The probiotic market is undoubtedly one of the fastest growing markets of new food development. According to the Food and Agricultural Organization (FAO) and the World Health Organization (WHO), probiotics are defined as 'live microorganisms which, when administered in adequate amounts, confer a health benefit to the host'. Although infections are rare, reports on isolation of bacteria used in probiotics from infections have raised some concern. Demonstration of the safety of probiotics used for human consumption is therefore of utmost importance. This PhD thesis aims to assess the safety of lactobacilli and enterococci, to investigate the immunomodulatory properties of lactobacilli, and to evaluate the effect of a probiotic Enterococcus on the gastro-intestinal flora. The first part addresses the safety of enterococci and lactobacilli. Different techniques were set up and evaluated to assess the safety of isolates belonging to these two genera. Firstly, enterococcal virulence, antibiotic resistance and genotypic delineation of E. faecium are studied. Secondly, infectivity of lactobacilli in a rat endocarditis model is investigated, as well as putative risk factors and their adhesion to extracellular matrix proteins and cell lines. In the second part, immunomodulatory properties of lactobacilli are explored. In the third part temporal changes of the human enterococcal flora as well as the effect of a probiotic E. faecium on this flora are investigated. The fourth part provides recommendations on the biosafety assessment of probiotics used for human consumption. Finally, we summarize and discuss the obtained results, and conclude with future perspectives.