Abstract
Strain CCM 2573 is a Gram-positive bacterium that has been intensively studied in the past due to its distinct chemotaxonomic properties, but its reliable taxonomic classification has not been satisfactorily clarified. Whole-genome sequencing and comparative genomic analyses performed in this study revealed that the strain belongs to the Macrococcus caseolyticus phylogenetic clade. Genome-to-genome comparisons confirmed the closest relationship to the type strains of M. caseolyticus subsp. hominis CCM 7927 T and M. caseolyticus subsp. caseolyticus DSM 20597 T . However, the strain harbored unique genomic elements distinguishing it from its nearest phylogenetic neighbors. Its accessory genome contains dozens of insertion sequences, a 92-kbp composite transposon with unique palindromic repeat loci associated with a CRISPR-Cas adaptive immune system, a pseudo-staphylococcal chromosome cassette, and several additional genomic islets. Unlike other macrococci, strain CCM 2573 exhibits a specific peptidoglycan composition (L-Lys-Gly₂-Ser₂-Gly) and shows a higher phylogenetic divergence of aminoacyltransferases (FemABX) involved in interpeptide bridge synthesis. In addition, it reveals distinct biochemical characteristics from both subspecies of M. caseolyticus , particularly in its ability to produce acid from galactose, cellobiose, melezitose, and turanose, as well as in its susceptibility to novobiocin. The MALDI-TOF mass spectra enable differentiation of the strain from other type strains of the genus Macrococcus . The results of polyphasic taxonomy obtained in this study showed that strain CCM 2573 belongs to the species M. caseolyticus , but it is distinct from both validly named M. caseolyticus subspecies. We propose to assign the analyzed strain as a new subspecies, Macrococcus caseolyticus subsp. lactis subsp. nov. The type strain is CCM 2573 T (= DSM 20227 T ).