Development of metagenomic methods for non-invasive health monitoring of endangered species: Unveiling hidden microbial threats in fecal samples

Metagenomic analysis of feces is emerging as a powerful tool for improving the monitoring of endangered species. A critical aspect in assessing the extinction risk of a species is the analysis of the burden of parasites and pathogens that can negatively affect the health of individuals. However, the identification of pathogens in non-model species using metagenomics is a major challenge due to the lack of reference genome sequences or data limited to distantly related species. In this study, we developed a pipeline for detecting potentially pathogenic bacteria from metagenomic sequences by mapping unassembled reads to available reference genomes. The approach uses the breadth of genome coverage rather than the number of mapped reads for species identification, thereby minimizing false positives due to conserved or repetitive genomic regions. We applied this method to fresh fecal samples of the Iberian desman (Galemys pyrenaicus), a critically endangered semi-aquatic mammal. Our analysis revealed the presence of 19 potentially pathogenic bacterial species, with prevalences ranging from a single individual to 30% of the samples. We also detected some desmans with elevated or altered pathogen loads, suggesting variations in individual health status or different environmental exposures. This work represents a novel application of metagenomic methods for species-level pathogen detection in wildlife using fecal samples. Application of this method across populations and over time for endangered species may provide essential health and epidemiological information to improve conservation strategies.