First Report on Multi-Drug Resistant Paraclostridium bifermentans in Avian Faeces: A Comparative Analysis with Co-Resident Clostridium perfringens

Introduction: Poultry production can serve as an important reservoir of antimicrobial-resistant bacteria. Although surveillance commonly focuses on prevalent organisms such as Clostridium perfringens, less prevalent anaerobic spore-forming bacteria may also contribute to the antimicrobial resistance reservoir. This study characterised Paraclostridium bifermentans recovered from broiler chicken faeces and compared its antimicrobial resistance profiles with those of co-resident C. perfringens. Methods: Ninety-six pooled avian faecal samples were screened for P. bifermentans. Isolates were identified using biochemical testing and 16S rRNA gene sequencing. Antimicrobial susceptibility was assessed against five antibiotic classes, and selected antimicrobial resistance genes (ARGs) were detected using conventional PCR. Phenotypic resistance profiles were compared with those of 52 previously characterised C. perfringens isolates recovered from the same sampling population. Results: Six P. bifermentans isolates were recovered from 96 pooled samples (6.25%) and showed 99.9% 16S rRNA gene sequence similarity to reference P. bifermentans sequences. All six isolates were multi-drug-resistant, compared with 24/52 (46.15%) C. perfringens isolates (Fisher’s exact test, p = 0.024). P. bifermentans showed significantly higher resistance to chloramphenicol (100% vs. 34.62%, p = 0.003) and clindamycin (100% vs. 46.15%, p = 0.024) than C. perfringens. Resistance genes were variably detected among the P. bifermentans isolates. All six isolates carried tet(A), while tet(W) was detected in 33.3%. The blaOXA gene was detected at 66.7%, and catIV and catII were detected in 66.7% and 33.3%, respectively. Phenotypic resistance was not consistently explained by the ARGs targeted in this study. Conclusions: The recovery of multi-drug-resistant P. bifermentans from broiler faeces indicates that this less frequently recovered organism may represent an under-recognised component of the antimicrobial resistance reservoir in the investigated poultry environment. However, the small number of isolates limits generalisation. Further studies using larger sample sizes and whole-genome or functional approaches are required to determine the diversity, genomic context, expression, and potential mobility of resistance determinants in P. bifermentans.