Enterococcus spp. in Brazilian artisanal cheeses: Occurrence and assessment of phenotypic and safety properties of a large set of strains through the use of high throughput tools combined with multivariate statistics

Traditionally fermented food generally has unique sensorial aspects due to its high microbial diversity, being broadly consumed worldwide. Despite this, some Vancomycin-Resistant-Enterococci (VRE) can occur in these products, which raises questions about their safety. Thus, this study tries to assess information about the occurrence of Enterococcus strains in traditional Brazilian cheese samples collected from the five main regions as well as the characterization of their phenotypic (proteolytic, lipolytic and bacteriogenic potential) and safety (hemolysin activity and growth at commercial VRE selecting media) aspects. A total of 1669 lactic acid bacteria were isolated, of which 605 (36.2%) were identified as Enterococcus by 16sRNA sequencing. Out of these, E. faecium (58.7%) was identified as the dominant species, followed by Enterococcus faecalis (31.8%), Enterococcus durans (4.8%), Enterococcus hermanniensis (1.6%) and Enterococcus gilvus (1.6%). All the data were analyzed by multivariate statistics, and the findings highlighted the presence of Enterococcus as a naturally component of indigenous microbiota of Brazilian artisanal cheeses, mostly in the cheeses of Marajo, Coalho, and Manteiga, possibly as a consequence of their heat resistance and the curd cooking step applied during the process in these regions. The majority of the identified Enterococcus (63.1%) did not produce beta-hemolysin and were not resistant to vancomycin. Interestingly some of these strains (21.4%) were able to inhibit Listeria monocytogenes ATCC 35152, mostly isolates from Serrano, Colonial, and Minas artisanal (from Cerrado microregion) cheeses. Also, 10.2 and 11.8% of them have shown moderate to high proteolytic and lipolytic activity, respectively. Antilisterial activity is of most interest as it can contribute to reduce risk of listeriosis due to dairy products consumption, while proteolysis and lipolysis are responsible for some remarkable features (e. g. flavor, taste, and texture) of fermented foods. In summary, this study represents a contribution towards revealing some phenotypic and safety traits of a large set of strains of Enterococcus strains through the use of multivariate statistics combined with high throughput tools.