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Germfree (GF) mice were inoculated with a cultured flora from 10-1, 10-3, 10-5, and 10-7dilutions of caecal contents from a 'normal' mouse. GF mice associated with a flora of a 'normal' mouse served as controls. The following intestinal parameters were determined: Colonization resistance (CR), Relative caecal weight (RCW), villus:crypt ratio (jejunum and ileum), IgA-producing cells (jejunum and ileum), ß-aspartyl glycine (faeces), volatile and non-volatile fatty acids (caecum) and bile acids (faeces). Only the 10-1culture was able to induce similar changes in the GF mice to a 'normal' flora. The GF + 10-5and GF + 10-7groups deviated markedly from the controls while the GF + 10-3group showed in general intermediate values between GF + SPF and GF + 10-1on the one hand and GF + 10-5and GF + 10-7on the other hand. ß-aspartyl glycine was present only in the GF + 10-7group.Scanning electron microscopy (SEM) of ileal contents revealed segmented filamentous organisms in the ileum of controls and the GF + 10-1group. The faecal flora consisted mainly of fusiform organisms. In the faeces of the 10-5and 10-7groups increasing amounts of non-bacterial matter were found, while in the faeces of the other groups virtually only bacteria were seen.

Attempts are described to 'normalize' germfree mice by association with 3, 21 and 71 different intestinal bacterial cultures isolated from mice with an SPF flora. Germfree mice associated naturally with an SPF flora served as controls. Vital bacterial counts were determined by aerobic and anaerobic culture. Stomach and small intestine contained fewer bacteria per gram than caecum and large intestine. Aerobic vital counts from caecum and large intestine were higher in the experimental groups than in control mice. The aerobic and anaerobic flora in stomach and small intestine comprised mainly Gram-positive nonfusiform shaped rods. In the caecum and colon Gram-positive cocci predominated in the aerobic culture while in the anaerobic culture fusiform-shaped rods were prominent. Scanning electron microscopy of oesophagus, ileum, caecum and faeces demonstrated colonization of the oesophageal epithelium only after association with 71 bacterial strains; the filamentous bacteria present in the ileum of SPF mice were not found in the experimental groups and caecum and faeces contained mainly fusiform-shaped bacteria. Nonbacterial matter decreased in the caecum and faeces with increase in the complexity of the flora.

Scanning electron micrographs are presented of the ileal epithelium of mice aged 5, 15, 20 and 25 days. During this period the villous pattern develops to full maturity. By the twentieth day of life a segmented filamentous micro-organism colonizes the ileal epithelium and is firmly attached via a small segment. During the first days of colonization the segmented filamentous micro-organisms themselves are subcolonized by small rod-shaped bacteria, presumably lactobacilli. At the age of 25 days this subcolonization was no longer observed.

Hysterectomy-derived germ-free rabbits were given colonization-resistant caecal flora (CRF) from mice, or microflora obtained from the caecum of an antibiotic-decontaminated conventional rabbit and compared with rabbits conventionally raised with the doe. Bodyweight and the following intestinal parameters were determined for the 3 groups: colonization resistance to E. coli, relative caecal weight, villus:crypt ratio (ileum), β-aspartylglycine (faeces), volatile fatty acids (caecum), and bile acids (faeces). Germ-free rabbits given mouse CRF-flora showed values quite different from control animals (or most parameters, indicating unsuitability of mouse CRF flora to 'normalize' rabbits. In germ-free rabbits given modified (antibiotic-treated) rabbit flora, values for most parameters were intermediate between those found for the other 2 groups. This species-specific caecal flora should be improved to provide full 'normalization' of germ-free rabbits.