Phenotypic and Whole-Genome Sequencing-Based Profiling of Antimicrobial Resistance and Virulence in Campylobacter spp. Isolated from Human Stool in a Croatian University Hospital.

Abstract

Campylobacter spp. is a leading cause of community-acquired gastroenteritis, with rising antimicrobial resistance (AMR) posing a significant public health challenge. This study aimed to characterize the genomic basis of resistance and virulence in Campylobacter spp. isolated from human stool at the University Hospital of Split, Croatia. A total of 115 unduplicated isolates (100 Campylobacter jejuni and 15 Campylobacter coli) were identified and tested for antimicrobial susceptibility, from January to June 2023. A representative number of isolates with phenotypically detected resistance were analyzed with whole-genome sequencing. Multilocus sequence typing (MLST) was determined using genomic sequences as input data against the standard MLST schemes. High rates of resistance were detected to tetracycline (45% of C. jejuni and 26.7% of C. coli) and ciprofloxacin (81% of C. jejuni and 46.7% of C. coli), while combined ciprofloxacin/tetracycline resistance was observed in 43% of C. jejuni and 7% of C. coli isolates. The majority of whole-genome sequenced isolates possess tet(O/32/O) gene and gyrA.T861 mutation, conferring resistance to tetracycline and fluoroquinolones. All isolates have at least one resistance gene for β-lactams (bla_OXA-61, bla_OXA-193, bla_OXA-450, bla_OXA-451, bla_OXA-452, bla_OXA-461, and bla_OXA-489), while 40% of isolates possess ant(6)-Ia for aminoglycoside resistance. In addition, a newly emerged, phenotypically identified multidrug-resistant C. coli, harbored tet(O/32/O) and tet(O) genes, point mutation in gyrA gene, and A2075G mutation in 23S rRNA. Virulence analysis highlighted C. jejuni's broad pathogenic potential, including motility, adherence, invasion, exotoxin production, and immune modulation (e.g., wlaN gene, linked to Guillain-Barré syndrome). On the contrary, C. coli isolates possess genes exclusively for motility. Nine different C. jejuni MLST sequence types were identified, while all C. coli isolates belong to the same 828 clonal complex. Escalating AMR and a broad spectrum of virulence in Campylobacter spp. highlight the importance of continuous surveillance on the phenotypic and genotypic level, thereby allowing more efficient health care management of these infections.