Staphylococcus microti Strains Isolated from an Italian Mediterranean Buffalo Herd

SIMPLE SUMMARY: Non-aureus staphylococci (NAS) are frequently found in bovine milk samples as well as on teat skin associated with subclinical mastitis (SCM). Staphylococcus microti is a new species among NAS resulted to recently be the predominant species in the milk of water buffaloes (Bubalus bubalis) affected by intramammary infection (IMI). The aim of this study was to assess the occurrence of S. microti in milk and milking parlor equipment samples of an Italian buffalo farm. No clinical mastitis was noticed, but S. microti-associated SCM and IMI were detected. The presence of S. microti was also revealed on milking parlor surfaces. The results of the antimicrobial susceptibility tests highlighted a complete resistance to tetracyclines, and 20.4% of the isolated S. microti exhibited a multidrug-resistant (MDR) profile. In conclusion, the presence of this bacterium in the milk of buffaloes in the absence of evident mastitis clinical signs underlines the need for further studies, and its finding on milking parlor surfaces suggests that the environmental quality of the milking parlor plays an important role in influencing both S. microti’s spread and the microbial communities of the milk. ABSTRACT: S. microti is a new species among non-aureus staphylococci (NAS) frequently found in bovine milk samples and associated with subclinical mastitis (SCM). The aim of this study was to analyze the presence of S. microti in 200 composite milk samples and 104 milking parlor surface swabs collected at a buffalo farm in Southern Italy to define its presence in milk and a milking parlor environment. The samples were inoculated onto different agar plates, and the isolates were identified by MALDI-TOF MS. The strains identified as S. microti (54/304 samples, 17.8%) were collected, and their purified genomic DNA was subjected to PCR amplification and whole 16S rRNA gene sequencing. Furthermore, their phenotypic resistance profiles were evaluated by a disk diffusion method, and the genotypic characterization of the tetracycline resistance was performed for the tetM and tetK genes by multiplex PCR. Four and forty-seven S. microti isolates from milk samples of lactating animals with subclinical mastitis (SCM) and intramammary infection (IMI), respectively, and three isolates from milking parlor surfaces were recovered. The genomic DNA was purified from the bacterial isolates, and the amplification and sequencing of the 16S gene further supported the proteomic identification as S. microti. No clinical mastitis was detected in the herd during the study period. The antimicrobial susceptibility testing revealed a worrisome 100% resistance to tetracyclines, genotypically mediated by the tetM gene for all strains. This study highlights that S. microti may be commonly isolated from dairy buffalo milk and milking parlor equipment. Its association with SCM or IMI remains to be established.