Full-length 16S ribosomal RNA sequencing of foal synovial fluid shows substantial agreement with microbial culture.

Full-length 16S rRNA sequencing substantially agrees with culture and may aid diagnosis of culture-negative septic arthritis in foals.

This retrospective observational study evaluated full-length 16S ribosomal RNA (rRNA) long-read sequencing as a diagnostic tool for septic arthritis in foals by comparing its findings to conventional microbial culture and cytology. Synovial fluid samples from 50 foals treated at a single equine referral center between 2018 and 2024 were analyzed using a MinION sequencer after DNA extraction. Culture-positive samples demonstrated significantly higher alpha diversity (Shannon index mean difference 0.42) compared to culture-negative samples. Microbiome composition differed significantly by culture and cytology status, though no specific taxa were associated with age, affected joint, or cytology status alone. Klebsiella were significantly more abundant in culture-positive samples. The agreement between positive microbial culture results and the predominant genus identified by sequencing was substantial, with a Gwet AC1 coefficient of 0.774, falling within the probabilistic benchmark interval of 0.6 to 0.8. Importantly, next-generation sequencing (NGS) and culture results were substantially aligned overall. The authors suggest that NGS may serve as a valuable adjunct diagnostic tool, particularly in clinically suspected septic arthritis cases where conventional culture yields negative results—a scenario that occurs due to fastidious organisms, prior antibiotic exposure, or sampling limitations. This study represents a novel application of long-read sequencing technology in equine joint diagnostics and supports further investigation into microbiome-based diagnostics for infectious musculoskeletal conditions in foals.

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