Spatioseasonal Comparison of Fecal Resistome and Pathogenome of Raccoon Dogs in Korea

The raccoon dog (Nyctereutes procyonoides) is a medium-sized omnivore native to Asia. Because they live close to human habitation, and therefore, there is likely bidirectional influence between raccoon dogs and humans, it is important to investigate their potential risks. Here, to identify potential risks of carriage of antimicrobial resistance (AMR) and human pathogens by raccoon dogs, we investigated spatioseasonal patterns of fecal resistome (collection of antimicrobial resistance genes: ARGs), pathogenome (collection of virulence factor genes: VFGs), and microbiome (collection of bacterial species) of raccoon dogs inhabiting an urban forest area and a rural rice paddy area in Korea. Metagenomic sequencing revealed that the compositions of fecal resistome, pathogenome, and microbiome were all patterned by season, and we hypothesize that the observed patterns are due to seasonal changes in the diet of omnivorous raccoon dogs. Furthermore, although less pronounced than the seasonal differences, we also observed the geographical differences in the resistome, with aminoglycoside resistance genes being more prevalent in the rural area, which may reflect the geographical difference in selective pressures for AMR emergence, such as the use of manure that may contain antimicrobials in rice cultivation in agricultural areas. Additionally, our network analysis revealed that specific antimicrobial resistance genes were linked to specific bacterial pathogens, e.g., tetA-P to Clostridium. Overall, our study successfully revealed for the first time that not only the microbiome, but also the resistome and pathogenome of wild animals change spatioseasonally, and that the risk of AMR in bacterial pathogens laden by raccoon dogs is spatioseasonal.