Corynebacterium rouxii is one of the recently described species of the Corynebacterium diphtheriae complex. As this species can potentially infect different hosts and harbor the tox gene, producing diphtheria toxin, we present its first pangenomic analysis in this work. A total of fifteen genomes deposited in online databases were included. After confirming the taxonomic position of the isolates by genomic taxonomy, the genomes were submitted to genomic plasticity, gene synteny, and pangenome prediction analyses. In addition, virulence and antimicrobial resistance genes were investigated. Finally, epidemiological data were obtained through molecular typing, clustering, and phylogenetic analysis. Our data demonstrated genetic diversity within the species with low synteny. However, the gene content is extensively conserved, and the pangenome is composed of 2606 gene families, of which 1916 are in the core genome and 80 are related to unique genes. Prophages, insertion sequences, and genomic islands were found. A type I-E CRISPR-Cas system was also detected. Besides the tox gene, determinants involved in adhesion and iron acquisition and two putative antimicrobial resistance genes were predicted. These findings provide valuable insight about this species’ pathogenicity, evolution, and diversity. In the future, our data can contribute to different areas, including vaccinology and epidemiology.

Background: Corynebacterium ulcerans is an emerging zoonotic pathogen capable of cau-sing diphtheria-like infections in humans. Objectives: we report, for the first time in Brazil, the detection and phenotypic/genomic characterization of three atoxigenic ST-339 strains isolated from domestic animals, including one with a ciprofloxacin resistance profile linked to double GyrA mutations (S89L, D93G). Methods: species identification was performed by MALDI-TOF MS, followed by in vitro antimicrobial susceptibility testing, whole-genome sequencing, and bioinformatic analyses to predict virulence determinants, antimicrobial resistance genes, CRISPR–Cas systems, mobile genetic elements, and in silico structural analysis as well as phylogenetic reconstruction. Results: whole-genome sequencing confirmed species identity, revealed high genetic similarity, and identified distinct phylogenetic subclades, suggesting potential international dissemination. Genomic analyses showed conserved virulence determinants, such as incomplete pilus clusters, iron acquisition systems, and the pld gene, with the absence of the tox gene. Molecular modeling and dynamics simulations indicated that GyrA mutations disrupt critical ciprofloxacin–magnesium–water interactions, reducing binding stability. Mobile genetic elements, prophages, and CRISPR–Cas systems underscored the genomic plasticity of these isolates. Conclusions: these findings document a little-studied antimicrobial resistance mechanism in zoonotic C. ulcerans, highlighting the need for strengthened surveillance and further research on virulence and resistance, even in ato-xigenic strains.

Corynebacterium striatum has been increasingly associated with nosocomial outbreaks and antimicrobial resistance. Objectives: This study presents the comparative analysis of 26 multidrug-resistant (MDR) C. striatum strains isolated in Brazil. Methods: Additional genomes from international sources were incorporated. The analyses encompassed in vitro antimicrobial susceptibility testing and an in silico workflow for genomic similarity comparison, phylogenetic reconstruction, genomic clustering, pangenome analysis, mobilome content, virulence prediction, and functional annotation of unique proteins and putative virulence clusters. Results: Strong in silico evidence of clonality among several Brazilian isolates was obtained at the same time that some strains consistently indicated a divergent genomic profile. There are 196 unique coding sequences (CDSs) across the Brazilian IHPs. Of particular interest, strain IHP2030 carried an exclusive fimbria, sharing less than 50% similarity with other fimbriae in the dataset. Yet, structural predictions suggested conservation of key structural domains typically associated with fimbrial proteins. Mobilome content analysis revealed that IHPs strains were overall similar, differing primarily in the number of insertion sequences and in the presence or absence of CRISPR-Cas defense systems. Regarding virulence, an exclusive cluster in IHP2050 and IHP2060 suggests adaptive advantages associated with their respective environments of isolation. Conclusion: This study reveals a complex genomic landscape among Brazilian MDR C. striatum strains, marked by clonal dissemination alongside strain-level genetic variation in accessory genomes, mobilome composition, and virulence-associated gene repertoires, providing genomic evidence of diversification within hospital-associated lineages.

Corynebacterium ulcerans is an emerging zoonotic pathogen that can cause diphtheria-like infections in humans. In this study, we report the novel detection and comprehensive phenotypic and genomic characterization of three atoxigenic C. ulcerans strains isolated from domestic animals in Brazil. Notably, all isolates belonged to the multilocus sequence type ST-339, which has been previously identified in both human and animal hosts from geographically distant regions, suggesting the potential for international dissemination. Whole-genome sequencing confirmed species identity and revealed high genetic simila-rity among isolates, although distinct phylogenetic subclades were observed. Genomic analyses identified conserved virulence-associated determinants, including incomplete pilus gene clusters, iron acquisition systems, and the pld gene encoding phospholipase D. In contrast, the tox gene was absent in all strains. Notably, one isolate exhibited ciproflo-xacin resistance associated with double mutations (S89L and D93G) in the quinolone- resistance-determining region of GyrA. Molecular modeling and dynamics simulations demonstrated that these mutations impair key interactions within the ciprofloxacin– magnesium–water complex, thereby compromising the stability of drug binding. Additionally, the presence of diverse mobile genetic elements, prophages, and CRISPR-Cas systems highlighted the genomic plasticity of these isolates. Our findings provide new insights into the zoonotic potential, antimicrobial resistance mechanisms, and genomic diversity of C. ulcerans, underscoring the need for strengthened surveillance and mole-cular monitoring of this emerging pathogen in both veterinary and public health contexts.