Population genetics as a tool to elucidate pathogen reservoirs: Lessons fromPseudogymnoascus destructans, the causative agent of white-nose disease in bats

Abstract

Emerging infectious diseases pose a major threat to human, animal, and plant health. The risk of species-extinctions increases when pathogens can survive in the absence of the host, for example in environmental reservoirs. However, identifying such reservoirs and modes of infection is often highly challenging. In this study, we investigated the presence and nature of an environmental reservoir for the ascomycete fungus Pseudogymnoascus destructans, the causative agent of white-nose disease. We also characterised the modes and timing of transmission of the pathogen; key elements to better understand the disease dynamics. Using 18 microsatellite markers, we determined the genotypic and genic (based on allele frequencies) differentiation between 1,497 P. destructans isolates collected from nine closely situated hibernacula in North-Eastern Germany. One hibernaculum was the focus of intensive sampling in which both the bats and walls of the site were sampled at regular intervals over five consecutive winter seasons (1,062 isolates). We found significant genic differentiation between sites and few multi-locus genotypes shared across hibernacula (genotypic differentiation). This demonstrates that each hibernaculum has an essentially unique population of the fungus. This would be expected if bats purge viable P. destructans over the summer, preventing the mixing and exchange of the pathogen in maternity colonies, where bats from all of the studied hibernacula meet. Results from the intensively sampled site show higher measures of genotypic richness on walls compared to bats, the absence of genic differentiation between bats and walls, and stable relative abundance of multi-locus genotypes over multiple winter seasons. This clearly implicates hibernacula walls as the main environmental reservoir of the pathogen, from which bats become re-infected annually.

Population genetics as a tool to elucidate pathogen reservoirs: Lessons fromPseudogymnoascus destructans, the causative agent of White‐Nose disease in bats

“…The tidyverse collection of packages (Wickham et al, 2019) was used to improve ease and efficiency in analyses and the corrplot package (version 0.90, Wei & Simko, 2017) was used for visualization of correlations of relative frequencies of genotypes occurrence. The R-script used for analyses as well as the raw data are available from Dryad (https://doi.org/10.5061/dryad.x0k6d jhhx; Fischer et al, 2021).…”

“…To examine this, we simulated bottlenecks of different strengths (i.e., sampled isolates; in steps of 1 from 2 to 3100) by subsampling the pool of genotyped isolates from the walls in Eldena, mimicking the subset of spores passed from the walls to the bats (each winter season investigated separately). As previous studies have shown that only a fraction of spores germinates, we subsequently randomly selected 17.5% of the previously subsampled isolates for further analyses (based on the mean germination rate for spores of P. destructans from the data in Fischer et al, 2020; mean = 17.5%). For each value of bottleneck strength (i.e., sample size), we then calculated the genotypic richness (i.e., number of genotypes) observed in the subsampled data set.…”

“…So far in nature, active growth of the fungus (after germination of spores) and spore production have only been observed on bats, which are the only known hosts of P. destructans (e.g., Fischer et al, 2020;Palmer et al, 2018;Puechmaille et al, 2011;Reynolds & Barton, 2014). Only asexual reproduction has been observed in this haploid species (e.g., Gargas et al, 2009), and the resulting spores are likely to remain viable in the environment (dormancy) for several years at least (Fischer et al, 2020). Therefore, it becomes important to integrate information about host behaviour and life cycle to better understand and characterise the location and mode of transmission of P. destructans (e.g., Fuller et al, 2020;Hoyt et al, 2020;Puechmaille et al, 2011; Figure 1).…”

“…This behaviour is hypothesised to result in bats becoming infected from the environment (Puechmaille et al, 2011). The fact that P. destructans spores persist on hibernacula walls (Vanderwolf et al, 2016), even when bats are absent, and that the seasonal pattern of P. destructans germination is synchronised with the presence of bats in hibernacula (Fischer et al, 2020), lends support to the inferred wall-to-bat mode of infection after summer (see also Hoyt et al, 2020). Additionally, sharp increases in P. destructans prevalence (estimated via qPCR) on bats in autumn further supports the inferred timing of infection (swarming behaviour at swarming/hibernation sites; see also although viability of the fungus on bats during this period remains to be confirmed.…”

“…There are still large gaps in our understanding of P. destructans biology. So far in nature, active growth of the fungus (after germination of spores) and spore production have only been observed on bats, which are the only known hosts of P. destructans (e.g., Fischer et al, 2020;Palmer et al, 2018;Puechmaille et al, 2011;Reynolds & Barton, 2014). Only asexual reproduction has been observed in this haploid species (e.g., Gargas et al, 2009), and the resulting spores are likely to remain viable in the environment (dormancy) for several years at least (Fischer et al, 2020).…”