Culture and Metagenomic Insights into the Ear Microbiota in Dogs with Healthy Ears and Otitis Externa

The canine ear microbiota plays an important role in ear health, and dysbiosis is associated with otitis externa (OE) and antimicrobial resistance (AMR). This study aims to investigate the ear microbiota of dogs with healthy ears and OE using bacterial culture-based methods and shotgun metagenomic sequencing, and to screen for AMR and virulence-associated genetic signatures. Ear swab specimens from 100 healthy and 100 OE-affected dogs were analyzed. The isolates obtained via bacterial culture were identified by MALDI-TOF MS and 16S rRNA sequencing. Metagenomic analysis was performed via Illumina shotgun sequencing. The most commonly defined species in healthy dogs in culture were Staphylococcus pseudintermedius (24.5%) and Staphylococcus epidermidis (5.7%); in dogs with OE, the most commonly defined species were S. pseudintermedius (30.5%), and Clostridium perfringens (4.5%). In healthy samples, metagenomic analysis revealed higher relative abundances of Bacteroides fragilis (15.8%) and Ezakiella coagulans (8.2%), while S. pseudintermedius (38.7%) dominated in OE. AMR profiling demonstrated diverse resistance determinants, including efflux pump systems and methicillin resistance-associated genes. In conclusion, the present study shows that S. pseudintermedius is a predominant member of canine ear microbiota, with higher presence in OE highlighting microbial shifts, and demonstrates that combining culture and metagenomic analyses provides a concise view of microbial communities and clinical relevance.