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. 2023 Nov 9:14:1285236.
doi: 10.3389/fmicb.2023.1285236. eCollection 2023.

Combined genomic-proteomic approach in the identification of Campylobacter coli amoxicillin-clavulanic acid resistance mechanism in clinical isolates

Francis Deforet  1 , Quentin Jehanne  2 , Lucie Bénéjat  2 , Johanna Aptel  2 , Roxane Prat  3 , Chloé Desbiolles  3 , Astrid Ducournau  2 , Marine Jauvain  2   4 , Richard Bonnet  5 , François Vandenesch  3 , Jérôme Lemoine  1 , Philippe Lehours  2   4
Affiliations

Combined genomic-proteomic approach in the identification of Campylobacter coli amoxicillin-clavulanic acid resistance mechanism in clinical isolates

Francis Deforet et al. Front Microbiol. .

Abstract

Introduction: Aminopenicillins resistance among Campylobacter jejuni and Campylobacter coli strains is associated with a single mutation in the promoting region of a chromosomal beta-lactamase blaOXA61, allowing its expression. Clavulanic acid is used to restore aminopenicillins activity in case of blaOXA61 expression and has also an inherent antimicrobial activity over Campylobacter spp. Resistance to amoxicillin-clavulanic acid is therefore extremely rare among these species: only 0.1% of all Campylobacter spp. analyzed in the French National Reference Center these last years (2017-2022).

Material and methods: Whole genome sequencing with bioinformatic resistance identification combined with mass spectrometry (MS) was used to identify amoxicillin-acid clavulanic resistance mechanism in Campylobacters.

Results: A G57T mutation in blaOXA61 promoting region was identified in all C. jejuni and C. coli ampicillin resistant isolates and no mutation in ampicillin susceptible isolates. Interestingly, three C. coli resistant to both ampicillin and amoxicillin-clavulanic acid displayed a supplemental deletion in the promoting region of blaOXA61 beta-lactamase, at position A69. Using MS, a significant difference in the expression of BlaOXA61 was observed between these three isolates and amoxicillin-clavulanic acid susceptible C. coli.

Conclusion: A combined genomics/proteomics approach allowed here to identify a rare putative resistance mechanism associated with amoxicillin-clavulanic acid resistance for C. coli.

Keywords: AMR; Campylobacter; amoxicillin-clavulanic acid; beta-lactamase; gene expression.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Alignment of blaOXA61 promoting sequences in 18 C. jejuni and 12 C. coli included in the present study. For each isolate, ampicillin and amoxicillin-clavulanic acid MICs are indicated. Promoting regions were extracted from every assembled genome using Nucleotide-Nucleotide BLAST 2.12.0+ (Altschul et al., 1997), and sequences were aligned using Muscle v3.8.1551 (Edgar, 2004). Relevant genotypes are here highlighted in white at positions −57 and −69.
Figure 2
Figure 2
BlaOXA61 relative quantification in ampicillin-susceptible and resistant C. jejuni isolates. (A) Each strain individually. (B) Strains grouped according to the blaOXA61-promoting sequences. Data are expressed in arbitrary units. In panel (B), the mean of triplicates was used to perform a Mann–Whitney U test. ***p value <0.001. wt, wild type; S, susceptible; R, resistant. No signal was observed for C. jejuni reference CCUG 11284.
Figure 3
Figure 3
Expression levels of BlaOXA61 in C. coli isolates with wild type, G57T or G57T + ∆A69 promoters. (A) Each strain individually. (B) Strains grouped according to the blaOXA61-promoting sequences. Data are expressed in arbitrary units. In panel (B), the mean of triplicates was used to perform one-way ANOVA. ***p-value <0.001. wt, wild type; AMP, ampicillin; AMC, amoxicillin + clavulanic acid; S, susceptible; R, resistant.
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