Antimicrobial-resistant genes in feces from otters (Lontra longicaudis) within the Peñas Blancas river basin, Costa Rica

Antimicrobial resistance poses a growing threat to human health, yet its implications for wildlife remain a subject of ongoing research. River otters inhabiting the Peñas Blancas River face exposure to various anthropogenic activities in their habitat, potentially leading to the accumulation of antibiotic-resistant genes (ARGs) with unknown consequences for their health. This study aimed to identify specific ARGs in otter feces from this river basin, employing quantitative polymerase chain reaction (qPCR), DNA sequencing of ARGs, and phylogenetic analysis techniques. Over the period from 2019 to 2022, we collected 102 fecal samples from otters through the Peñas Blancas River watershed, spanning its upper and middle basins. We assessed the bacterial presence via the 16S rRNA gene through qPCR analysis and screened for 12 ARGs. Sequences of 16 ARG-positive samples were subsequently analyzed using Maximum-likelihood-base taxonomic placement. In total, 56 samples tested positive for the 16S rRNA gene, with 24 exhibiting at least one ARG. Notably, three samples showcased a "multi-resistance microbiome". qPCR analyses identified seven distinct ARGs: <i>tetB</i> (in 26.8 % of the samples), <i>sulI</i> (21.4 %), <i>sulII</i> (21.4 %), <i>qnrS</i> (10.7 %), <i>tetQ</i> (8.9 %), <i>tetW</i> (7.1 %), and <i>tetA</i> (3.6 %). Phylogenetic analysis confirmed the taxonomic association of all detected ARGs, which were compared with The Comprehensive Antibiotic Resistance Database. Our findings underscore the importance of comprehending the spread of ARGs in wildlife populations, with river otters serving as potential sentinels for ARG dissemination. Moreover, they highlight the potential impact of anthropogenic activities on the health of aquatic ecosystems, emphasizing the need for proactive measures to mitigate antimicrobial resistance in natural environments.