Aquatic Soundscape Recordings Reveal Diverse Vocalizations and Nocturnal Activity of an Endangered Frog.

AbstractAutonomous sensors provide opportunities to observe organisms across spatial and temporal scales that humans cannot directly observe. By processing large data streams from autonomous sensors with deep learning methods, researchers can make novel and important natural history discoveries. In this study, we combine automated acoustic monitoring with deep learning models to observe breeding-associated activity in the endangered Sierra Nevada yellow-legged frog (Rana sierrae), a behavior that current surveys do not measure. By deploying inexpensive hydrophones and developing a deep learning model to recognize breeding-associated vocalizations, we discover three undocumented R. sierrae vocalization types and find an unexpected temporal pattern of nocturnal breeding-associated vocal activity. This study exemplifies how the combination of autonomous sensor data and deep learning can shed new light on species' natural history, especially during times or in locations where human observation is limited or impossible.

Host density has limited effects on pathogen invasion, disease-induced declines and within-host infection dynamics across a landscape of disease.

1. Host density is hypothesized to be a major driver of variability in the responses and outcomes of wildlife populations following pathogen invasion. While the effects of host density on pathogen transmission have been extensively studied, these studies are dominated by theoretical analyses and small-scale experiments. This focus leads to an incomplete picture regarding how host density drives observed variability in disease outcomes in the field. 2. Here, we leveraged a dataset of hundreds of replicate amphibian populations that varied by orders of magnitude in host density. We used these data to test the effects of host density on three outcomes following the arrival of the amphibian-killing fungal pathogen Batrachochytrium dendrobatidis (Bd): the probability that Bd successfully invaded a host population and led to a pathogen outbreak, the magnitude of the host population-level decline following an outbreak and within-host infection dynamics that drive population-level outcomes in amphibian-pathogen systems. 3. Based on previous small-scale transmission experiments, we expected that populations with higher densities would be more likely to experience Bd outbreaks and would suffer larger proportional declines following outbreaks. To test these predictions, we developed and fitted a Hidden Markov Model that accounted for imperfectly observed disease outbreak states in the amphibian populations we surveyed. 4. Contrary to our predictions, we found minimal effects of host density on the probability of successful Bd invasion, the magnitude of population decline following Bd invasion and the dynamics of within-host infection intensity. Environmental conditions, such as summer temperature, winter severity and the presence of pathogen reservoirs, were more predictive of variability in disease outcomes. 5. Our results highlight the limitations of extrapolating findings from small-scale transmission experiments to observed disease trajectories in the field and provide strong evidence that variability in host density does not necessarily drive variability in host population responses following pathogen arrival. In an applied context, we show that feedbacks between host density and disease will not necessarily affect the success of reintroduction efforts in amphibian-Bd systems of conservation concern.

Cryptic diversity of a widespread global pathogen reveals expanded threats to amphibian conservation.

Biodiversity loss is one major outcome of human-mediated ecosystem disturbance. One way that humans have triggered wildlife declines is by transporting disease-causing agents to remote areas of the world. Amphibians have been hit particularly hard by disease due in part to a globally distributed pathogenic chytrid fungus (Batrachochytrium dendrobatidis [Bd]). Prior research has revealed important insights into the biology and distribution of Bd; however, there are still many outstanding questions in this system. Although we know that there are multiple divergent lineages of Bd that differ in pathogenicity, we know little about how these lineages are distributed around the world and where lineages may be coming into contact. Here, we implement a custom genotyping method for a global set of Bd samples. This method is optimized to amplify and sequence degraded DNA from noninvasive skin swab samples. We describe a divergent lineage of Bd, which we call BdASIA3, that appears to be widespread in Southeast Asia. This lineage co-occurs with the global panzootic lineage (BdGPL) in multiple localities. Additionally, we shed light on the global distribution of BdGPL and highlight the expanded range of another lineage, BdCAPE. Finally, we argue that more monitoring needs to take place where Bd lineages are coming into contact and where we know little about Bd lineage diversity. Monitoring need not use expensive or difficult field techniques but can use archived swab samples to further explore the history-and predict the future impacts-of this devastating pathogen.

Divergent regional evolutionary histories of a devastating global amphibian pathogen.

Emerging infectious diseases are a pressing threat to global biological diversity. Increased incidence and severity of novel pathogens underscores the need for methodological advances to understand pathogen emergence and spread. Here, we use genetic epidemiology to test, and challenge, key hypotheses about a devastating zoonotic disease impacting amphibians globally. Using an amplicon-based sequencing method and non-invasive samples we retrospectively explore the history of the fungal pathogen Batrachochytrium dendrobatidis (Bd) in two emblematic amphibian systems: the Sierra Nevada of California and Central Panama. The hypothesis in both regions is the hypervirulent Global Panzootic Lineage of Bd (BdGPL) was recently introduced and spread rapidly in a wave-like pattern. Our data challenge this hypothesis by demonstrating similar epizootic signatures can have radically different underlying evolutionary histories. In Central Panama, our genetic data confirm a recent and rapid pathogen spread. However, BdGPL in the Sierra Nevada has remarkable spatial structuring, high genetic diversity and a relatively older history inferred from time-dated phylogenies. Thus, this deadly pathogen lineage may have a longer history in some regions than assumed, providing insights into its origin and spread. Overall, our results highlight the importance of integrating observed wildlife die-offs with genetic data to more accurately reconstruct pathogen outbreaks.

Stepping into the past to conserve the future: Archived skin swabs from extant and extirpated populations inform genetic management of an endangered amphibian.

Moving animals on a landscape through translocations and reintroductions is an important management tool used in the recovery of endangered species, particularly for the maintenance of population genetic diversity and structure. Management of imperiled amphibian species rely heavily on translocations and reintroductions, especially for species that have been brought to the brink of extinction by habitat loss, introduced species, and disease. One striking example of amphibian declines and associated management efforts is in California's Sequoia and Kings Canyon National Parks with the mountain yellow-legged frog species complex (Rana sierrae/muscosa). Mountain yellow-legged frogs have been extirpated from more than 93% of their historic range, and limited knowledge of their population genetics has made long-term conservation planning difficult. To address this, we used 598 archived skin swabs from both extant and extirpated populations across 48 lake basins to generate a robust Illumina-based nuclear amplicon data set. We found that samples grouped into three main genetic clusters, concordant with watershed boundaries. We also found evidence for historical gene flow across watershed boundaries with a north-to-south axis of migration. Finally, our results indicate that genetic diversity is not significantly different between populations with different disease histories. Our study offers specific management recommendations for imperiled mountain yellow-legged frogs and, more broadly, provides a population genetic framework for leveraging minimally invasive samples for the conservation of threatened species.

Reduced skin bacterial diversity correlates with increased pathogen infection intensity in an endangered amphibian host.

The fungal pathogen Batrachochytrium dendrobatidis (Bd) infects the skin of amphibians and has caused severe declines and extinctions of amphibians globally. In this study, we investigate the interaction between Bd and the bacterial skin microbiome of the endangered Sierra Nevada yellow-legged frog, Rana sierrae, using both culture-dependent and culture-independent methods. Samples were collected from two populations of R. sierrae that likely underwent Bd epizootics in the past, but that continue to persist with Bd in an enzootic disease state, and we address the hypothesis that such "persistent" populations are aided by mutualistic skin microbes. Our 16S rRNA metabarcoding data reveal that the skin microbiome of highly infected juvenile frogs is characterized by significantly reduced species richness and evenness, and by strikingly lower variation between individuals, compared to juveniles and adults with lower infection levels. Over 90% of DNA sequences from the skin microbiome of highly infected frogs were derived from bacteria in a single order, Burkholderiales, compared to just 54% in frogs with lower infection levels. In a culture-dependent Bd inhibition assay, the bacterial metabolites we evaluated all inhibited the growth of Bd. Together, these results illustrate the disruptive effects of Bd infection on host skin microbial community structure and dynamics, and suggest possible avenues for the development of anti-Bd probiotic treatments.

Large-scale recovery of an endangered amphibian despite ongoing exposure to multiple stressors.

Amphibians are one of the most threatened animal groups, with 32% of species at risk for extinction. Given this imperiled status, is the disappearance of a large fraction of the Earth's amphibians inevitable, or are some declining species more resilient than is generally assumed? We address this question in a species that is emblematic of many declining amphibians, the endangered Sierra Nevada yellow-legged frog (Rana sierrae). Based on >7,000 frog surveys conducted across Yosemite National Park over a 20-y period, we show that, after decades of decline and despite ongoing exposure to multiple stressors, including introduced fish, the recently emerged disease chytridiomycosis, and pesticides, R. sierrae abundance increased sevenfold during the study and at a rate of 11% per year. These increases occurred in hundreds of populations throughout Yosemite, providing a rare example of amphibian recovery at an ecologically relevant spatial scale. Results from a laboratory experiment indicate that these increases may be in part because of reduced frog susceptibility to chytridiomycosis. The disappearance of nonnative fish from numerous water bodies after cessation of stocking also contributed to the recovery. The large-scale increases in R. sierrae abundance that we document suggest that, when habitats are relatively intact and stressors are reduced in their importance by active management or species' adaptive responses, declines of some amphibians may be partially reversible, at least at a regional scale. Other studies conducted over similarly large temporal and spatial scales are critically needed to provide insight and generality about the reversibility of amphibian declines at a global scale.

Resistance, tolerance and environmental transmission dynamics determine host extinction risk in a load-dependent amphibian disease.

While disease-induced extinction is generally considered rare, a number of recently emerging infectious diseases with load-dependent pathology have led to extinction in wildlife populations. Transmission is a critical factor affecting disease-induced extinction, but the relative importance of transmission compared to load-dependent host resistance and tolerance is currently unknown. Using a combination of models and experiments on an amphibian species suffering extirpations from the fungal pathogen Batrachochytrium dendrobatidis (Bd), we show that while transmission from an environmental Bd reservoir increased the ability of Bd to invade an amphibian population and the extinction risk of that population, Bd-induced extinction dynamics were far more sensitive to host resistance and tolerance than to Bd transmission. We demonstrate that this is a general result for load-dependent pathogens, where non-linear resistance and tolerance functions can interact such that small changes in these functions lead to drastic changes in extinction dynamics.

Epidemic and endemic pathogen dynamics correspond to distinct host population microbiomes at a landscape scale.

Infectious diseases have serious impacts on human and wildlife populations, but the effects of a disease can vary, even among individuals or populations of the same host species. Identifying the reasons for this variation is key to understanding disease dynamics and mitigating infectious disease impacts, but disentangling cause and correlation during natural outbreaks is extremely challenging. This study aims to understand associations between symbiotic bacterial communities and an infectious disease, and examines multiple host populations before or after pathogen invasion to infer likely causal links. The results show that symbiotic bacteria are linked to fundamentally different outcomes of pathogen infection: host-pathogen coexistence (endemic infection) or host population extirpation (epidemic infection). Diversity and composition of skin-associated bacteria differed between populations of the frog, Rana sierrae, that coexist with or were extirpated by the fungal pathogen, Batrachochytrium dendrobatidis (Bd). Data from multiple populations sampled before or after pathogen invasion were used to infer cause and effect in the relationship between the fungal pathogen and symbiotic bacteria. Among host populations, variation in the composition of the skin microbiome was most strongly predicted by pathogen infection severity, even in analyses where the outcome of infection did not vary. This result suggests that pathogen infection shapes variation in the skin microbiome across host populations that coexist with or are driven to extirpation by the pathogen. By contrast, microbiome richness was largely unaffected by pathogen infection intensity, but was strongly predicted by geographical region of the host population, indicating the importance of environmental or host genetic factors in shaping microbiome richness. Thus, while both richness and composition of the microbiome differed between endemic and epidemic host populations, the underlying causes are most likely different: pathogen infection appears to shape microbiome composition, while microbiome richness was less sensitive to pathogen-induced disturbance. Because higher richness was correlated with host persistence in the presence of Bd, and richness appeared relatively stable to Bd infection, microbiome richness may contribute to disease resistance, although the latter remains to be directly tested.

Enzootic and epizootic dynamics of the chytrid fungal pathogen of amphibians.

Chytridiomycosis, the disease caused by the chytrid fungus, Batrachochytrium dendrobatidis (Bd), has contributed to amphibian population declines and extinctions worldwide. The impact of this pathogen, however, varies markedly among amphibian species and populations. Following invasion into some areas of California's Sierra Nevada, Bd leads to rapid declines and local extinctions of frog populations (Rana muscosa, R. sierrae). In other areas, infected populations of the same frog species have declined but persisted at low host densities for many years. We present results of a 5-year study showing that infected adult frogs in persistent populations have low fungal loads, are surviving between years, and frequently lose and regain the infection. Here we put forward the hypothesis that fungal load dynamics can explain the different population-level outcomes of Bd observed in different areas of the Sierra Nevada and possibly throughout the world. We develop a model that incorporates the biological details of the Bd-host interaction. Importantly, model results suggest that host persistence versus extinction does not require differences in host susceptibility, pathogen virulence, or environmental conditions, and may be just epidemic and endemic population dynamics of the same host-pathogen system. The different disease outcomes seen in natural populations may result solely from density-dependent host-pathogen dynamics. The model also shows that persistence of Bd is enhanced by the long-lived tadpole stage that characterize these two frog species, and by nonhost Bd reservoirs.

Dynamics of an emerging disease drive large-scale amphibian population extinctions.

Epidemiological theory generally suggests that pathogens will not cause host extinctions because the pathogen should fade out when the host population is driven below some threshold density. An emerging infectious disease, chytridiomycosis, caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd) is directly linked to the recent extinction or serious decline of hundreds of amphibian species. Despite continued spread of this pathogen into uninfected areas, the dynamics of the host-pathogen interaction remain unknown. We use fine-scale spatiotemporal data to describe (i) the invasion and spread of Bd through three lake basins, each containing multiple populations of the mountain yellow-legged frog, and (ii) the accompanying host-pathogen dynamics. Despite intensive sampling, Bd was not detected on frogs in study basins until just before epidemics began. Following Bd arrival in a basin, the disease spread to neighboring populations at approximately 700 m/yr in a wave-like pattern until all populations were infected. Within a population, infection prevalence rapidly reached 100% and infection intensity on individual frogs increased in parallel. Frog mass mortality began only when infection intensity reached a critical threshold and repeatedly led to extinction of populations. Our results indicate that the high growth rate and virulence of Bd allow the near-simultaneous infection and buildup of high infection intensities in all host individuals; subsequent host population crashes therefore occur before Bd is limited by density-dependent factors. Preventing infection intensities in host populations from reaching this threshold could provide an effective strategy to avoid the extinction of susceptible amphibian species in the wild.

Effectiveness of antifungal treatments during chytridiomycosis epizootics in populations of an endangered frog.

The recently-emerged amphibian chytrid fungus Batrachochytrium dendrobatidis (Bd) has had an unprecedented impact on global amphibian populations, and highlights the urgent need to develop effective mitigation strategies. We conducted in-situ antifungal treatment experiments in wild populations of the endangered mountain yellow-legged frog during or immediately after Bd-caused mass die-off events. The objective of treatments was to reduce Bd infection intensity ("load") and in doing so alter frog-Bd dynamics and increase the probability of frog population persistence despite ongoing Bd infection. Experiments included treatment of early life stages (tadpoles and subadults) with the antifungal drug itraconazole, treatment of adults with itraconazole, and augmentation of the skin microbiome of subadults with Janthinobacterium lividum, a commensal bacterium with antifungal properties. All itraconazole treatments caused immediate reductions in Bd load, and produced longer-term effects that differed between life stages. In experiments focused on early life stages, Bd load was reduced in the 2 months immediately following treatment and was associated with increased survival of subadults. However, Bd load and frog survival returned to pre-treatment levels in less than 1 year, and treatment had no effect on population persistence. In adults, treatment reduced Bd load and increased frog survival over the entire 3-year post-treatment period, consistent with frogs having developed an effective adaptive immune response against Bd. Despite this protracted period of reduced impacts of Bd on adults, recruitment into the adult population was limited and the population eventually declined to near-extirpation. In the microbiome augmentation experiment, exposure of subadults to a solution of J. lividum increased concentrations of this potentially protective bacterium on frogs. However, concentrations declined to baseline levels within 1 month and did not have a protective effect against Bd infection. Collectively, these results indicate that our mitigation efforts were ineffective in causing long-term changes in frog-Bd dynamics and increasing population persistence, due largely to the inability of early life stages to mount an effective immune response against Bd. This results in repeated recruitment failure and a low probability of population persistence in the face of ongoing Bd infection.

The amphibian microbiome exhibits poor resilience following pathogen-induced disturbance.

Infectious pathogens can disrupt the microbiome in addition to directly affecting the host. Impacts of disease may be dependent on the ability of the microbiome to recover from such disturbance, yet remarkably little is known about microbiome recovery after disease, particularly in nonhuman animals. We assessed the resilience of the amphibian skin microbial community after disturbance by the pathogen, Batrachochytrium dendrobatidis (Bd). Skin microbial communities of laboratory-reared mountain yellow-legged frogs were tracked through three experimental phases: prior to Bd infection, after Bd infection (disturbance), and after clearing Bd infection (recovery period). Bd infection disturbed microbiome composition and altered the relative abundances of several dominant bacterial taxa. After Bd infection, frogs were treated with an antifungal drug that cleared Bd infection, but this did not lead to recovery of microbiome composition (measured as Unifrac distance) or relative abundances of dominant bacterial groups. These results indicate that Bd infection can lead to an alternate stable state in the microbiome of sensitive amphibians, or that microbiome recovery is extremely slow-in either case resilience is low. Furthermore, antifungal treatment and clearance of Bd infection had the additional effect of reducing microbial community variability, which we hypothesize results from similarity across frogs in the taxa that colonize community vacancies resulting from the removal of Bd. Our results indicate that the skin microbiota of mountain yellow-legged frogs has low resilience following Bd-induced disturbance and is further altered by the process of clearing Bd infection, which may have implications for the conservation of this endangered amphibian.

Nonnative trout impact an alpine-nesting bird by altering aquatic-insect subsidies.

Adjacent food webs may be linked by cross-boundary subsidies: more-productive donor systems can subsidize consumers in less-productive neighboring recipient systems. Introduced species are known to have direct effects on organisms within invaded communities. However, few studies have addressed the indirect effects of nonnative species in donor systems on organisms in recipient systems. We studied the direct role of introduced trout in altering a lake-derived resource subsidy and their indirect effects in altering a passerine bird's response to that subsidy. We compared the abundance of aquatic insects and foraging Gray-crowned Rosy-Finches (Leucosticte tephrocotis dawsoni, "Rosy-Finch") at fish-containing vs. fishless lakes in the Sierra Nevada Mountains of California (USA). Introduced trout outcompeted Rosy-Finches for emerging aquatic insects (i.e., mayflies). Fish-containing lakes had 98% fewer mayflies than did fishless lakes. In lakes without fish, Rosy-Finches showed an aggregative response to emerging aquatic insects with 5.9 times more Rosy-Finches at fishless lakes than at fish-containing lakes. Therefore, the introduction of nonnative fish into the donor system reduced both the magnitude of the resource subsidy and the strength of cross-boundary trophic interactions. Importantly, the timing of the subsidy occurs when Rosy-Finches feed their young. If Rosy-Finches rely on aquatic-insect subsidies to fledge their young, reductions in the subsidy by introduced trout may have decreased Rosy-Finch abundances from historic levels. We recommend that terrestrial recipients of aquatic subsidies be included in conservation and restoration plans for ecosystems with alpine lakes.

Allee effect limits colonization success of sexually reproducing zooplankton.

Understanding the dynamics of populations at low density and the role of Allee effects is a priority due to concern about the decline of rare species and interest in colonization/invasion dynamics. Despite well-developed theory and observational support, experimental examinations of the Allee effect in natural systems are rare, partly because of logistical difficulties associated with experiments at low population density. We took advantage of fish introduction and removal in alpine lakes to experimentally test for the Allee effect at the whole-ecosystem scale. The large copepod Hesperodiaptomus shoshone is often extirpated from the water column by fish and sometimes fails to recover following fish disappearance, despite the presence of a long-lived egg bank. Population growth rate of this dioecious species may be limited by mate encounter rate, such that below some critical density a colonizing population will fail to establish. We conducted a multi-lake experiment in which H. shoshone was stocked at densities that bracketed our hypothesized critical density of 0.5-5 copoepods/m3. Successful recovery by the copepod was observed only in the lake with the highest initial density (3 copepods/m3). Copepods stocked into small cages at 3000 copepods/m3 survived and reproduced at rates comparable to natural populations, confirming that the lakes were suitable habitat for this species. In support of mate limitation as the mechanism underlying recovery failure, we found a significant positive relationship between mating success and density across experimental and natural H. shoshone populations. Furthermore, a mesocosm experiment provided evidence of increased per capita population growth rate with increasing population density in another diaptomid species, Skistodiaptomus pallidus. Together, these lines of evidence support the importance of the Allee effect to population recovery of H. shoshone in the Sierra Nevada, and to diaptomid copepods in general.

Recovery after local extinction: factors affecting re-establishment of alpine lake zooplankton.

The introduction of fishes into naturally fishless mountain lakes often results in the extirpation of large-bodied zooplankton species. The ability to predict whether or not particular species will recover following fish removal is critically important for the design and implementation of lake restoration efforts but is currently not possible because of a lack of information on what factors affect recovery. The objective of this study was to identify the factors influencing recovery probability in two large-bodied zooplankton species following fish removal. We predicted that (1) Daphnia melanica would have a higher probability of recovery than Hesperodiaptomus shoshone due to differences in reproductive mode (D. melanica is parthenogenetic, H. shoshone is obligately sexual), (2) recovery probability would be a decreasing function of fish residence time due to the negative relationship between fish residence time and size of the egg bank, and (3) recovery probability would be an increasing function of lake depth as a consequence of a positive relationship between lake depth and egg bank size. To test these predictions, we sampled contemporary zooplankton populations and collected paleolimnological data from 44 naturally fishless lakes that were stocked with trout for varying lengths of time before reverting to a fishless condition. D. melanica had a significantly higher probability of recovery than did H. shoshone (0.82 vs. 0.54, respectively). The probability of recovery for H. shoshone was also significantly influenced by lake depth, fish residence time, and elevation, but only elevation influenced the probability of recovery in D. melanica. These results are consistent with between-species differences in reproductive mode combined with the much greater longevity of diapausing eggs in D. melanica than in H. shoshone. Our data also suggest that H. shoshone will often fail to recover in lakes with fish residence times exceeding 50 years.

Rapid evolution in response to introduced predators II: the contribution of adaptive plasticity.

BACKGROUND: Introductions of non-native species can significantly alter the selective environment for populations of native species, which can respond through phenotypic plasticity or genetic adaptation. We examined phenotypic and genetic responses of Daphnia populations to recent introductions of non-native fish to assess the relative roles of phenotypic plasticity versus genetic change in causing the observed patterns. The Daphnia community in alpine lakes throughout the Sierra Nevada of California (USA) is ideally suited for investigation of rapid adaptive evolution because there are multiple lakes with and without introduced fish predators. We conducted common-garden experiments involving presence or absence of chemical cues produced by fish and measured morphological and life-history traits in Daphnia melanica populations collected from lakes with contrasting fish stocking histories. The experiment allowed us to assess the degree of population differentiation due to fish predation and examine the contribution of adaptive plasticity in the response to predator introduction. RESULTS: Our results show reductions in egg number and body size of D. melanica in response to introduced fish. These phenotypic changes have a genetic basis but are partly due to a direct response to chemical cues from fish via adaptive phenotypic plasticity. Body size showed the largest phenotypic change, on the order of nine phenotypic standard deviations, with approximately 11% of the change explained by adaptive plasticity. Both evolutionary and plastic changes in body size and egg number occurred but no changes in the timing of reproduction were observed. CONCLUSION: Native Daphnia populations exposed to chemical cues produced by salmonid fish predators display adaptive plasticity for body size and fecundity. The magnitude of adaptive plasticity was insufficient to explain the total phenotypic change, so the realized change in phenotypic means in populations exposed to introduced fish may be the result of a combination of initial plasticity and subsequent genetic adaptation. Our results suggest that immediately following the introduction of fish predators, adaptive plasticity may reduce the impact of selection through "Baldwin/Bogert effects" by facilitating the movement of populations toward new fitness optima. Our study of the response of a native species to an introduced predator enhances our understanding of the conditions necessary for rapid adaptive evolution and the relationship between rapid evolution and adaptive phenotypic plasticity.

Rapid evolution in response to introduced predators I: rates and patterns of morphological and life-history trait divergence.

BACKGROUND: Introduced species can have profound effects on native species, communities, and ecosystems, and have caused extinctions or declines in native species globally. We examined the evolutionary response of native zooplankton populations to the introduction of non-native salmonids in alpine lakes in the Sierra Nevada of California, USA. We compared morphological and life-history traits in populations of Daphnia with a known history of introduced salmonids and populations that have no history of salmonid introductions. RESULTS: Our results show that Daphnia populations co-existing with fish have undergone rapid adaptive reductions in body size and in the timing of reproduction. Size-related traits decreased by up to 13 percent in response to introduced fish. Rates of evolutionary change are as high as 4,238 darwins (0.036 haldanes). CONCLUSION: Species introductions into aquatic habitats can dramatically alter the selective environment of native species leading to a rapid evolutionary response. Knowledge of the rates and limits of adaptation is an important component of understanding the long-term effects of alterations in the species composition of communities. We discuss the evolutionary consequences of species introductions and compare the rate of evolution observed in the Sierra Nevada Daphnia to published estimates of evolutionary change in ecological timescales.

Multiple stressors and amphibian declines: dual impacts of pesticides and fish on yellow-legged frogs.

More than 40% of Earth's 5700+ amphibian species have undergone recent declines. Despite the likely involvement of multiple factors in driving these declines, most studies continue to focus on single stressors. In California (USA), separate studies have implicated either introduced fish or pesticides as causal agents. To date, however, no study has simultaneously evaluated the respective roles of these two potential stressors nor attempted to assess their relative importance, information critical for the development of effective conservation efforts and environmental policies. We examined the role and relative effect of fish and pesticides on the mountain yellow-legged frog (Rana muscosa) using unusually detailed data sets for a large portion of R. muscosa's historic range in California's Sierra Nevada. Habitat characteristics and presence/absence of R. muscosa and fish were quantified at each of 6831 sites during field surveys. Pesticide use upwind of each site was calculated from pesticide application records and predominant wind directions. Using generalized additive models, we found that, after accounting for habitat effects, the probability of R. muscosa presence was significantly reduced by both fish and pesticides, with the landscape-scale effect of pesticides much stronger than that of fish. The degree to which a site was sheltered from the predominant wind (and associated pesticides) was also a significant predictor of R. muscosa presence. Taken together, these results represent the strongest evidence to date that windborne pesticides are contributing to amphibian declines in pristine locations. Our results suggest that amphibian declines may have complex multi-factorial causes, and caution that single-factor studies that demonstrate the importance of one factor should not be used as evidence against the importance of other factors.

Removal of nonnative fish results in population expansion of a declining amphibian (mountain yellow-legged frog, Rana muscosa).

The mountain yellow-legged frog (Rana muscosa) was once a common inhabitant of the Sierra Nevada (California, USA), but has declined precipitously during the past century due in part to the introduction of nonnative fish into naturally fishless habitats. The objectives of the current study were to describe (1) the effect of fish removal from three lakes (located in two watersheds) on the small, remnant R. muscosa populations inhabiting those lakes, and (2) the initial development of metapopulation structure in each watershed as R. muscosa from expanding populations in fish-removal lakes dispersed to adjacent habitats. At all three fish-removal lakes, R. muscosa population densities increased significantly following the removal of predatory fish. The magnitude of these increases was significantly greater than that observed over the same time period in R. muscosa populations inhabiting control lakes that remained in their natural fishless condition. Following these population increases, R. muscosa dispersed to adjacent suitable (but unoccupied) sites, moving between 200 and 900 m along streams or across dry land. Together, these results suggest that large-scale removal of introduced fish could result in at least partial reversal of the decline of R. muscosa. Continued monitoring of R. muscosa at the fish-removal sites will be necessary to determine whether the positive effects of fish eradication are sustained over the long-term, especially in light of the increasingly important role played by an emerging infectious disease (chytridiomycosis, caused by Batrachochytrium dendrobatidis) in influencing R. muscosa populations.