Aim-To investigate the colonisation with Bifidobacterium breve of the bowels of very low birthweight (VLBW) infants. Methods-The adverse eVects of B breve were examined in 66 VLBW infants (preliminary study). A prospective randomised clinical study of 91 VLBW infants was also completed and these infants were followed up for three years. Precise viable bacterial counts of serial stool specimens were examined for the first eight weeks after birth in 10 infants. The colonisation rates of administered bacteria were examined using immunohistochemical staining of stool specimens with a B breve specific monoclonal antibody. Results-In the preliminary study there were no side eVects attributable to the bacteria. Immunohistochemical staining of stool specimens showed that the colonisation rates of the administered bacteria were 73% at 2 weeks of age, but only 12% in the control group. Early administration of B breve significantly decreased aspirated air volume from the stomach and improved weight gain. Conclusions-B breve can colonise the immature bowel very eVectively and is associated with fewer abnormal abdominal signs and better weight gain in VLBW infants, probably as a result of stabilisation of their intestinal flora and accelerated feeding schedules. (Arch Dis Child 1997;76:F101-F107)

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Selective adhesion to only certain epithelia is particularly common among the bacterial members of the indigenous microflora of mammals. We have found that the stratified squamous epithelium of the nonsecreting area of horse stomach is colonized by gram-positive rods. The microscopic features of a dense layer of these bacteria on the epithelium were found to be similar to those reported in mice, rats, and swine. Adhering microorganisms were isolated and identified as Lactobacillus salivarius, L. crispatus, L. reuteri, and L. agilis by DNA-DNA hybridization and 16S rRNA gene sequencing techniques. These lactobacilli associated with the horse, except for L. reuteri, were found to adhere to horse epithelial cells in vitro but not to those of rats. A symbiotic relationship of these lactobacilli with the horse is suggested.In a series of studies on the relationships between intestinal microflora and host animals, we have demonstrated that lactobacilli indigenous to and dominant in the upper digestive tract control the population levels of other bacterial species in the stomach and the upper small intestine (13,25). We have also shown that indigenous lactobacilli can attach host specifically to keratinized epithelial cells of the rat stomach in vitro (20).In our recent study examining microbial colonization of the intestinal tract in newborn foals (15), we noticed a dense layer of gram-positive rods on the stratified squamous epithelium in the nonsecreting area of the stomach of the horse. This is the first report on the isolation and identification of indigenous Lactobacillus species adhering to the horse stomach. MATERIALS AND METHODSPreparation of specimens for isolation of bacteria and histology. Samples of the nonsecreting area of the stomach, obtained from a healthy 7-year-old female thoroughbred immediately after euthanasia, were washed three times with vigorous agitation in buffered saline (BS; 0.8% NaCl, 0.121% K 2 HPO 4 , 0.034% KH 2 PO 4 ; pH 7.2). A schematic drawing of the sampling locations of the horse stomach is shown in Fig. 1. After washing the specimen, some parts of the tissue were fixed with formalin and then embedded in paraffin, processed for histology, and stained with hematoxylin and eosin and the Gram stain. Epithelial cells were recovered from the remaining portion of fresh tissue by scraping and then homogenized in a Teflon grinder. The homogenates were plated at appropriate dilutions on MRS agar (Difco Laboratories, Detroit, Mich.). The plates were incubated anaerobically (AnaeroPack; Mitsubishi Gas Chemical Co., Inc., Tokyo, Japan) at 37°C for 72 to 96 h. Before homogenization, a portion of epithelial cells were placed on slides, stained with Giemsa stain, and examined for the detection of adherent bacteria by microscope.DNA preparation. Bacteria were grown in MRS broth overnight at 37°C. Chromosomal DNA to be used as the template for RAPD [random(ly) amplified polymorphic DNA] PCR and 16S rRNA gene amplification was prepared from bacterial strains by the method of Zhu et al. (27). Intact...

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Lactobacillus equi sp. nov. is described on the basis of 18 strains isolated as one of the predominant intestinal lactobacilli from horse faecal specimens. These 18 strains were isolated from 10 horses of 6 different farms out of 20 horses of 10 farms examined. They were Gram-positive, facultatively anaerobic, catalase-negative, non-spore-forming, non-motile, lactic-acidhomofermentative rods. The DNA GMC content was 389 O08 mol %. DNA-DNA hybridization failed to associate these strains closely with any of the validly described type strains used. Analysis of the 16S rRNA gene sequence of representative strain YIT 0455 T revealed that the new isolates represent a new Lactobacillus species, for which the name Lactobacillus equi is proposed. The type strain is YIT 0455 T (l ATCC BAA-261 T l JCM 10991 T ).

We evaluated the effects of ingestion of a synbiotic fermented milk beverage containing Lactobacillus casei strain Shirota (LcS) at 3 × 10 10 and transgalactosylated oligosaccharides (GOS) at 2.5 g per 80 ml (once a day, 2 weeks) on the defecation frequency in 35 female university students with constipation as well as the defecation frequency, intestinal microflora, and intestinal environment in elderly persons in whom the intestinal microflora and the levels of putrefactive metabolites were abnormal in a placebo-controlled double-blind study. In the female students, the defecation frequency after 1 week of synbiotic fermented milk beverage ingestion was significantly higher than that after 1 week of placebo ingestion or before ingestion. In the elderly persons, the fecal Bifidobacterium and Lactobacillus bacterial counts after 1 and 2 weeks of synbiotic fermented milk beverage ingestion were significantly higher than those after placebo ingestion or before ingestion (p<0.05 and p<0.01, respectively). The fecal lecithinasepositive Clostridium bacterial count after 1 week of synbiotic fermented milk beverage ingestion and the fecal Enterobacteriaceae bacterial counts after 1 and 2 weeks of synbiotic fermented milk beverage ingestion were significantly lower than those after placebo ingestion (p<0.05). LcS at 10 7 CFU per gram of stool was collected during the ingestion period. The acetic acid levels after 1 and 2 weeks of synbiotic fermented milk beverage ingestion were significantly higher than those after placebo ingestion (p<0.01). The stool pH values after 1 and 2 weeks of synbiotic fermented milk beverage ingestion and the ammonia and phenol levels after 2 weeks of synbiotic fermented milk beverage ingestion were significantly lower than those after placebo ingestion (p<0.05). These results suggest that ingestion of the synbiotic fermented milk beverage containing LcS and GOS improves the stool quality, intestinal microflora, and intestinal environment.