Non-aureus Staphylococci and Mammaliicocci (NASM) in bovine milk: antimicrobial resistance, virulence, and associations with management practices in Brazil

Non-aureus staphylococci and mammaliicocci (NASM) are frequently isolated from bovine milk in cases of mastitis and intramammary infection, as well as from extramammary environments. Despite advances in species-level identification of NASM, it is not consistently performed across studies, which may limit a comprehensive understanding of their epidemiological relevance. This study aimed to provide a comprehensive characterization of NASM isolated from bovine milk across Brazil's 5 major dairy-producing regions. Using MALDI-ToF MS, 309 NASM isolates obtained from cows with low and high somatic cell counts (SCC) and from clinical mastitis cases were identified at the species level. All isolates were evaluated for antimicrobial susceptibility and screened for the presence of methicillin resistance genes (mecA, mecC), disinfectant resistance genes (qacAB, qacC), and enterotoxin genes (sea, seb, sec, sed, see, seg, seh, sei). In addition, the association between NASM occurrence and herd management practices was assessed. Eighteen NASM species were identified, with Staphylococcus chromogenes (58.6%), S. simulans (8.4%), and S. haemolyticus (8.1%) being the most frequently isolated species. Forty-one isolates (13.3%) were classified as multidrug-resistant, and 15 carried the mecA gene. The most frequently detected enterotoxin gene was seb, present in 15.2% of isolates. Management-related analyses indicated that S. chromogenes occurred widely across farms with adequate milking hygiene, whereas non-chromogenes species were more frequently associated with herds where hygienic practices were less consistent. This study provides the first large-scale, species-level evaluation of NASM across multiple Brazilian regions, integrating phenotypic, genotypic, and management-related data. The findings highlight the importance of NASM as common intramammary pathogens and their potential roles in the persistence of mastitis, the dissemination of antimicrobial resistance, and challenges in dairy herd management.