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.

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.

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.

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.

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.

Exposure of juvenile green frogs (Lithobates clamitans) in littoral enclosures to a glyphosate-based herbicide.

The majority of studies on the toxicity of glyphosate-based herbicides to amphibians have focused on larval life stages exposed in aqueous media. However, adult and juvenile amphibians may also be exposed directly or indirectly to herbicides. The potential for such exposures is of particular interest in the littoral zone surrounding wetlands as this is preferred habitat for many amphibian species. Moreover, it may be argued that potential herbicide effects on juvenile or adult amphibians could have comparatively greater influence on overall recruitment, reproductive potential and thus stability of local populations than effects on larvae. In this experiment, juvenile green frogs (Lithobates clamitans) were exposed to two concentrations (2.16 and 4.27 kg a.e./ha) of a glyphosate-based herbicide formulation (VisionMax®), which were based on typical application scenarios in Canadian forestry. The experimental design employed frogs inhabiting in situ enclosures established at the edge of small naturalized wetlands that were split in half using an impermeable plastic barrier. When analyzed using nominal target application rates, exposure to the glyphosate-based herbicide had no significant effect on survival, body condition, liver somatic index or the observed rate of Batrachochytrium dendrobatidis infection. However, there were marginal trends in both ANOVA analysis and post-hoc regressions regarding B. dendrobatidis infection rates and liver somatic index in relation to measured exposure estimates. Results from this study highlight the importance of field research and the need to include multiple endpoints when examining potential effects of a contaminant on non-target organisms.

Life-history strategies associated with local population variability confer regional stability.

A widely held ecological tenet is that, at the local scale, populations of K-selected species (i.e. low fecundity, long lifespan and large body size) will be less variable than populations of r-selected species (i.e. high fecundity, short lifespan and small body size). We examined the relationship between long-term population trends and life-history attributes for 185 bird species in the Czech Republic and found that, at regional spatial scales and over moderate temporal scales (100-120 years), K-selected bird species were more likely to show both large increases and decreases in population size than r-selected species. We conclude that life-history attributes commonly associated with variable populations at the local scale, confer stability at the regional scale.