Michael Zarske (Berlin / DE), Christiane Werckenthin (Oldenburg / DE), Julia Golz (Berlin / DE), Kerstin Stingl (Berlin / DE)
Introduction:
Pheno- and genotypic analysis of thermotolerant Campylobacter spp. identified a C. coli isolate from turkey caecal content to exhibit resistance to gentamicin (GEN) without harbouring known resistance determinants.
Goals:
The study aimed to identify the molecular determinant conferring resistance to GEN in the C. coli isolate. Also, cross-resistance to other aminoglycosides, transferability of the resistance among Campylobacter spp. and its persistence upon in vitro passaging were characterized.
Materials & Methods:
Antimicrobial susceptibility was tested using microdilution. Whole genome sequencing (WGS) was performed with Illumina NextSeq and Oxford Nanopore Technology. Genomic DNA (gDNA) or a PCR fragment from the resistant donor, harbouring the 16S rRNA gene, was transferred from the resistant isolate to sensitive recipient C. spp. isolates using natural transformation. Transformants were selected on 8-16 mg/L tobramycin (TOB) and reanalysed by WGS. Sanger sequencing was performed to assess transitions in 16S rRNA copies in the transformants upon passaging.
Results:
C. coli BfR-CA-15687 had been isolated during the zoonosis monitoring from turkey caecal content in 2018. Antimicrobial susceptibility tests showed resistance to apramycin (APR), GEN, kanamycin (KAN) and TOB, while the isolate was sensitive to streptomycin (STR). The resistance was naturally transformable to sensitive recipient C. jejuni and C. coli using BfR-CA-15687 gDNA. The transformation rate was ~2 log10 lower compared to the control rpsL-A128G point mutation, conferring STR resistance. SNP analysis of WGS data revealed a novel A1387G mutation in the 16S rRNA gene associated with APRR-GENR-KANR-TOBR resistance. Transformation of this mutation within a 16S rRNA PCR fragment showed causal relationship between 16S rRNA_A1387G and resistance. Sanger sequencing revealed A1387G transition in all three or less copies of the 16S rRNA genes in transformants, impacting resistance stability.
Summary:
We unveiled a novel, rare APRR-GENR-KANR-TOBR resistance mechanism in C. coli, mediated by the point mutation 16S rRNA_ A1387G, which was transferable via natural transformation.