Establishment of specific pathogen-free (SPF) rat colonies using gnotobiotic techniques

Gnotobiotic Wistar rats were produced using gnotobiotic techniques, which were established in the production of a SPF mouse colony, in order to establish a barrier-sustained colony. One strain of Escherichia coli, 28 strains of Bacteriodaceae (B-strains), three strains of Lactobacillus (L-strains) and a chloroform-treated fecal suspension (CHF, Clostridium mixture) were prepared from conventional Wistar rats as the microflora source. Two groups of limited-flora rats, E. coli plus B-strains and E. coli plus CHF, were produced. After confirmation that Clostridium difficile was not detected in the CHF-inoculated rats, two groups of limited-flora rats were transferred to an isolator and housed together in a cage. These rats were then orally inoculated with L-strains. The gnotobiotic rats showed colonization resistance to Pseudomonas aeruginosa, and the number of E. coli in the feces was 10(5) to 10(6)/g. The gnotobiotic rats were transferred to a barrier room as a source of intestinal flora for SPF colonies. In the SPF rats, basic cecal flora was mainly composed of Bacteroidaceae, clostridia, fusiform-shaped bacteria and lactobacilli, and did not change over a long period. Their flora became similar to that of conventional rats.