Comparative Genomics of Bacteroides fragilis Group Isolates Reveals Species-Dependent Resistance Mechanisms and Validates Clinical Tools for Resistance Prediction

Bacteroides fragilis group (BFG) are the most frequently recovered anaerobic bacteria from human infections, and resistance to frontline antibiotics is emerging. In the absence of routine antimicrobial susceptibility testing (AST) for BFG in most clinical settings, we assessed the utility of clinical and modern genomics tools to determine BFG species-level identification and resistance patterns. A total of 174 BFG clinical isolates supplemented with 20 archived carbapenem-resistant B. fragilis sensu stricto (BFSS) isolates underwent antimicrobial susceptibility testing, MALDI-ToF mass-spectrometry, and whole-genome sequencing (WGS). Bruker BioTyper and VITEK-MS MALDI-ToF systems demonstrated accurate species-level identifications (91% and 90% agreement, respectively) compared to average nucleotide identity (ANI) analysis of WGS data. Distinct β-lactamase gene profiles were observed between BFSS and non-fragilis Bacteroides species, with significantly higher MICs to piperacillin-tazobactam in B. vulgatus and B. thetaiotaomicron relative to BFSS (P ≤ 0.034). We also uncovered phylogenetic diversity at the genomospecies level between division I and division II BFSS (ANI <0.95) and demonstrate that division II BFSS strains harbor an increased capacity to achieve carbapenem resistance through chromosomal activation of the CfiA carbapenemase. Finally, we report that CfiA detection by the Bruker BioTyper Subtyping Module accurately detected carbapenem resistance in BFSS with positive and negative percent agreement of 94%/90% and 95%/95% compared to ertapenem and meropenem susceptibility, respectively. These comparative analyses indicate that resistance mechanisms are distinct at both the phenotypic and genomic level across species within the BFG and that modern MALDI-ToF identification systems can be used for accurate species-level identification and resistance prediction of the BFG.