Sex‐biased disease dynamics increase extinction risk by impairing population recovery

The periodicity of life‐cycle events (phenology) modulates host availability to pathogens in a repeatable pattern. The effects of sexual differences in host phenology have been little explored in wildlife epidemiological studies. A recent series of ranavirosis outbreaks led to serious declines of Boscas’ newt populations at Serra da Estrela (Portugal). The peculiar phenology of this species, in which a large number of females remain in the aquatic habitat after the breeding season, turns it into a suitable model to test how sex‐biased mortality can affect host population persistence in the context of infectious diseases. We investigated how the phenology of Bosca's newt (i.e. biased number of females) mediated the impact of Ranavirus. We then evaluated the risk of extinction of the population under different scenarios of sex‐biased mortality using a population viability analysis. Two newt populations (one subject to yearly outbreaks and a comparative site where outbreaks have not been recorded) were tracked for trends over time following emergence of ranaviral disease, allowing us to assess the differential impact of the disease on both sexes. In addition to a significant decline in abundance of adult newts, our data suggest that phenology can affect disease dynamics indirectly, leading to reduction in females and a reversal of the sex ratio of the breeding population. Our models suggest that female‐biased mortality does not exacerbate Ranavirus‐driven population declines in the short‐term, but is likely to have a deleterious impact during the recovery process once the lethal effect of disease is removed from the system

Chromosomal copy number variation, selection and uneven rates of recombination reveal cryptic genome diversity linked to pathogenicity

This project was funded by the UK Natural Environmental Research Council (NERC) grant NE/E006701/1, the European Research Council (ERC) grant 260801-BIG_IDEA, the Swiss National Science Foundation grant 31-125099 and the Biodiversa project RACE: Risk Assessment of Chytridiomycosis to European Amphibian Biodiversity (http://www.bd-maps.eu). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer reviewedPublisher PD

Expression Profiling the Temperature-Dependent Amphibian Response to Infection by Batrachochytrium dendrobatidis

Amphibians are experiencing a panzootic of unprecedented proportions caused by the emergence of Batrachochytrium dendrobatidis (Bd). However, all species are not equally at risk of infection, and risk is further modified by environmental variables, specifically temperature. In order to understand how, and when, hosts mount a response to Bd we analysed infection dynamics and patterns of gene expression in the model amphibian species Silurana (Xenopus) tropicalis. Mathematical modelling of infection dynamics demonstrate the existence of a temperature-dependent protective response that is largely independent of the intrinsic growth-rate of Bd. Using temporal expression-profiling by microarrays and qRT-PCR, we characterise this response in the main amphibian lymphoid tissue, the spleen. We demonstrate that clearance of Bd at the host-optimal temperature is not clearly associated with an adaptive immune response, but rather is correlated with the induction of components of host innate immunity including the expression of genes that are associated with the production of the antimicrobial skin peptide preprocareulein (PPCP) as well as inflammatory responses. We find that adaptive immunity appears to be lacking at host-optimal temperatures. This suggests that either Bd does not stimulate, or suppresses, adaptive immunity, or that trade-offs exist between innate and adaptive limbs of the amphibian immune system. At cold temperatures, S. tropicalis loses the ability to mount a PPCP-based innate response, and instead manifests a more pronounced inflammatory reaction that is characterised by the production of proteases and higher pathogen burdens. This study demonstrates the temperature-dependency of the amphibian response to infection by Bd and indicates the influence that changing climates may exert on the ectothermic host response to pathogens

Pathological and Phylogenetic characterisation of Amphibiothecum sp. infection in an isolated amphibian (Lissotriton helveticus) population on the island of Rum (Scotland)

Outbreaks of cutaneous infectious disease in amphibians are increasingly being attributed to an overlooked group of fungal-like pathogens, the Dermocystids. During the last 10 years on the Isle of Rum, Scotland, palmate newts (Lissotriton helveticus) have been reportedly afflicted by unusual skin lesions. Here we present pathological and molecular findings confirming that the pathogen associated with these lesions is a novel organism of the order Dermocystida, and represents the first formally reported, and potentially lethal, case of amphibian Dermocystid infection in the UK. Whilst the gross pathology and the parasite cyst morphology were synonymous to those described in a study from infectedL. helveticusin France, we observed a more extreme clinical outcome on Rum involving severe subcutaneous oedema. Phylogenetic topologies supported synonymy between Dermocystid sequences from Rum and France and as well as their distinction fromAmphibiocystidiumspp. Phylogenetic analysis also suggested that the amphibian-infecting Dermocystids are not monophyletic. We conclude that theL. helveticus-infecting pathogen represents a single, novel species;Amphibiothecum meredithae

Successful elimination of a lethal wildlife infectious disease in nature

Methods to mitigate the impacts of emerging infectious diseases affecting wildlife are urgently needed to combat loss of biodiversity. However, the successful mitigation of wildlife pathogens in situ has rarely occurred. Indeed, most strategies for combating wildlife diseases remain theoretical, despite the wealth of information available for combating infections in livestock and crops. Here, we report the outcome of a 5-year effort to eliminate infection with Batrachochytrium dendrobatidis affecting an island system with a single amphibian host. Our initial efforts to eliminate infection in the larval reservoir using a direct application of an antifungal were successful ex situ but infection returned to previous levels when tadpoles with cleared infections were returned to their natal sites. We subsequently combined antifungal treatment of tadpoles with environmental chemical disinfection. Infection at four of the five pools where infection had previously been recorded was eradicated, and remained so for 2 years post-application.This work was funded by Fundacio´n General CSIC, Banco Santander and BiodivERsA project RACE.Peer reviewe

Evidence for gene flow differs from observed dispersal patterns in the Humboldt penguin, Spheniscus humboldti

Erworben im Rahmen der Schweizer Nationallizenzen (http://www.nationallizenzen.ch)The Humboldt penguin, once common throughout its range, is today listed as Vulnerable by the IUCN. Mark-recapture and telemetry studies indicate that adult Humboldt penguins are sedentary, suggesting strong genetic differentiation between colonies. We developed genotypes for 336 individuals at 12 microsatellite loci sampled at four different localities spanning the entire range of this species. Results show that long-term gene flow has occurred but appears to be affected by geographic distance as pairwise F ST comparisons involving the colony at Punta San Juan (Peru) and the two colonies at Algarrobo (central Chile) and Puñihuil (southern Chile) are significant. Bayesian estimates of recent migration rates indicate substantial dispersal among all colonies. Despite the dramatic decline in numbers, we did not observe a bottleneck in any population. Furthermore, we did not detect a founder effect in the recently discovered colony at Puñihuil. As our indirect estimates signal strong gene flow between populations, we suggest that Humboldt penguin colonies need to be managed as a metapopulation rather than as discrete management units

Multiple emergences of genetically diverse amphibian-infecting chytrids include a globalized hypervirulent recombinant lineage

Batrachochytriumdendrobatidis (Bd) is a globally ubiquitous fungal infection that has emerged to become a primary driver of amphibian biodiversity loss. Despite widespread effort to understand the emergence of this panzootic, the origins of the infection, its patterns of global spread, and principle mode of evolution remain largely unknown. Using comparative population genomics, we discovered three deeply diverged lineages of Bd associated with amphibians. Two of these lineages were found in multiple continents and are associated with known introductions by the amphibian trade.We found that isolates belonging to one clade, the global panzootic lineage (BdGPL) have emerged across at least five continents during the 20th century and are associated with the onset of epizootics in North America, Central America, the Caribbean, Australia, and Europe. The two newly identified divergent lineages, Cape lineage (BdCAPE) and Swiss lineage (BdCH), were found to differ in morphological traits when compared against one another and BdGPL, and we show that BdGPL is hypervirulent. BdGPL uniquely bears the hallmarks of genomic recombination, manifested as extensive intergenomic phylogenetic conflict and patchily distributed heterozygosity. Wepostulate that contact between previously genetically isolated allopatric populations of Bd may have allowed recombination to occur, resulting in the generation, spread, and invasion of the hypervirulent BdGPL leading to contemporary disease-driven losses in amphibian biodiversity.Peer Reviewe

Exposure to Batrachochytrium dendrobatidis affects chemical defences in two anuran amphibians, Rana dalmatina and Bufo bufo

BACKGROUND: Batrachochytrium dendrobatidis (Bd) is the causative agent of chytridiomycosis, one of the major causes of worldwide amphibian biodiversity loss. Many amphibians exhibit skin-based chemical defences, which may play an important role against invading pathogens, but whether the synthesis of these chemical compounds is enhanced or suppressed in the presence of pathogens is largely unknown. Here we investigated direct and indirect effects of larval exposure to the globally distributed and highly virulent Bd-GPL strain on skin secreted chemical defences and life history traits during early ontogeny of agile frogs (Rana dalmatina) and common toads (Bufo bufo). RESULTS: Exposure to Bd during the larval stage did not result in enhanced synthesis of the antimicrobial peptide Brevinin-1 Da in R. dalmatina tadpoles or in increased production of bufadienolides in B. bufo tadpoles. However, exposure to Bd during the larval stage had a carry-over effect reaching beyond metamorphosis: both R. dalmatina and B. bufo froglets contained smaller quantities of defensive chemicals than their Bd-naïve conspecifics in the control treatment. Prevalence of Bd and infection intensities were very low in both larvae and metamorphs of R. dalmatina, while in B. bufo we observed high Bd prevalence and infection intensities, especially in metamorphs. At the same time, we did not find a significant effect of Bd-exposure on body mass or development rate in larvae or metamorphs in either species. CONCLUSIONS: The lack of detrimental effect of Bd-exposure on life history traits, even parallel with high infection intensities in the case of B. bufo individuals, is surprising and suggests high tolerance of local populations of these two species against Bd. However, the lowered quantity of defensive chemicals may compromise antimicrobial and antipredatory defences of froglets, which may ultimately contribute to population declines also in the absence of conspicuous mass-mortality events

Microbiome function predicts amphibian chytridiomycosis disease dynamics

[Background] The fungal pathogenBatrachochytrium dendrobatidis (Bd) threatens amphibian biodiversity and ecosystem stability worldwide. Amphibian skin microbial community structure has been linked to the clinical outcome of Bd infections, yet its overall functional importance is poorly understood. [Methods] Microbiome taxonomic and functional profiles were assessed using high-throughput bacterial 16S rRNA and fungal ITS2 gene sequencing, bacterial shotgun metagenomics and skin mucosal metabolomics. We sampled 56 wild midwife toads (Alytes obstetricans) from montane populations exhibiting Bd epizootic or enzootic disease dynamics. In addition, to assess whether disease-specific microbiome profiles were linked to microbe-mediated protection or Bd-induced perturbation, we performed a laboratory Bd challenge experiment whereby 40 young adult A. obstetricans were exposed to Bd or a control sham infection. We measured temporal changes in the microbiome as well as functional profiles of Bd-exposed and control animals at peak infection. [Results] Microbiome community structure and function differed in wild populations based on infection history and in experimental control versus Bd-exposed animals. Bd exposure in the laboratory resulted in dynamic changes in microbiome community structure and functional differences, with infection clearance in all but one infected animal. Sphingobacterium, Stenotrophomonas and an unclassified Commamonadaceae were associated with wild epizootic dynamics and also had reduced abundance in laboratory Bd-exposed animals that cleared infection, indicating a negative association with Bd resistance. This was further supported by microbe-metabolite integration which identified functionally relevant taxa driving disease outcome, of which Sphingobacterium and Bd were most influential in wild epizootic dynamics. The strong correlation between microbial taxonomic community composition and skin metabolome in the laboratory and field is inconsistent with microbial functional redundancy, indicating that differences in microbial taxonomy drive functional variation. Shotgun metagenomic analyses support these findings, with similar disease-associated patterns in beta diversity. Analysis of differentially abundant bacterial genes and pathways indicated that bacterial environmental sensing and Bd resource competition are likely to be important in driving infection outcomes. [Conclusions] Bd infection drives altered microbiome taxonomic and functional profiles across laboratory and field environments. Our application of multi-omics analyses in experimental and field settings robustly predicts Bd disease dynamics and identifies novel candidate biomarkers of infection. [MediaObject not available: see fulltext.]K.A.B. was funded by a CASE studentship from NERC, NERC Biomolecular Analysis Facility grant (NBAF939) and an E.P. Abraham Junior Research Fellowship from St Hilda’s College, University of Oxford. M.C.F and T.W.J.G. were funded by NERC award NE/E006701/1 and the Biodiversa project RACE: Risk Assessment of Chytridiomycosis to European Amphibian Biodiversity. T.W.J.G was also funded by Research England and NERC NE/S000062/1. D.S.S. and A.L. received funding through the project People, Pollution, and Pathogens financed through the call “Mountains as Sentinels of Change” by the Belmont-Forum (ANR-15-MASC-0001 - P3, DFG-SCHM3059/6-1, NERC-1633948, NSFC-41661144004). D.S.S. holds the AXA Chair for Functional Mountain Ecology funded by the AXA Research Fund through the project GloMEc and M.C.F. is a fellow in the CIFAR ‘Fungal Kingdoms’ Program