Influence of a Human Milk Diet on Colonisation Resistance Mechanisms AgainstSalmonella typhimuriumin Human Faecal Bacteria-Associated Mice

Abstract

Previous studies in our laboratory showed that gnotobiotic mice associated with human infant intestinal microorganisms were more resistant to colonisation with Salmonella typhimurium when consuming human milk than when consuming bovine milk, cows' milk based formula (CMBF) or formula modifications. To determine mechanisms of protection, we examined the effects of human milk and CMBF consumption on several putative protective host factors including gastric acidity, caecal pH levels and caecal short chain fatty acid (SCFA) concentrations. When consuming human milk, the pH of the stomach contents of the mice was significantly lower (2.18) than when consuming CMBF (3.24). S. typhimurium survived for less than 30 min in tryptic soy broth adjusted to pH 2.18 but survived for more than 2.5 h in broth adjusted to pH 3.24. Caecal SCFA concentrations were greater in mice consuming CMBF (1968 μg/g) than in mice consuming human milk (1404 μg/g), however, the pH of caecal contents of mice consuming human milk (5.69) was significantly lower than that of mice consuming CMBF (6.44). Caecal contents, obtained from germfree mice, were adjusted to simulate the SCFA and pH levels measured in the caeca of test mice, and were then inoculated with S. typhimurium . While SCFA and pH at levels present in caecal contents of human milk-consuming mice were bactericidal for S. typhimurium , multiplication of the organism occurred at levels present in contents of CMBF-consuming mice. Lowering the pH level of caecal contents obtained from germfree mice from 6.69 to 5.69, without concomitant SCFA addition, did not retard the multiplication of S. typhimurium , indicating that hydrogen-ion activity alone was not responsible for inhibition. Raising the pH level of caecal contents obtained from human milk-consuming mice containing SCFA to 6.44, the level determined in the contents of CMBF-consuming mice, or to 7.50 to neutralise the acids, reversed inhibition. To confirm the importance of low caecal pH in S. typhimurium inhibition, groups of mice were fed either human milk, CMBF, or buffered human milk before challenge with the pathogen. Consumption of buffered human milk, as compared with the unbuffered control, resulted in an increase in the pH of caecal contents and in caecal S. typhimurium concentrations to levels approaching those observed in mice consuming CMBF. The results suggest that the low gastric pH and non-ionised SCFA, present in the acidic environment of the caecum during human milk consumption, impede colonisation of the gastrointestinal tract of the human faecal microbiota associated mice with S. typhimurium . Keywords: human milk diet, Salmonella typhimurium , resistance, faecal bacteria