with temperature, due to symbionts having higher optimal growth temperatures than parasites. We found that a temperature increase over the range measured in bumblebee colonies would favour symbionts over parasites. As predicted by our hypothesis, symbionts reduced the optimal growth temperature for parasites, both in direct competition and when parasites were exposed to symbiont spent medium. Inhibitory effects of the symbiont increased with temperature, reflecting accelerated growth and acid production by symbionts. Our results indicate that high temperatures, whether due to host endothermy or environmental factors, can enhance the inhibitory effects of symbionts on parasites. Temperature-modulated manipulation of microbiota could be one explanation for fever- and heat-induced reductions of infection in animals, with consequences for diseases of medical and conservation concern.

Agrochemicals increase risk of human schistosomiasis by supporting higher densities of intermediate hosts.

Schistosomiasis is a snail-borne parasitic disease that ranks among the most important water-based diseases of humans in developing countries. Increased prevalence and spread of human schistosomiasis to non-endemic areas has been consistently linked with water resource management related to agricultural expansion. However, the role of agrochemical pollution in human schistosome transmission remains unexplored, despite strong evidence of agrochemicals increasing snail-borne diseases of wildlife and a projected 2- to 5-fold increase in global agrochemical use by 2050. Using a field mesocosm experiment, we show that environmentally relevant concentrations of fertilizer, a herbicide, and an insecticide, individually and as mixtures, increase densities of schistosome-infected snails by increasing the algae snails eat and decreasing densities of snail predators. Epidemiological models indicate that these agrochemical effects can increase transmission of schistosomes. Identifying agricultural practices or agrochemicals that minimize disease risk will be critical to meeting growing food demands while improving human wellbeing.

Thermal Performance Curves and the Metabolic Theory of Ecology-A Practical Guide to Models and Experiments for Parasitologists.

Climate change will affect host-parasite dynamics in complex ways. The development of forecast models is necessary for proactive disease management, but past studies have frequently reported thermal performance data in idiosyncratic ways that have limited use for parameterizing thermal host-parasite models. Development of improved forecast models will require strong collaborations between experimental parasitologists and disease modelers. The purpose of this article is to facilitate such collaborations by reviewing practical considerations for describing thermal performance curves of parasite and host performance traits, and using them to predict climate change impacts on host-parasite systems. In the first section, we provide an overview of how thermal performance curves can be embedded in life-cycle-based dynamical models of parasitism, and we outline how such models can capture the net effect of multiple nonlinear temperature dependencies affecting the host-parasite dynamics. We also discuss how macroecological generalities based on the metabolic theory of ecology (MTE) can be used to determine a priori parameter estimates for thermal performance curves to derive null models for data-deficient species, but we note that most of the generalities suggested by MTE remain to be tested for parasites. In the second section, we discuss empirical knowledge gaps for the temperature dependence of parasite and host performance traits, and we outline the types of data that need to be collected to inform MTE-based models for data-deficient species. We specifically emphasize the importance of (1) capturing the entire thermal response of performance traits, including lower and upper temperature thresholds, and (2) experimentally or statistically separating out the thermal responses of different performance traits (e.g., development and mortality) rather than only reporting composite measures (e.g., apparent development). Not adhering to these principles can lead to biased climate change impact predictions. In the third section, we provide a practical guide outlining how experimentalists can contribute to fill data gaps by measuring the temperature dependence of host and parasite performance traits in ways that are systematic, statistically rigorous, and consistent with the requirements of life cycle-based host-parasite models. This guide includes recommendations and practical examples illustrating (1) the use of perturbation analyses to determine experimental priorities, (2) experimental design tips for quantifying thermal response curves, and (3) statistical methods for estimating the parameters of thermal performance curves. Our hope is that this article helps researchers to maximize the value and use of future data collections for both empirical and modelling studies investigating the way in which temperature influences parasitism.

Bottom-up and trait-mediated effects of resource quality on amphibian parasitism.

Leaf litter subsidies are important resources for aquatic consumers like tadpoles and snails, causing bottom-up effects on wetland ecosystems. Recent studies have shown that variation in litter nutritional quality can be as important as litter quantity in driving these bottom-up effects. Resource subsidies likely also have indirect and trait-mediated effects on predation and parasitism, but these potential effects remain largely unexplored. We generated predictions for differential effects of litter nutrition and secondary polyphenolic compounds on tadpole (Lithobates sylvatica) exposure and susceptibility to Ribeiroia ondatrae, based on ecological stoichiometry and community-ecology theory. We predicted direct and indirect effects on key traits of the tadpole host (rates of growth, development and survival), the trematode parasite (production of the cercaria infective stages) and the parasite's snail intermediate host (growth and reproduction). To test these predictions, we conducted a large-scale mesocosm experiment using a natural gradient in the concentrations of nutrients (nitrogen) and toxic secondary compounds (polyphenolics) of nine leaf litter species. To differentiate between effects on exposure vs. susceptibility to infection, we included multiple infection experiments including one with constant per capita exposure. We found that increased litter nitrogen increased tadpole survival, and also increased cercaria production by the snail intermediate hosts, causing opposing effects on tadpole per capita exposure to trematode infection. Increased litter polyphenolics slowed tadpole development, leading to increased infection by increasing both their susceptibility to infection and the length of time they were exposed to parasites. Based on these results, recent shifts in forest composition towards more nitrogen-poor litter species should decrease trematode infection in tadpoles via density- and trait-mediated effects on the snail intermediate hosts. However, these shifts also involve increased abundance of litter species with high polyphenolic levels, which should increase trematode infection via trait-mediated effects on tadpoles. Future studies will be needed to determine the relative strength of these opposing effects in natural wetland communities. [Correction added after online publication on 5 January 2017: wording changed to 'which should increase trematode infection via trait-mediated effects on tadpoles'.].