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The aim was to determine whether orally administered ovine serum immunoglobulins modulate the gut microbiota in the growing rat. Thirty Sprague-Dawley male rats were used in a 21-d study and fed either a basal control diet (control; no immunoglobulin) or a similar diet containing freeze-dried ovine immunoglobulin (ovine Ig) with 15 individually fed rats per diet. Bacterial DNA isolated from ileal and colonic digesta were subjected to PCR-denaturing gradient gel electrophoresis (PCR-DGGE). In the ileum, the DGGE band number and diversity index were greater (P < 0.05) for rats fed the ovine Ig than those fed the control diet. The DNA sequencing of a selected DGGE band in the ovine Ig-fed rats revealed 99% similarity to the Lactobacillus strains. The quantitative PCR data revealed that supplementation of the diet with the ovine Ig fraction supported the growth of Lactobacillus and conversely decreased the number of enterobacteria in ileal and colonic digesta. Inclusion of the ovine Ig fraction led to a greater (P < 0.05) ratio for total Lactobacillus to total bacteria and total Lactobacillus to enterobacteria. The results from the present study show that dietary supplementation with ovine Ig may alter the intestinal environment by a specific enrichment of Lactobacillus strains and depletion of enterobacteria.

Tyrosine-mediated repression of aroF and tyrP was studied by inserting DNA sequences between the two adjacent TYR R boxes which, in each case, overlap the respective RNA polymerase binding sites of these genes. In both cases, repression was greatest when homologous regions of these two TYR R boxes were on the same face of the DNA helix and the boxes were directly adjacent. An insertion of 3 bases was sufficient to abolish repression, which was reestablished as the boxes became separated by one full turn of the helix. These observations, coupled with the results of in vitro DNase I protection studies, supported the hypothesis that the binding of TyrR protein to the downstream boxes required cooperative interaction with TyrR protein already bound to the upstream boxes. In the case of tyrP, moving the upstream box also affected activation. Maximal activation was observed when the box was moved 3 or 12 to 14 residues upstream. Practically no activation was seen at intermediate positions, such as +7 and -4. It is hypothesized that these results indicate positions allowing maximal interaction between TyrR protein bound to the upstream box and RNA polymerase bound to the RNA polymerase binding site.

In a previous report it had been suggested that the tyrP gene of Escherichia coli may be expressed from two separate promoters. We have endeavored to confirm this suggestion by primer extension studies and the separate subcloning of each of these promoters. In these studies, we found a single promoter whose expression was repressed by TyrR protein in the presence of tyrosine and activated by TyrR protein in the presence of phenylalanine. Two adjacent TYR R boxes, with the downstream one overlapping the tyrP promoter, are the likely targets for the action of TyrR protein. Mutational analysis showed that both TYR R boxes were required for tyrosine-mediated repression but that only the upstream box was required for phenylalanine-mediated activation. In vitro DNase protection studies established that whereas in the absence of tyrosine TyrR protein protected the region of DNA represented by the upstream box, at low TyrR protein concentrations both tyrosine and ATP were required to protect the region of DNA involving the downstream box and overlapping the RNA polymerase binding site.