Project description:Aliarcobacter butzleri is an emerging gastrointestinal pathogen found in many countries worldwide. In France, it has become the third most commonly isolated bacterial species from the stools of patients with intestinal infections. No interpretative criteria for antimicrobial susceptibility testing have been proposed for A. butzleri, and most strains are categorized using the recommendations of the Clinical and Laboratory Standards Institute or the European Committee on Antimicrobial Susceptibility Testing for Campylobacter or Enterobacterales. In the present study, the genomes of 30 resistant A. butzleri isolates were analyzed to propose specific epidemiological cut-off values for ampicillin, ciprofloxacin, erythromycin, and tetracycline. The identification of a β-lactamase and the T85I GyrA mutation associated with ampicillin and ciprofloxacin resistance, respectively, allowed us to adjust the disk diffusion (DD) and MIC cut-off values for these molecules. However, epidemiological cut-off values for erythromycin and tetracycline could not be estimated due to the absence of known resistance mechanisms. The present study paves the way for building a consensus for antimicrobial susceptibility testing for this concerning pathogen. IMPORTANCE Aliarcobacter butzleri is an emerging and concerning intestinal pathogen. Very few studies have focused on this particular species, and antimicrobial susceptibility testing (AST) is based on methods that have been mostly developed for Campylobacter spp. In fact, no disk diffusion and E-tests adapted cut-offs for A. butzleri are available which leads to misinterpretations. We have shown here that NGS approach to identify genes and mutations in close relation to phenotypic resistance levels is a robust way to solve that issue and precisely differentiate WT and NWT A. butzleri isolates for frequently used antimicrobials. MIC and DD cut-off values have been significantly adjusted and answer the need for a global consensus regarding AST for A. butzleri.
Project description:Aliarcobacter butzleri is an emergent enteropathogen that can be found in a range of environments. This bacterium presents a vast repertoire of efflux pumps, such as the ones belonging to the resistance nodulation cell division family, which may be associated with bacterial resistance, as well as virulence. Thus, this work aimed to evaluate the contribution of three RND efflux systems, AreABC, AreDEF and AreGHI, in the resistance and virulence of A. butzleri. Mutant strains were constructed by inactivation of the gene that encodes the inner membrane protein of these systems. The bacterial resistance profile of parental and mutant strains to several antimicrobials was assessed, as was the intracellular accumulation of the ethidium bromide dye. Regarding bacterial virulence, the role of these three efflux pumps on growth, strain fitness, motility, biofilm formation ability, survival in adverse conditions (oxidative stress and bile salts) and human serum and in vitro adhesion and invasion to Caco-2 cells was evaluated. We observed that the mutants from the three efflux pumps were more susceptible to several classes of antimicrobials than the parental strain and presented an increase in the accumulation of ethidium bromide, indicating a potential role of the efflux pumps in the extrusion of antimicrobials. The mutant strains had no bacterial growth defects; nonetheless, they presented a reduction in relative fitness. For the three mutants, an increase in the susceptibility to oxidative stress was observed, while only the mutant for AreGHI efflux pump showed a relevant role in bile stress survival. All the mutant strains showed an impairment in biofilm formation ability, were more susceptible to human serum and were less adherent to intestinal epithelial cells. Overall, the results support the contribution of the efflux pumps AreABC, AreDEF and AreGHI of A. butzleri to antimicrobial resistance, as well as to bacterial virulence.