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Background  Accurate assessment of probiotics with targeted anti- Salmonella  activity requires suitable models accounting for both, microbe-microbe and host-microbe interactions in gut environments. Here we report the combination of two original  in vitro  intestinal models closely mimicking the complex  in vivo  conditions of the large intestine. Effluents from continuous  in vitro  three-stage fermentation colonic models of  Salmonella  Typhimurium infection inoculated with immobilized child microbiota and  Salmonella  were directly applied to confluent mucus-secreting HT29-MTX cell layers. The effects of  Salmonella , addition of two bacteriocinogenic strains,  Bifidobacterium thermophilum  RBL67 (thermophilicin B67) and  Escherichia coli  L1000 (microcin B17), and inulin were tested on  Salmonella  growth and interactions with epithelial cell layers.  Salmonella  adhesion and invasion were investigated and epithelial integrity assessed by transepithelial electrical resistance (TER) measurements and confocal microscopy observation. Data from complex effluents were compared with pure  Salmonella  cultures.    Results   Salmonella  in effluents of all reactors of the colonic fermentation model stabilized at mean values of 5.3 ± 0.8 log 10  cfu/ml effluent. Invasion of cell-associated  Salmonella  was up to 50-fold lower in complex reactor samples compared to pure  Salmonella  cultures. It further depended on environmental factors, with 0.2 ± 0.1% being measured with proximal, 0.6 ± 0.2% with transverse and 1.3 ± 0.7% with distal reactor effluents, accompanied by a similar high decrease of TER across cell monolayers (minus 45%) and disruption of tight junctions. Subsequent addition of  E. coli  L1000 stimulated  Salmonella  growth (6.4 ± 0.6 log 10  cfu/ml effluent of all 3 reactors) and further decreased TER, but led to 10-fold decreased invasion efficiency when tested with distal reactor samples. In contrast, presence of  B. thermophilum  RBL67 revealed a protective effect on epithelial integrity compared to previous  E. coli  L1000 periods, as reflected by a significant mean increase of TER by 58% in all reactors. Inulin addition enhanced  Salmonella  growth and invasion when tested with distal and proximal reactor samples, respectively, but induced a limited decrease of TER (minus 18%) in all reactors.    Conclusions  Our results highlight the benefits of combining suitable cellular and colonic fermentation models to assess strain-specific first-level host protection properties of probiotics during  Salmonella  infection, providing an efficient system biology tool for preclinical development of new antimicrobials.

Protective effect of probiotics on Salmonella infectivity assessed with combined in vitro gut fermentation-cellular models