Weak evidence of density dependent population regulation when using the ability of two simple density dependent models to predict population size.

The relative importance of density dependence regulation in natural population fluctuations has long been debated. The concept of density dependence implies that current abundance is determined by historical abundance. We have developed four models-two density dependent and two density independent-to predict population size one year beyond the training set and used predictive performance on more than 16,000 populations from 14 datasets to compare the understanding captured by those models. For 4 of 14 datasets the density dependent models make better predictions (i.e., density dependent regulated) than either of the density independent models. However, neither of the density dependent models is statistically significantly superior to density independent models for any of the 14 datasets. We conclude that the evidence for widespread density dependent population regulation in the forms represented by these two simple density-dependent models is weak. However, the density dependent models used here-the Logistic and Gompertz models-are simple representations of how population density might regulate natural populations and only examine density-dependent effects on population size. A comprehensive assessment of the relative importance of density-dependent population regulation will require testing the predictive ability of a wider range of density-dependent models including models examining effects on population characteristics other than population size.

Compensatory indirect effects of an herbicide on wetland communities.

The direct effects of large-scale disturbances are readily studied because their effects are often apparent and result in large changes to ecosystems. Direct effects can cascade through the ecosystem, leading to indirect effects that are often subtle and difficult to detect. Managing anthropogenic disturbances, such as chemical contamination, requires an understanding of both direct and indirect effects to predict, measure, and characterize the impact. Using a replicated whole-ecosystem experiment and path analyses (assesses the effects of a set of variables on a specified outcome, similar to multiple regression), we examined the direct and indirect effects of a glyphosate-based herbicide and nutrient enrichment on wetland communities. The latter did not impact any measured endpoints. The strongest drivers of macrophyte, benthic invertebrate, and amphibian assemblages were the ephemerality and the size of wetlands, factors which were not altered by herbicide applications. The herbicide had a direct negative effect on macrophyte cover, amphibian larval abundance, and the proportion of predatory benthic invertebrates. However, both amphibians and invertebrates were positively affected by the reduction in the macrophyte cover caused by the herbicide applications. The opposing directions of the direct and indirect effects lead to no net change in either group. The compensatory dynamics observed herein highlight the need for a better understanding of indirect effects pathways to determine whether common anthropogenic disturbances alter the ecological communities in small wetland ecosystems.

Designing better frog call recognition models.

Advances in bioacoustic technology, such as the use of automatic recording devices, allow wildlife monitoring at large spatial scales. However, such technology can produce enormous amounts of audio data that must be processed and analyzed. One potential solution to this problem is the use of automated sound recognition tools, but we lack a general framework for developing and validating these tools. Recognizers are computer models of an animal sound assembled from "training data" (i.e., actual samples of vocalizations). The settings of variables used to create recognizers can impact performance, and the use of different settings can result in large differences in error rates that can be exploited for different monitoring objectives. We used Song Scope (Wildlife Acoustics Inc.) to build recognizers and vocalizations of the wood frog (Lithobates sylvaticus) to test how different settings and amounts of training data influence recognizer performance. Performance was evaluated using precision (the probability of a recognizer match being a true match) and sensitivity (the proportion of vocalizations detected) based on a receiver operating characteristic (ROC) curve-determined score threshold. Evaluations were conducted using recordings not used to build the recognizer. Wood frog recognizer performance was sensitive to setting changes in four out of nine variables, and small improvements were achieved by using additional training data from different sites and from the same recording, but not from different recordings from the same site. Overall, the effect of changes to variable settings was much greater than the effect of increasing training data. Additionally, by testing the performance of the recognizer on vocalizations not used to build the recognizer, we discovered that Type I error rates appear idiosyncratic and do not recommend extrapolation from training to new data, whereas Type II errors showed more consistency and extrapolation can be justified. Optimizing variable settings on independent recordings led to a better match between recognizer performance and monitoring objectives. We provide general recommendations for application of this methodology with other species and make some suggestions for improvements.

General and histological indicators of health in wild fishes from a biological mercury hotspot in northeastern North America.

Kejimkujik National Park and National Historic Site, Nova Scotia, Canada, is considered a biological mercury (Hg) hotspot because the tissues of yellow perch (Perca flavescens) and common loons (Gavia immer) inhabiting the lakes frequently exceed so-called safe levels of Hg. In the present study, the relationships between Hg and overall health of males and females of 3 forage fish species (brown bullhead Ameirus nebulosus, banded killifish Fundulus diaphanus, and golden shiner Notemigonus crysoleucas; n = 6-18/sex/lake) in 6 lakes at the park were assessed using condition factor, liversomatic index (LSI), and macrophage aggregates (MAs; indicators of tissue damage). Mean muscle total Hg (THg) concentrations of brown bullhead, banded killifish, and golden shiner across lakes were 0.32 µg/g, 0.27 µg/g, and 0.34 µg/g, respectively. Condition was negatively related to muscle THg in golden shiner and banded killifish, LSI was not related to THg in any species, and all species showed evidence of increasing MA prevalence (counts and area) with increasing THg concentrations. The MAs were most prevalent in spleen tissues of golden shiner, with mean percentage cover ranging from 0.36% to 5.59% across lakes. In addition, the area of MAs appeared to be better predicted by THg concentration than was the number of MAs in the same tissue. These findings suggest that Hg is affecting the health of wild fishes in Kejimkujik National Park and National Historic Site and that other populations with similar or higher concentrations of this metal may also be at risk. Environ Toxicol Chem 2017;36:976-987. © 2016 SETAC.

How Effective Is Road Mitigation at Reducing Road-Kill? A Meta-Analysis.

Road traffic kills hundreds of millions of animals every year, posing a critical threat to the populations of many species. To address this problem there are more than forty types of road mitigation measures available that aim to reduce wildlife mortality on roads (road-kill). For road planners, deciding on what mitigation method to use has been problematic because there is little good information about the relative effectiveness of these measures in reducing road-kill, and the costs of these measures vary greatly. We conducted a meta-analysis using data from 50 studies that quantified the relationship between road-kill and a mitigation measure designed to reduce road-kill. Overall, mitigation measures reduce road-kill by 40% compared to controls. Fences, with or without crossing structures, reduce road-kill by 54%. We found no detectable effect on road-kill of crossing structures without fencing. We found that comparatively expensive mitigation measures reduce large mammal road-kill much more than inexpensive measures. For example, the combination of fencing and crossing structures led to an 83% reduction in road-kill of large mammals, compared to a 57% reduction for animal detection systems, and only a 1% for wildlife reflectors. We suggest that inexpensive measures such as reflectors should not be used until and unless their effectiveness is tested using a high-quality experimental approach. Our meta-analysis also highlights the fact that there are insufficient data to answer many of the most pressing questions that road planners ask about the effectiveness of road mitigation measures, such as whether other less common mitigation measures (e.g., measures to reduce traffic volume and/or speed) reduce road mortality, or to what extent the attributes of crossing structures and fences influence their effectiveness. To improve evaluations of mitigation effectiveness, studies should incorporate data collection before the mitigation is applied, and we recommend a minimum study duration of four years for Before-After, and a minimum of either four years or four sites for Before-After-Control-Impact designs.

The combined influence of two agricultural contaminants on natural communities of phytoplankton and zooplankton.

Concentrations of glyphosate observed in the environment are generally lower than those found to exert toxicity on aquatic organisms in the laboratory. Toxicity is often tested in the absence of other expected co-occurring contaminants. By examining changes in the phytoplankton and zooplankton communities of shallow, partitioned wetlands over a 5 month period, we assessed the potential for direct and indirect effects of the glyphosate-based herbicide, Roundup WeatherMax(©) applied at the maximum label rate, both in isolation and in a mixture with nutrients (from fertilizers). The co-application of herbicide and nutrients resulted in an immediate but transient decline in dietary quality of phytoplankton (8.3 % decline in edible carbon content/L) and zooplankton community similarity (27 % decline in similarity and loss of three taxa), whereas these effects were not evident in wetlands treated only with the herbicide. Thus, even at a worst-case exposure, this herbicide in isolation, did not produce the acutely toxic effects on plankton communities suggested by laboratory or mesocosm studies. Indirect effects of the herbicide-nutrient mixture were evident in mid-summer, when glyphosate residues were no longer detectable in surface water. Zooplankton abundance tripled, and zooplankton taxa richness increased by an average of four taxa in the herbicide and nutrient treated wetlands. The lack of significant toxicity of Roundup WeatherMax alone, as well as the observation of delayed interactive or indirect effects of the mixture of herbicide and nutrients attest to the value of manipulative field experiments as part of a comprehensive, tiered approach to risk assessments in ecotoxicology.

Experimental study designs to improve the evaluation of road mitigation measures for wildlife.

An experimental approach to road mitigation that maximizes inferential power is essential to ensure that mitigation is both ecologically-effective and cost-effective. Here, we set out the need for and standards of using an experimental approach to road mitigation, in order to improve knowledge of the influence of mitigation measures on wildlife populations. We point out two key areas that need to be considered when conducting mitigation experiments. First, researchers need to get involved at the earliest stage of the road or mitigation project to ensure the necessary planning and funds are available for conducting a high quality experiment. Second, experimentation will generate new knowledge about the parameters that influence mitigation effectiveness, which ultimately allows better prediction for future road mitigation projects. We identify seven key questions about mitigation structures (i.e., wildlife crossing structures and fencing) that remain largely or entirely unanswered at the population-level: (1) Does a given crossing structure work? What type and size of crossing structures should we use? (2) How many crossing structures should we build? (3) Is it more effective to install a small number of large-sized crossing structures or a large number of small-sized crossing structures? (4) How much barrier fencing is needed for a given length of road? (5) Do we need funnel fencing to lead animals to crossing structures, and how long does such fencing have to be? (6) How should we manage/manipulate the environment in the area around the crossing structures and fencing? (7) Where should we place crossing structures and barrier fencing? We provide experimental approaches to answering each of them using example Before-After-Control-Impact (BACI) study designs for two stages in the road/mitigation project where researchers may become involved: (1) at the beginning of a road/mitigation project, and (2) after the mitigation has been constructed; highlighting real case studies when available.

The response of amphibian larvae to exposure to a glyphosate-based herbicide (Roundup WeatherMax) and nutrient enrichment in an ecosystem experiment.

Herbicides and fertilizers are widely used throughout the world and pose a threat to aquatic ecosystems. Using a replicated, whole ecosystem experiment in which 24 small wetlands were split in half with an impermeable barrier we tested whether exposure to a glyphosate-based herbicide, Roundup WeatherMax™, alone or in combination with nutrient enrichment has an effect on the survival, growth or development of amphibians. The herbicide was applied at one of two concentrations (low=210 µg a.e./L, high=2880 µg a.e./L) alone and in combination with nutrient enrichment to one side of wetlands and the other was left as an untreated control. Each treatment was replicated with six wetlands, and the experiment was repeated over two years. In the high glyphosate and nutrient enrichment treatment the survival of wood frog (Lithobates sylvaticus) larvae was lower in enclosures placed in situ on the treated sides than the control sides of wetlands. However, these results were not replicated in the second year of study and they were not observed in free swimming wood frog larvae in the wetlands. In all treatments, wood frog larvae on the treated sides of wetlands were slightly larger (<10%) than those on the control side, but no effect on development was observed. The most dramatic finding was that the abundance of green frog larvae (Lithobates clamitans) was higher on the treated sides than the control sides of wetlands in the herbicide and nutrient treatments during the second year of the study. The results observed in this field study indicate that caution is necessary when extrapolating results from artificial systems to predict effects in natural systems. In this experiment, the lack of toxicity to amphibian larvae was probably due to the fact the pH of the wetlands was relatively low and the presence of sediments and organic surfaces which would have mitigated the exposure duration.

Variation in amphibian response to two formulations of glyphosate-based herbicides.

Variation in toxicity among formulations and species makes it difficult to extrapolate results to all species and all formulations of herbicides. The authors exposed larval wood frogs (Lithobates sylvaticus) from 4 populations to 2 glyphosate-based herbicides, Roundup Weed and Grass Control® and Roundup WeatherMax®. The 96-h median lethal concentration values for both formulations varied among the populations (Roundup Weed and Grass Control, 0.14 mg acid equivalents (a.e.)/L to 1.10 mg a.e./L; Roundup WeatherMax, 4.94 mg a.e./L to 8.26 mg a.e./L), demonstrating that toxicity varies among the formulations and that susceptibility may differ among populations.

The direct and indirect effects of a glyphosate-based herbicide and nutrients on Chironomidae (Diptera) emerging from small wetlands.

Laboratory and mesocosm experiments have demonstrated that some glyphosate-based herbicides can have negative effects on benthic invertebrate species. Although these herbicides are among the most widely used in agriculture, there have been few multiple-stressor, natural system-based investigations of the impacts of glyphosate-based herbicides in combination with fertilizers on the emergence patterns of chironomids from wetlands. Using a replicated, split-wetland experiment, the authors examined the effects of 2 nominal concentrations (2.88 mg acid equivalents/L and 0.21 mg acid equivalents/L) of the glyphosate herbicide Roundup WeatherMax, alone or in combination with nutrient additions, on the emergence of Chironomidae (Diptera) before and after herbicide-induced damage to macrophytes. There were no direct effects of treatment on the structure of the Chironomidae community or on the overall emergence rates. However, after macrophyte cover declined as a result of herbicide application, there were statistically significant increases in emergence in all but the highest herbicide treatment, which had also received no nutrients. There was a negative relationship between chironomid abundance and macrophyte cover on the treated sides of wetlands. Fertilizer application did not appear to compound the effects of the herbicide treatments. Although direct toxicity of Roundup WeatherMax was not apparent, the authors observed longer-term impacts, suggesting that the indirect effects of this herbicide deserve more consideration when assessing the ecological risk of using herbicides in proximity to wetlands.

Assessing gene network stability and individual variability in the fathead minnow (Pimephales promelas) transcriptome.

Transcriptomics is increasingly used to assess biological responses to environmental stimuli and stressors such as aquatic pollutants. However, fundamental studies characterizing individual variability in mRNA levels are lacking, which currently limits the use of transcriptomics in environmental monitoring assessments. To address individual variability in transcript abundance, we performed a meta-analysis on 231 microarrays that were conducted in the fathead minnow (FHM), a widely used toxicological model. The mean variability for gene probes was ranked from most to least variable based upon the coefficient of variation. Transcripts that were the most variable in individual tissues included NADH dehydrogenase flavoprotein 1, GTPase IMAP family member 7-like and v-set domain-containing T-cell activation inhibitor 1-like while genes encoding ribosomal proteins (rpl24 and rpl36), basic transcription factor 3, and nascent polypeptide-associated complex alpha subunit were the least variable in individuals across a range of microarray experiments. Gene networks that showed high variability (based upon the variation in expression of individual members within the network) included cell proliferation, metabolism (steroid, lipids, and glucose), cell adhesion, vascularization, and regeneration while those that showed low variability (more stability) included mRNA and rRNA processing, regulation of translational fidelity, RNA splicing, and ribosome biogenesis. Real-time PCR was conducted on a subset of genes for comparison of variability collected from the microarrays. There was a significant positive relationship between the two methods when measuring individual variability, suggesting that variability detected in microarray data can be used to guide decisions on sample sizes for measuring transcripts in real-time PCR experiments. A power analysis revealed that measuring estrogen receptor ba (esrba) requires fewer biological replicates than that of estrogen receptor bb (esrbb) in the gonad and samples sizes required to detect a 50% change for reproductive-related transcripts is between 12 and 20. Characterizing individual variability at the molecular level will prove necessary as efforts are made toward integrating molecular tools into environmental risk assessments.

Laboratory and field exposure of two species of juvenile amphibians to a glyphosate-based herbicide and Batrachochytrium dendrobatidis.

Herbicides are commonly used in agriculture and silviculture to reduce interspecific competition among plants and thereby enhance crop growth, quality, and volume. Internationally, formulations of glyphosate-based herbicides are the most widely used herbicides in both these sectors. A large amount of work has focused on the effects of these herbicides on amphibians. Several laboratory and mesocosm studies have demonstrated that various formulations of glyphosate herbicides can be acutely toxic to larval and juvenile amphibians at concentrations at the upper end of environmental realism. However, to date there has been little work done investigating such effects in natural systems, limited work on juvenile amphibians, and only a few studies have investigated interactions with other stressors. We conducted a 16 day field experiment in which juveniles of two amphibian species (Lithobates clamitans and Lithobates pipiens) were exposed to the herbicide Roundup WeatherMax™ at four application rates (0, 2.16, 4.32 and 8.64 kg a.e./ha) to investigate effects on survival, liver somatic index (LSI), body condition, and incidence of disease caused by Batrachochytrium dendrobatidis (Bd). In a separate 16 day laboratory experiment, we exposed juvenile L. clamitans to both the herbicide and Bd. Results of our studies showed that this particular herbicide formulation had no effect on juvenile survival, LSI, body condition, or disease incidence, nor was there an interaction between exposure to herbicide and exposure to the disease in tests which closely mimic real world exposure scenarios. These experiments suggest that Roundup WeatherMax as typically used in agriculture is unlikely to cause significant deleterious effects on juvenile amphibians under real world exposure conditions.

Optimizing α for better statistical decisions: a case study involving the pace-of-life syndrome hypothesis: optimal α levels set to minimize Type I and II errors frequently result in different conclusions from those using α = 0.05.

Setting optimal significance levels that minimize Type I and Type II errors allows for more transparent and well-considered statistical decision making compared to the traditional α = 0.05 significance level. We use the optimal α approach to re-assess conclusions reached by three recently published tests of the pace-of-life syndrome hypothesis, which attempts to unify occurrences of different physiological, behavioral, and life history characteristics under one theory, over different scales of biological organization. While some of the conclusions reached using optimal α were consistent to those previously reported using the traditional α = 0.05 threshold, opposing conclusions were also frequently reached. The optimal α approach reduced probabilities of Type I and Type II errors, and ensured statistical significance was associated with biological relevance. Biologists should seriously consider their choice of α when conducting null hypothesis significance tests, as there are serious disadvantages with consistent reliance on the traditional but arbitrary α = 0.05 significance level.

Negative consequences of using α = 0.05 for environmental monitoring decisions: a case study from a decade of Canada's Environmental Effects Monitoring Program.

Using the traditional α = 0.05 significance level for null hypothesis significance tests makes assumptions about relative costs of Type I vs relevant Type II errors and inflates their combined probabilities. We have examined the results of 1254 monitoring tests conducted under the Canadian Environmental Effects Monitoring (EEM) program from 1992 to 2003, focusing on how the choice of α affected the relative probabilities and implied costs of Type I and Type II errors. Using α = 0.05 resulted in implied relative costs of Type I vs Type II errors that were both inconsistent among monitoring end points and also inconsistent with the philosophy of the monitoring program. Using α = 0.05 also resulted in combinations of Type I and II error that were 15-17% larger than those for "optimal" α levels set to minimize Type I and II errors for each study, and 12% of all monitoring tests would have reached opposite conclusions had they used these optimal α levels for decision-making. Thus, if the Canadian EEM program used study-specific optimal α levels, they would reduce the incidence of relevant errors and eliminate inconsistent implied relative costs of these errors. Environmental research and monitoring programs using α = 0.05 as a decision-making threshold should re-evaluate the usefulness of this "one-size-fits-all" approach.

A silviculture application of the glyphosate-based herbicide VisionMAX to wetlands has limited direct effects on amphibian larvae.

Herbicides are commonly used in agriculture and silviculture to reduce interspecific competition among plants and thereby enhance crop growth, quality, and volume. Internationally, glyphosate-based herbicides are the most widely used herbicides in both of these sectors. Laboratory and mesocosm studies have demonstrated that some formulations are toxic to amphibian larvae below concentrations that approximate predicted maximal or "worst-case" exposure scenarios. However, field studies have not found evidence of toxicity at these concentrations. The authors conducted a replicated field experiment involving 10 naturalized wetlands split in half with an impermeable plastic barrier to assess the direct toxicity of a glyphosate formulation commonly used in silviculture (VisionMAX™). The herbicide formulation was applied directly to the surface of one side of each wetland at one of two target aqueous exposure rates (high = 2,880, low = 550 µg acid equivalents [a.e.]/L), and the other side was left as an untreated control. The survival and growth of green frog larvae (Lithobates clamitans) were assessed for two years following herbicide treatment. The herbicide did not have a negative impact on survival or growth of L. clamitans larvae at either treatment level. In fact, mean larval abundance was typically greater in the treated sides than in control sides within the year of herbicide application. These results indicate that typical silviculture use of VisionMAX poses negligible risk to larval amphibians, likely because the combined effects of sorption and degradation in natural wetlands limit the exposure magnitude and duration.

Setting an optimal α that minimizes errors in null hypothesis significance tests.

Null hypothesis significance testing has been under attack in recent years, partly owing to the arbitrary nature of setting α (the decision-making threshold and probability of Type I error) at a constant value, usually 0.05. If the goal of null hypothesis testing is to present conclusions in which we have the highest possible confidence, then the only logical decision-making threshold is the value that minimizes the probability (or occasionally, cost) of making errors. Setting α to minimize the combination of Type I and Type II error at a critical effect size can easily be accomplished for traditional statistical tests by calculating the α associated with the minimum average of α and ß at the critical effect size. This technique also has the flexibility to incorporate prior probabilities of null and alternate hypotheses and/or relative costs of Type I and Type II errors, if known. Using an optimal α results in stronger scientific inferences because it estimates and minimizes both Type I errors and relevant Type II errors for a test. It also results in greater transparency concerning assumptions about relevant effect size(s) and the relative costs of Type I and II errors. By contrast, the use of α = 0.05 results in arbitrary decisions about what effect sizes will likely be considered significant, if real, and results in arbitrary amounts of Type II error for meaningful potential effect sizes. We cannot identify a rationale for continuing to arbitrarily use α = 0.05 for null hypothesis significance tests in any field, when it is possible to determine an optimal α.

Tadpole mortality varies across experimental venues: do laboratory populations predict responses in nature?

Laboratory experiments are widely used to study how populations in nature might respond to various biological interactions, but the relevance of experiments in artificial venues is not known. We compiled mortality and growth data from 424 anuran populations carried out under laboratory, mesocosm, field enclosure, and field settings to determine if major differences exist amongst experimental venues and how this might influence experimental responses of tadpoles amongst venues. Our results show that there are fundamental differences in survival amongst venues, with the highest mortality occurring in field populations and the lowest in laboratory populations. Separation of mesocosm and field enclosure data based on the possibility of predatory interactions indicates that predation is an important factor leading to increased mortality in natural populations. Comparisons of size distributions across venues (although size data were limited for field populations) suggest that variation in tadpole size is low in natural populations compared to populations in artificial venues. We infer from this that mortality has a homogenizing effect on size in nature, resulting in natural populations that are not a random sample of hatched individuals. This finding suggests that populations reared under controlled laboratory conditions in the absence of predation (and other selective pressures) may not be representative of natural populations.

Varying responses of northeastern North American amphibians to the chytrid pathogen Batrachochytrium dendrobatidis.

Chytridiomycosis, caused by Batrachochytrium dendrobatidis (Bd), is widespread among amphibians in northeastern North America. It is unknown, however, whether Bd has the potential to cause extensive amphibian mortalities in northeastern North America as have occurred elsewhere. In the laboratory, we exposed seven common northeastern North American amphibian species to Bd to assess the likelihood of population-level effects from the disease. We exposed larval wood frogs (Lithobates sylvaticus) and postmetamorphic frogs of six other species to two different strains of Bd, a northeastern strain (JEL404) and a strain that caused die-offs of amphibians in Panama (JEL423), under ideal in vitro growth conditions for Bd. Exposed American toads (Anaxyrus americanus) all died; thus, this species may be the most likely to die from Bd-caused disease in the wild. Both Bd strains were associated with mortalities of wood frogs, although half the metamorphs survived. The Bd strain from Panama killed metamorphic green frogs (L. clamitans), whereas the northeastern strain did not, which means novel strains of Bd may lead to death even when local strains may not. No mortality was observed in four species (bullfrogs [L. catesbeianus], northern leopard frogs [L. pipiens], spring peepers [Pseudacris crucifer], and blue-spotted salamanders [Ambystoma laterale]) and in some individuals of green frogs and wood frogs that we exposed. This finding suggests these six species may be Bd vectors. Our results show that systematic exposures of amphibian species to Bd in the laboratory may be a good first step in the identification of species susceptible to Bd-caused declines and in directing regional conservation efforts aimed at susceptible species.

Effects of chytrid fungus and a glyphosate-based herbicide on survival and growth of wood frogs (Lithobates sylvaticus).

Anthropogenic-derived stressors in the environment, such as contaminants, are increasingly considered important cofactors that may decrease the immune response of amphibians to pathogens. Few studies, however, have integrated amphibian disease and contaminants to test this multiple-stressor hypothesis for amphibian declines. We examined whether exposure to sublethal concentrations of a glyphosate-based herbicide and two strains of the pathogenic chytrid fungus, Batrachochrytrium dendrobatidis (Bd) could: (1) sublethally affect wood frogs (Lithobates sylvaticus) by altering the time to and size at metamorphosis, and (2) directly affect survivability of wood frogs after metamorphosis. Neither Bd strain nor herbicide exposure alone significantly altered growth or time to metamorphosis. The two Bd strains did not differ in their pathogenicity, and both caused mortality in post-metamorphic wood frogs. There was no evidence of an interaction between treatments, indicating a lack of herbicide-induced susceptibility to Bd. However, the trends in our data suggest that exposure of wood frogs to a high concentration of glyphosate-based herbicide may reduce Bd-caused mortality compared to animals exposed to Bd alone. These results exemplify the complexities inherent when populations are coping with multiple stressors. In this case, the perceived stressor, glyphosate-based herbicide, appeared to affect the pathogen more than the host's immune system, relieving the host from disease-caused effects. This suggests caution when invoking multiple stressors as a cause for increased disease susceptibility and indicates that the effects of multiple stressors on disease outcome depend on the interrelationships of stressors to both the pathogen and the host.