In vivo assessment of potential probiotic Lactobacillus salivarius strains: evaluation of their establishment, persistence, and localisation in the murine gastrointestinal tract

The enteric flora comprise approximately 95% of the total number of cells in the human body. Numerous studies have investigated potentially beneficial members of this microbial community due to their ability to elicit immune responses while also protecting against microbial pathogens. We have previously reported on the isolation and identification, from surgically-resected segments of the human gastrointestinal tract (GIT), of potential probiotic lactic acid bacteria (LAB). These bacterial strains exhibit potentially beneficial probiotic traits in vitro such as bile tolerance in the absence of deconjugation; gastric acid resistance; and adherence to epithelial cell lines. The objective of this study was to administer two strains of the previously-isolated LAB to mice over a period of 7 or 14 days in order to assess their ability to establish themselves within specific regions of the GIT. Throughout this feeding period, and for 4 days following cessation of feeding, the numbers of total culturable lactobacilli and of the administered LAB present in faeces were monitored. Spontaneous rifampicin resistant derivatives (50 mg:ml) of Lactobacillus salivarius subsp. salivarius UCC1(LM5) and Lb. salivarius subsp. salivarius UCC118(LM2) were generated to facilitate enumeration of the strains in GIT and faecal samples. Each potential probiotic strain was individually administered to Balb:c mice at a daily concentration of approximately 4.0 109 CFU. After 1 day of feeding, strains UCC1(LM5) and UCC118(LM2) were recovered from murine faeces at Log10 6.95 (1.18) CFU:g and Log10 6.33 (0.37) CFU:g, respectively. Interestingly, UCC118(LM2), which was originally isolated from the ileal-caecal region of the human GIT, was found to have become established in the corresponding region of the murine GIT regardless of the length of the feeding period. UCC118(LM2) was also found to persist in faeces for a period of up to 3 days following cessation of feeding. Administration of UCC1(LM5) and UCC118(LM2) did not result in any significant changes in the levels of indigenous bacteria culturable from faeces. In conclusion, human isolate Lb. salivarius subsp. sali6arius UCC118(LM2) was found to effectively colonise, and survive transit through, the murine gastrointestinal tract. Lactobacillus salivarius subsp. sali6arius UCC118 has been deposited at The National Collections of Industrial and Marine Bacteria (NCIMB) and accorded the accession number NCIMB40829

A randomised controlled trial of a probiotic Lactobacillus strain in healthy adults: assessment of its delivery, transit and in fluence on microbial flora and enteric immunity

In severa intestinal disease states, altered microflora, impaired gut barrier and:or intestinal in ammation offer a rationale for the effective therapeutic use of probiotic microorganisms. However , for most candidate probiotic organisms there is a lack of evidence detailing their characterisation and effects on host flora and immunity. We have previously reported the isolation and characterisation, from surgically resected segments of the human gastrointestinal tract (GIT), of potential probiotic lactic acid bacteria (LAB). We have also described subsequent animal experiments that evaluated the establishment , persistence and localisation of speci fic probiotic Lactobacillus strains within the murine intestinal tract, in addition to their ability to influence the development of murine in ammatory disorders. In these studies, transit and survival of Lactobacillus salivarius UCC118 at the ileum was demonstrated using enteral tube sampling of six healthy volunteer s following consumption of a single dose (150 ml) of fermented milk-borne probiotic (108 colony forming units per ml (CFU:ml)). Subsequently, we performed a randomised controlled trial of 80 volunteer s fed strain UCC118 (108 CFU:day for 21 d), using two oral delivery vehicles (fresh milk, n¾20 vs. fermented milk, n¾20; controls, n¾20 for each). Throughout this feeding period, and for up to 100 days following cessation of feeding, the number s of total culturable lactobacilli and of the administered Lactobacillus UCC118 present in faeces were monitored. Five subjects (5:40; fresh milk, four; fermented milk, one) were still excreting the probiotic lactobacilli 21 days post-cessation of feeding, while one subject (fermented milk) was still colonised up to 100 days after feeding. Consumption of fermented milk-borne UCC118 cells resulted in signi cantly increased levels of faecal-borne enterococci and lactobacilli. Numbers of bifidobacteria, coliforms and bacteroides were not significantly altered. In addition, changes in salivary IgA levels against UCC118 cells and increased granulocyte phagocytic activity were observed following consumption of the fermented milk-borne probiotic. In summary, Lactobacillus UCC118 was found to effectively transit (and persist within) the human intestinal tract, to modify the faecal flora and to engage the immune system

Mechanisms of adherence of a probiotic lactobacillus strain during and after in vivo assessment in ulcerative colitis patients

In a pilot-scale, open-label study to determine the ability of well-characterized probiotic Lactobacillus salivarius UCC118 cells to adhere to human epithelial cells in situ , the bacterial strain was administered to ulcerative colitis patients at approximately 109 CFU/day for 12 days. Microbiological analysis of biopsy specimens demonstrated that the ingested bacteria effectively adhered to both inflamed and non-inflamed mucosa of the large bowel in significant numbers. In previous reports, we have described the ability of the lactobacilli to adhere to enterocytic epithelial cells in vitro. In this study, we found that the bacteria adhered at higher levels to differentiated rather than undifferentiated epithelial monolayers; and that stationary phase lactobacilli were found to adhere to eukaryotic HT-29 and Caco-2 epithelial cells at greater levels than log phase bacterial cells. Pretreatment of the Lactobacillus cells with proteolytic enzymes abolished attachment, indicating the potential involvement of surface/exposed protein(s) as bacterial adhesin(s). SDS-PAGE (denaturing) techniques determined that the proteolytic treatment resulted in degradation of a cell wall-associated protein of approximately 84 kDa. The proteinaceous factor was purified by both anion-exchange chromatography and by gel extraction after SDS-PAGE electrophoresis, and under in vitro assay conditions proved capable of adherence and significant inhibition of bacterial attachment to enterocytic epithelial cells