As normal inhabitants of diverse ecosystems, including the human gastrointestinal tract, the enterococci, and especially the two speciesEnterococcus faecalisandEnterococcus faecium, can be considered ubiquitous with regard to our natural environment.E. faeciumhas gained special importance thanks to beneficial strains marketed as probiotics, and because of its beneficial role in traditional fermented foods such as artisanal cheeses in some Southern European countries. Yet, following reports on the increasing association of some enterococcal strains with nosocomial infections such as endocarditis and bacteraemia, it became evident that strains from clinical origin are frequently highly resistant to ‘last-defence-line’ antibiotics such as the glycopeptide derivatives. For this reason enterococci have been classified in risk group 2 in the European Directive 93/88. With this paper it is intended to clarify the uncertain situation around the safety of the speciesE. faecium, also with referring to intra-species heterogeneity. In fact, well established scientific and surveillance data support the safety of some probioticE. faeciumstrains for both human and animal applications. As a model, summarising yet extensive information is provided on the efficacy and safety ofE. faeciumSF68®, a pharmaceutical probiotic with a long history of safe use. We propose the approach presented in this review as a model for the evaluation of safety of probiotic strains of this species.

Aims The objective of this study was to isolate multifunctional bacteriocin-producing strains; to characterize the expressed bacteriocin for the control of Listeria monocytogenes and vancomycin-resistant Enterococcus; to evaluate safety of studied strains; and to explore their antifungal activity. Methods and Results Two Pediococcus strains were isolated from silage samples obtained from an organic farm in Belogratchick, Bulgaria. The strains were identified by 16S rRNA sequencing analysis and characterized as bacteriocins producers. Strong antimicrobial activity was detected against more than 74 different strains of Listeria monocytogenes, 27 different vancomycin-resistant Enterococcus strains. In addition studied strains were able to inhibit the growth of strains of Alternaria alternate, Aspergillus flavus, Asp. niger, Cladosporium sphaerospermum, Penicillium chrysogenum and Pen. expansum. Some aspects of the antimicrobial mode of action were evaluated, including killing curves and aggregation properties. Both strains generated positive PCR results for the presence of pediocin PA-1, but not for other bacteriocins evaluated in this screening process. Metabolomic analysis of the cell-free supernatants from both strains was performed in order to explain the observed antifungal activity against different molds. According to PCA and PLS-DA score plot, P. acidilactici ST3522BG and P. pentosaceus ST3633BG were clearly clustered from control (MRS). Increases in the production of benzoic acid, 2-hydroxyisocaproic acid, β-phenyl-lactic acid, α-hydroxybutyric acid and 1,3-butanediol were recorded, these metabolites were previously described as antifungic. Conclusions P. acidilactici ST3522BG and P. pentosaceus ST3633BG were evaluated as producing bacteriocin strains with high specificity against Listeria and vancomycin-resistant Enterococcus species. In addition both investigated Pediococcus strains were evaluated as producer of effective antifungal metabolite with potential for inhibition of mycotoxin producing molds. Significance and Impact of the Study To the best of our knowledge, this report is a pioneer in evaluation of Pediococcus strains isolated from silage with highly specific bacteriocinogenic antimicrobial activity against Listeria spp. and vancomycin-resistant Enterococcus spp., and antifungal activity against mycotoxin producing molds.

Gut microbiota play a key role in the development of metabolic disorders. Defining and correlating structural shifts in gut microbial assemblages with conditions related to metabolic syndrome have, however, been proven difficult. Results from 16S genomic DNA and 16S ribosomal RNA analyses of fecal samples may differ widely, leading to controversial information on the whole microbial community and metabolically active microbiota. Using a C57BL/6J murine model, we compared data from 16S genomic DNA and ribosomal RNA of the fecal microbiota. The study included three groups of experimental animals comprising two groups with high fat diet induced obesity (DIO) while a third group (control) received a low fat diet. One of the DIO groups was treated with the probiotic Lactobacillus rhamnosus GG (LGG). Compared to the data obtained by DNA analysis, a significantly higher abundance of OTUs was accounted for by RNA analysis. Moreover, rRNA based analysis showed a modulation of the active gut microbial population in the DIO group receiving LGG, thus reflecting a change in the induced obesity status of the host. As one of the most widely studied probiotics the functionality of LGG has been linked to the alleviation of metabolic syndrome, and, in some cases, to an impact on the microbiome. Yet, it appears that no study has reported thus far on modulation of the active microbiota by LGG treatment. It is postulated that the resulting impact on calorie consumption affects weight gain concomitantly with modulation of the functional structure of the gut microbial population. Using the 16S rRNA based approach therefore decisively increased the precision of gut microbiota metagenome analysis.