Genomic characterisation of Campylobacter jejuni Cj26: A high-level ciprofloxacin/erythromycin-resistant strain isolated from a poultry carcass in southern Brazil.

OBJECTIVES: The draft genome sequence of Campylobacter jejuni (Cj26) was analysed to investigate genetic mechanisms of antimicrobial resistance, virulence-associated genes, and phylogenetic context. METHODS: Antimicrobial resistance was assessed by agar dilution and disk diffusion. Cj26 was sequenced using NovaSeq 6000 technology. The genome was assembled and annotated. Resistance genes and chromosomal mutations were analysed using the Center for Genomic Epidemiology services, and the multilocus sequence type, SVR-flaA, and porA were determined. The virulome was determined using the Virulence Factor Database. Plasmid detection and assembly were performed using Unicycler v0.5.0 software. To infer the core genome phylogeny, prokka v1.14.5 was employed in conjunction with IQtree v2.0.3. RESULTS: The Cj26 strain showed a high level of resistance to ciprofloxacin (32 µg/mL) and erythromycin (>128 µg/mL) and resistance to tetracycline and ampicillin. Multilocus sequence typing revealed that the strain belonged to sequence type 353. The substitutions Tre-86-Ile in gyrA and A2075G in 23s RNA were detected, along with the genes tetO, aph(3')-III, ant(6)-Ia, and blaOXA 460. A consistent relationship among accessory and core genes was identified. When compared to other sequence type 353 genomes from Brazil, Cj26 clustered with strains that had more antimicrobial resistance genes than the other clusters. CONCLUSION: This report provides insight into the antimicrobial resistance determinants found in a C. jejuni strain and offers a valuable resource for further studies on Campylobacter genomics and antimicrobial resistance.

SVR-flaA typing of erythromycin- and ciprofloxacin-resistant Campylobacter jejuni strains isolated from poultry slaughterhouses in southern Brazil.

The emergence of fluoroquinolone and macrolide resistance in C. jejuni, a recognized zoonotic pathogen, has increased worldwide. This study aimed to investigate phenotypic resistance to ciprofloxacin and erythromycin, the molecular mechanisms involved, and the strain of C. jejuni isolated from broiler carcasses. Eighty C. jejuni isolates from broiler carcasses in southern Brazil were investigated for their susceptibility to ciprofloxacin and erythromycin at minimal inhibitory concentrations. Mismatch amplification mutation assay-polymerase chain reaction (MAMA-PCR) was performed to detect substitutions of Thr-86-Ile, A2074C, and A2075G of domain V in the 23S rRNA. The presence of ermB gene and CmeABC operon were investigated by PCR. DNA sequencing was used to detect substitutions in the L4 and L22 proteins of the erythromycin-resistant strains. The Short Variable Region (SVR) of flaA was used to type all the strains resistant to both antimicrobials. Ciprofloxacin and erythromycin resistance were detected in 81.25% and 30.00% of the strains, respectively, and minimal inhibitory concentration values ranged from ≤ 0.125 to 64 µg/mL for ciprofloxacin and 0.5 to > 128 µg/mL for erythromycin. The Thr-86-Ile mutation in gyrA was observed in 100% of the ciprofloxacin-resistant strains. Mutations in both the A2074C and A2075G positions of 23S rRNA were observed in 62.5% of the erythromycin-resistant strains, while 37.5% had only the mutation A2075G. None of the strains harbored CmeABC operon, and ermB was not detected. Using DNA sequencing, the amino acid substitution T177S was detected in L4, and substitutions I65V, A103V, and S109A were detected in L22. Twelve flaA-SVR alleles were identified among the strains, with the most common SVR-flaA allele, type 287, covering 31.03% of ciprofloxacin- and erythromycin-resistant isolates. The present study revealed a high incidence and high levels of resistance to ciprofloxacin and erythromycin, as well as broad molecular diversity in C. jejuni isolates from broiler carcasses.

Molecular markers associated with antimicrobial resistance and genotypes of Campylobacter jejuni and Campylobacter coli isolated from broiler and swine flocks in southeast Brazil.

This study aimed to identify molecular markers associated with antimicrobial resistance and genotype isolates of Campylobacter spp. from broiler and swine flocks due to its importance to one-health. C. jejuni (n=27) and C. coli (n = 35) strains were screened for the antimicrobial genetic markers C257T in gyrA, A2074C and A2075G in 23S rRNA, CmeABC, ermB, tetO and blaOXA61 by PCR. Fifteen strains had SVR-flaA and porA genes sequenced to evaluate their genetic diversity. Among C. jejuni strains 62.96% had C257T mutation and only one strain had A2075G mutation. CmeA, cmeB, cmeC, tetO and blaOXA61 were detected respectively in 92.59%, 100%, 100%, 85.19%, 85.19% of the strains. All C. coli had C257T mutation; 48.75% had A2075G and cmeA, cmeB, cmeC, tetO, blaOXA61 were detected in 8.57%, 94.29%, 91.43%, 91.43%, 80%, respectively. Twelve porA and SVR-flaA alleles were detected, with a Simpson index of diversity value of 0.962.