Emerging Clostridioides difficile strains belonging to PCR ribotype 955 in Serbia are distinct from metronidazole resistant RT955 outbreak isolates from the UK

End 2023, the UK Health Security Agency sent an alert about a new hypervirulent Clostridioides difficile PCR ribotype, ribotype 955 (RT955), causing slowly progressing infection clusters in hospitals in the Midlands. Between March 2018 and February 2022, CDI surveillance was performed in southern Serbia with centers providing medical services for approximately 750,000 inhabitants. Using the ECDC recommended protocol, clinical, epidemiological and microbiological data were collected. C. difficile RT955 was identified in 27 (7%) of 383 surveyed patients with CDI. Of 27 patients, 16 (59%) was older than 60 years and 19 (70%) were male. CDI was always associated with previous antibiotic therapy and had a hospital onset in 23 (85%) patients. The clinical presentaiton was milder than reported in UK and at 90 days follow up, no CDI related mortality was found. All sequenced strains belonged to multilocus sequence type (ST) 1 and were highly similar, with 0-1 alleles differences in a cgMLST analysis. The strains differed clearly from the UK RT955 outbreak strain by whole genome sequencing and phenotypic resistance to lincosamides and rifampicin. Interestingly, a high level erythromycin resistance was observed without a genetic resistance determinant. Both the UK and Serbian RT955 strains contained gyrA_p.T82I associated with resistance to fluoroquinolone antimicrobials and carried the PnimBG promoter mutation, suggestive for haem-dependent metronidazole resistance. We conclude that C. difficile RT955 is present in Serbia since 2018, without presentation of large outbreaks. The Serbian RT955 strains differed clearly from a representative UK cluster strain, but shared its haem dependent metronidazole resistance.