Coagulase-negative staphylococci (CNS) represent common skin commensals in humans and other mammals. They are opportunistic microorganisms and, in dairy production, are considered as minor pathogens, often associated to subclinical mastitis. It has been recently demonstrated that CNS can produce Staphylococcus aureus superantigens, as well as biofilm. In our study, 74 CNS were isolated from Tuscan (Central Italy) bovine bulk tank milk and phenotypically identified as Staphylococcus xylosus (28.4 %), Staphylococcus chromogenes (20.3 %), Staphylococcus sciuri (12.2 %), Staphylococcus hominis (9.4 %), Staphylococcus haemolyticus (6.8 %), Staphylococcus capitis (5.4 %), Staphylococcus cohnii ssp. cohnii (4.1 %), Staphylococcus saprophyticus (4.1 %), and Staphylococcus simulans (4.1 %). Moreover, one isolate (1.3 %) was detected for each of the following species: Staphylococcus caprae, Staphylococcus epidermidis, Staphylococcus lentus, and Staphylococcus warneri. All isolates were screened by PCR for the presence of genes encoding classical enterotoxins (sea-see), toxic shock syndrome toxin-1 (tst), and biofilm (icaA/D). Forty isolates (54.1 %) presented at least one gene encoding for enterotoxin production, 15 (20.3 %) harbored two or more genes in association. The most frequently detected gene was sea (41.9 %), followed by sec-1 (25.7 %). None of the tested isolates presented genes encoding for enterotoxin B or D. Fifteen isolates (20.3 %) presented genes encoding for biofilm production. Among these, 11 isolates also harbored genes encoding for enterotoxin production. Our results suggest that enterotoxigenic CNS could represent a potential reservoir of pathogenicity factors, which could substantially contribute to enhance S. aureus virulence when present in milk and milk products. Further investigations are required for the evaluation of enterotoxins and biofilm production.