228 may be that compared with a single strain probiotic a wider range of health beneﬁts could be accomplished. In contrast, mixing of probiotic strains might also result in reduced efﬁcacy since individual strains may have opposite effects or even inhibit each other. There are, however, a limited number of in vivo studies available that compare the effects of a probiotic mixture with those of the individual strains (Chapman et al. 2011). This means that the evidence for the hypothesis that probiotic mixtures are more effective than the individual strains is still limited. The inﬂuence of probiotics on the composition and diversity of the intestinal microbiota Modulation of the intestinal microbiota (part of level 1 probiotic action) is one of the potential health-beneﬁcial effects of probiotics. The mechanisms by which probiotic microorganisms are able to modify the intestinal micro- biota include reduction of luminal pH, competition for nutrients, secretion of anti-microbial compounds (bacteri- ocins), prevention of bacterial adhesion and evasion of epithelial cells, and induction of the production of anti- microbial compounds (defensins) by the host (Fooks and Gibson 2002; Ng et al. 2009). By these mechanisms, pro- biotics can not only potentially modulate the intestinal microbiota composition, but also prevent pathogenic bacterial overgrowth. Up to this date, many studies have been performed that examine the effects of probiotics on the composition and diversity of the intestinal microbiota, both in diseased and healthy individuals. For a given disease, the desired out- come of probiotic intervention is the modulation of the intestinal microbiota in such a way that a healthy micro- biota composition is achieved. However, also other parameters have been addressed such as stabilisation of the microbiota as in the case of IBS and a multispecies pro- biotic, determined with the use of a phylogenetic micro- array (Kajander et al. 2008). The interpretation of the effects of probiotics on the intestinal microbiota composi- tion in healthy individuals are, however, more difﬁcult to interpret (Table 7). Those studies do provide information on the effects of probiotics on the intestinal microbiota without a potential bias caused by disease effects. How- ever, this does not imply that in a diseased situation these probiotic products will have the same inﬂuence on the intestinal microbiota. Until recently, in most of the probiotic studies conven- tional culture-based methods have been used to study the inﬂuence of probiotics on the intestinal microbiota. How- ever, since a few years culture-independent methods are now also being applied in probiotic research (Table 7). In general, demonstrating the colonisation of the supple- mented probiotic microorganism(s) has been the primary 123 Genes Nutr (2011) 6:209–240 aim of most studies in healthy individuals. In most cases, a transient colonisation of the probiotic microorgan- ism(s) has been observed. It is still questionable, however, whether probiotic strains would need to colonise in order to be effective or whether transient presence would also sufﬁce to exert health-beneﬁcial effects. The probiotic studies performed in humans have almost exclusively examined the effect of probiotic administration on the composition of the faecal microbiota, whereas other niches of the GI tract have hardly been studied thus far (Table 7). As already indicated, even major local changes in microbiota composition in speciﬁc niches of the GI tract might not be reﬂected in the faeces. This means that there is still a major gap in knowledge on the inﬂuence of pro- biotic microorganisms on the intestinal microbiota. In addition, the inﬂuence of probiotic microorganisms on mucosa-associated intestinal microbiota is also not well studied. However, these interactions are possibly of key importance in relation to disease pathogenesis, since mucosa-associated microorganisms are in more close contact with the intestinal barrier and immune system. One of the few examples of a study on the in vivo effects of probiotics on the human host is a recent study by Van Baarlen et al. (2009). The authors examined the inﬂuence of a probiotic microorganism on human duodenal mucosal gene expression and they showed that changes in gene expression patterns, especially in the NF-jb dependent pathways, induced by Lactobacillus plantarum WCFS1 could be linked to the establishment of immunotolerance in human adults. In contrast with most human probiotic studies, animal studies have focused on the spatial inﬂuence of probiotics on the intestinal microbiota (Table 7). However, to which extent these results reﬂect the human situation has to be determined. Administration of a given probiotic strain will result in the (temporarily) increase of that strain the GI tract, but may also change the overall composition of the intestinal microbiota. Indeed, the results of relevant experiments performed thus far demonstrate that probiotic- induced changes in microbiota composition are not restricted to the administered species. Which probiotic microorganisms are able to inﬂuence the relative abun- dance of which speciﬁc intestinal microorganisms are questions that are currently under study. It should be realised, however, that a change in composition or diversity of the intestinal microbiota by probiotic intervention is not a health beneﬁt by itself. As discussed previously, dysbiosis of the intestinal microbiota has been associated with a growing number of (intestinal) diseases. Since modulation of the composition of intestinal microbiota by probiotics was demonstrated to be possible, probiotic intervention has the potential to counterbalance intestinal dysbiosis and thus restore health.