Background and aims-The gastrointestinal microflora exerts a barrier eVect against enteropathogens. The aim of this study was to examine if bifidobacteria, a major species of the human colonic microflora, participates in the barrier eVect by developing antimicrobial activity against enterovirulent bacteria. Methods-Antibacterial activity was examined in vitro against a wide range of Gram negative and Gram positive pathogens. Inhibition of Salmonella typhimurium SL1334 cell association and cell invasion was investigated in vitro using Caco-2 cells. Colonisation of the gastrointestinal tract in vivo by bifidobacteria was examined in axenic C3/He/Oujco mice. Antimicrobial activity was examined in vivo in axenic C3/He/Oujco mice infected by the lethal S typhimurium C5 strain. Results-Fourteen human bifidobacterium strains isolated from infant stools were examined for antimicrobial activity. Two strains (CA1 and F9) expressed antagonistic activity against pathogens in vitro, inhibited cell entry, and killed intracellular S typhimurium SL1344 in Caco-2 cells. An antibacterial component(s) produced by CA1 and F9 was found to be a lipophilic molecule(s) with a molecular weight of less than 3500. In the axenic C3/He/Oujco mice, CA1 and F9 strains colonised the intestinal tract and protected mice against S typhimurium C5 lethal infection. Conclusion-Several bifidobacterium strains from resident infant human gastrointestinal microflora exert antimicrobial activity, suggesting that they could participate in the "barrier eVect" produced by the indigenous microflora. (Gut 2000;47:646-652)

The spent culture supernatant of the human Lactobacillus acidophilus strain LB produces an antibacterial activity against a wide range of gram-negative and gram-positive pathogens. It decreased the in vitro viability of Staphylococcus aureus, Listeria monocytogenes, Salmonella typhimurium, Shigella flexneri, Escherichia coli, Klebsiella pneumoniae, Bacillus cereus, Pseudomonas aeruginosa, and Enterobacter spp. In contrast, it did not inhibit lactobacilli and bifidobacteria. The activity was heat stable and relatively sensitive to enzymatic treatments and developed under acidic conditions. The antimicrobial activity was independent of lactic acid production. Activity against S. typhimurium SL1344 infecting human cultured intestinal Caco-2 cells was observed as it was in the conventional C3H/He/oujco mouse model with S. typhimurium C5 infection and oral treatment with the LB spent culture supernatant.

The aim of this study was to compare the antagonistic properties of Lactobacillus casei GG exerted in vitro against Salmonella typhimurium C5 in a cellular model, cultured enterocyte-like Caco-2 cells, to those exerted in vivo in an animal model, C3H/He/Oujco mice. Our results show that a 1-h contact between the invading strain C5 and either the culture or the supernatant of L. casei GG impeded the invasion by the Salmonella strain in Caco-2 cells, without modifying the viability of the strain. After neutralization at pH 7, no inhibition of the invasion by C5 was observed. The antagonistic activity of L. casei GG was examined in C3H/He/Oujco mice orally infected with C5 as follows: (i) L. casei GG was given daily to conventional animals as a probiotic, and (ii) it was given once to germ-free animals in order to study the effect of the population of L. casei GG established in the different segments of the gut. In vivo experiments show that after a single challenge with C5, this strain survives and persists at a higher level in the feces of the untreated conventional mice than in those of the treated group. In L. casei GG germ-free mice, establishment of L. casei GG in the gut significantly delayed the occurrence of 100% mortality of the animals (15 days after C5 challenge versus 9 days in germ-free mice [P < 0.01]). Cecal colonization level and translocation rate of C5 to the mesenteric lymph nodes, spleen, and liver were significantly reduced during the first 2 days post-C5 challenge, although the L. casei GG population level in the gut dramatically decreased in these animals. Recently, Klaenhammer (21) proposed that Lactobacillus strains with well-defined properties should be selected and characterized for specific use. Among Lactobacillus strains used in fermented milks, Lactobacillus casei GG has been shown to promote clinical recovery from acute rotavirus diarrhea in infants (17-20, 24) and from either antibiotic-associated or traveller's diarrhea in adults (27, 36). It has been previously reported that L. casei GG adheres to enterocyte-like cells in culture (11). It exerts an antagonistic effect against several bacteria and produces an unknown antimicrobial substance against Escherichia coli (37). But the mechanism of action of L. casei GG in vivo remains unknown. We have recently reported that selected, adhering human L. acidophilus strains inhibit both cell association and cell entry of a variety of enterovirulent bacteria within human cultured enterocyte-like Caco-2 cells (5-8). In the present work, we have compared the antagonistic effect observed in vitro in a cellular model (Caco-2 cells) to that observed in an in vivo animal model (C3H/He/ Oujco mice). First, we studied whether L. casei GG can inhibit the invasion of cultured enterocyte-like Caco-2 cells by Salmonella typhimurium C5 (28). Second, the antagonistic activity of L. casei GG was examined in C3H/He/Oujco mice orally infected with S. typhimurium C5. Two experimental designs were used: (i) L. casei GG given daily to conventional animals as a ...