Rearing rats in a germ-free environment eliminates their odors of individuality

In order to test the hypothesis that commensal bacteria influence the urinary odors of individuality, we collected urine from PVG and PVG.R1 male rats born by cesarian section and reared in a germ-free environment. Using the habituation-dishabituation test with PVG.RT1 ^u and Lister hooded rats as subjects, we found that urine from the germ-free rats was not discriminated, while urine from conventionally housed rats of the same strains could be discriminated (experiment 1). When the germ-free rats were moved to a conventional animal house after recolonization with commensural flora and their urine collected, it was discriminated, indicating an essential role of bacteria in determining the unique urinary odors of MHC-congenic rats (experiment 2). The conventionally housed and germ-free rats did not differ in the amount of class I antigen in their urine (experiment 3). Finally, urines of PVG and PVG.R1 donors inoculated with a defined and highly restricted flora to render them specific-pathogen-free (SPF) could not be discriminated. Urine from SPF donors moved to a conventional animal house could be discriminated (experiment 4). These results indicate that commensal bacteria are essential for the production of the unique individual odor of the urine of MHC-congenic strains of rats.