# Genomic and phenotypic diversity among taxonomically ambiguous clinical Corynebacterium isolates

## TL;DR

This study examines 56 clinical corynebacteria isolates that could not be clearly identified, revealing significant genomic and phenotypic diversity, including potential new species and high antimicrobial resistance.

## Contribution

The study identifies 28 distinct corynebacterial species from clinical isolates and highlights potential novel species and high antimicrobial resistance.

## Key findings

- 56 isolates belonged to 28 distinct corynebacterial species, with 13 showing ambiguous species identification.
- Two isolates may represent novel species due to low ANI to known genomes.
- High antimicrobial resistance was observed, particularly against clindamycin, penicillin, and ciprofloxacin.

## Abstract

Corynebacterium is a widespread and abundant bacterial genus on human skin. Occasionally, corynebacteria are isolated from clinical specimens associated with infection. In this study, 56 bacterial isolates were examined. These isolates were obtained from 52 patients with diverse infections such as keratitis, osteitis/osteomyelitis, mastitis, (suspected) foreign body associated infections (spine, prosthetic joint), suspected meningitis, post-operative infections, among others. These isolates were identified as corynebacteria by MALDI-TOF mass spectrometry but could not be reliably assigned to a specific species. To resolve this issue, the isolates were genome-sequenced, and species identification was done with different approaches, including digital DNA–DNA hybridization, phylogenomic tree placement and Average Nucleotide Identity (ANI) calculations. A subset of 34 strains was further investigated by biochemical characterization and antimicrobial susceptibility testing (AST).

The 56 isolates belonged to 28 distinct corynebacterial species. Species identification was particularly ambiguous for 13 isolates as the ANIs were below 95% to the closest identified reference genomes. Two isolates represented potentially novel species, since no close relative could be identified (ANI < 90%). The majority of isolates belonged to the Corynebacterium marquesiae/tuberculostearicum (n = 10) and Corynebacterium kroppenstedtii/parakroppenstedtii (n = 9) complexes. Biochemical tests and AST revealed species- and strain-level variability. AST demonstrated extensive antimicrobial resistance (AMR), particularly among C. marquesiae, C. tuberculostearicum, C. lehmanniae, C. hesseae and C. resistens, with resistances observed against penicillin, clindamycin, ciprofloxacin, and rifampin. Resistance was frequently associated with acquired AMR genes, such as erm(X), tet(W) and genes encoding aminoglycoside-modifying enzymes. Among the tested antibiotics, clindamycin resistance was most common, detected in 23 of 34 tested strains (64.7%).

This study expands our knowledge of Corynebacterium isolates derived from clinical specimens, particularly those differing from well-characterized species. It underscores the extensive geno- and phenotypic variability within most Corynebacterium species and challenges current species boundary definitions. The extensive level of detected AMR may complicate treatment of underlying infections. However, it remains uncertain whether these isolates represent true infectious agents or contaminants derived from the skin of the patients.

The online version contains supplementary material available at 10.1186/s12866-025-04619-8.