Biodiversity, occurrence and antibiotic resistance profile of enterococci isolated in bovine raw milk and feces.

Enterococci are ubiquitous lactic acid bacteria that are widely distributed in raw-milk. They are commensals of the gastrointestinal tract of animals, thus, via fecal contamination, could reach raw milk and dairy products. The aims of this study were: 1) to determine the genotypic diversity of enterococci isolated from individual cow milk and bovine fecal samples; 2) to investigate if cow feces are the source of milk enterococcal contamination and 3) to evaluate the antibiotic resistance (AR) pattern of dairy-related enterococci and their ability to transfer resistance genes. Thirty-one multiparous Italian Holstein dairy cows was considered, and 310 presumptive Enterococcus were isolated at three different time points from milk (n: 177) and feces (n. 133) samples Differences in enterococcal community structure were observed between the milk and feces samples. E. faecalis (59.9%), E. faecium (18.6%) and E. lactis (12.4%) were the most frequently isolated species from milk, while E. faecium (84.2%) and E. hirae (15.0%) were the predominant ones in bovine feces. RAPD-PCR highlighted a high number of Enterococcus biotypes (45 from milk and 37 from cow feces) and none of the strains present in milk exhibited genetic profiles similar to those of strains from cow feces. These results suggest that the bovine feces are not a source of enterococci for milk. All the strains were susceptible to ampicillin, daptomycin, gentamicin, teicoplanin and vancomycin. Milk biotypes showed high percentages of AR to streptomycin (86.7%), tetracycline (73.3%), erythtomycin (46.6%) and tigecycline (33.3%), while linezolid (45.9%), quinupristin/dalfopristin (36.1%) and tigecycline (32.4%) were the most prevalent AR phenotypes in cow feces. In both milk and feces biotypes the tetracycline resistance was conferred by the tetM and tetL genes, and the Tn916/Tn1545 family transposons were detected in E. faecalis, E. malodoratus and E. hirae biotypes. Only the E. faecalis biotypes were able to transfer the tetM gene to the recipient strain, and the frequency from donors to Lb. delbrueckii ranged from 103 to 106 transconjugants per recipient. Our results indicated that Enterococcus biotypes from milk and bovine feces belong to different community and the ability of these microorganisms to transfer AR genes is strain-dependent.