Genotypic and phenotypic diversity of carbapenem-resistant Bacteroides fragilis strains collected from different clinical origins.

Abstract

Objective: Strains of carbapenem-resistant Bacteroides fragilis have frequently emerged in recent years. In China, data on the genotypic and phenotypic characteristics of these antimicrobial-resistant anaerobic bacteria are scarce. Thus, the aim of this study was to characterize clinical isolates of carbapenem-resistant B. fragilis collected from a tertiary hospital in China using whole genome sequencing (WGS), phenotypic susceptibility tests, and a biofilm formation assay.

Methods: We analyzed 49 B. fragilis strains with different antimicrobial resistance profiles. Antimicrobial susceptibility was determined using the agar dilution method and biofilm formation using a crystal violet assay. Genomic characteristics were analyzed using WGS, and the transcription level of cfiA, which is responsible for carbapenem resistance, was determined using quantitative reverse transcription polymerase chain reaction (PCR). Carbapenem-sensitive isolates were used as controls.

Results: All 49 B. fragilis isolates were biofilm producers and the percentage of carbapenem-resistant isolates was 42.86 % (21/49). The percentage of carbapenem-resistant isolates with medium-to-strong biofilm production ability was significantly lower than that of carbapenem-sensitive isolates (19.1 % vs. 88.9 %, p < 0.01). None of the carbapenem-resistant B. fragilis isolates carried bft. In contrast, 53.6 % (15/28) of the carbapenem-sensitive isolates carried bft, and all of them were fpn(+). All carbapenem-resistant isolates (21/21, 100 %) harbored cfiA and its upstream insertion sequence (IS) element. Three isolates (BF058, BF059, and BF060) carried the IS613 element, which was not immediately adjacent upstream to cfiA but was separated by a 1000-kb sequence encoding vatD. The quantitative PCR assay results revealed the elevated expression of cfiA mRNA among carbapenem-resistant isolates, although the relative expression levels varied greatly among isolates. However, a significant correlation between the relative expression level of cfiA mRNA and phenotypic carbapenem resistance was observed.

Conclusions: Carbapenem-resistant B. fragilis isolates carried a low frequency of virulence-related genes and showed weaker biofilm formation ability compared with carbapenem-sensitive B. fragilis isolates. CfiA was the dominant mediator of carbapenem resistance in B. fragilis. This study was the first to identify the structural plasticity of the cfiA-IS element, emphasizing the diverse and complex evolution of carbapenem resistance in B. fragilis, which warrants further investigation.