Objectives: Bacterial colonization of the infant gut may have important influences on the development of gastrointestinal, respiratory, and allergic disease. Early diet is a major determinant of the gut microflora. It is very difficult to carry out studies in human infants that can investigate the interaction of diet, flora, and mucosa. In this study we have developed an infant human flora-associated (IHFA) rat model to allow such investigation. Methods: Germ-free infant rats were infected with fecal bacteria from exclusively breast-fed infants and were maintained on a modified infant formula for 8 weeks. The fecal and cecal contents were collected and compared with feces of breast-fed infants for bacterial populations, bacterial metabolites, and enzymes and for the ability to inhibit adhesion of pathogenic bacteria to human mucosal cells. Results: The IHFA cecum and feces were dominated by lactic acid bacteria, Bifidobacterium, and lactobacilli, which were representative of the infant feces. The fecal short-chain fatty acid profile was dominated by acetic and lactic acid in a similar manner to human infant feces. Other bacterial metabolites were similar to those of the human infant. Rat intestinal samples were able to inhibit the adhesion of pathogens to mucosal cells, but to a lesser extent than the human samples. Conclusions: This IHFA infant model of the intestinal flora of the breast-fed infant is considered valid for studying the effect of diet on bacterial colonization and metabolism. Chemicals/CAS: acetic acid, 127-08-2, 127-09-3, 64-19-7, 71-50-1; ammonia, 14798-03-9, 51847-23-5, 7664-41-7; beta glucosidase, 51683-43-3, 9001-22-3; beta glucuronidase, 9001-45-0; cholesterol, 57-88-5; lactic acid, 113-21-3, 50-21-5; para cresol, 106-44-5; phenol, 108-95-2, 3229-70-7; tryptase, 97501-93-4