Short-term modifications in the distal gut microbiota of weaning mice induced by a high-fat diet.

The gut microbiota has been shown to be involved in host energy homeostasis and diet-induced metabolic disorders. To gain insight into the relationships among diet, microbiota and the host, we evaluated the effects of a high-fat (HF) diet on the gut bacterial community in weaning mice. C57BL/6 mice were fed either a control diet or a diet enriched with soy oil for 1 and 2 weeks. Administration of the HF diet caused an increase in plasma total cholesterol levels, while no significant differences in body weight gain were observed between the two diets. Denaturing gradient gel electrophoresis (DGGE) profiles indicated considerable variations in the caecal microbial communities of mice on the HF diet, as compared with controls. Two DGGE bands with reduced intensities in HF-fed mice were identified as representing Lactobacillus gasseri and an uncultured Bacteroides species, whereas a band of increased intensity was identified as representing a Clostridium populeti-related species upon sequencing. Quantitative real-time PCR confirmed a statistically significant 1-log decrease in L. gasseri cell numbers after HF feeding, and revealed a significantly lower level of Bifidobacterium spp. in the control groups after 1 and 2 weeks compared with that in the HF groups. These alterations of intestinal microbiota were not associated with caecum inflammation, as assessed by histological analysis. The observed shifts of specific bacterial populations within the gut may represent an early consequence of increased dietary fat.

Impact of antibiotics on the gut microbiota of critically ill patients.

We evaluated the relationship between the intestinal microbiota composition and clinical outcome in a group of 15 high-risk patients admitted for acute infection and/or surgical/accidental trauma who were treated with systemic antibiotics according to standard intensive care unit (ICU) protocols. There was a high mortality rate amongst these patients, each of whom had a considerable organ failure score at admission, respiratory assistance during the most of their ICU stay and a long length of stay. All of these individuals received sedation and enteral nutrition, and the majority also received insulin, vasoactive drugs and some stress-ulcer prophylaxis agents. The intestinal microbiota composition was assessed using denaturing gradient gel electrophoresis (DGGE), a molecular biology tool used to characterize bacterial ecosystems. As all of the patient subjects were in good health prior to their acute illness and admission to the ICU, the first faecal samples obtained from this group showed a DGGE banding pattern that was similar to that of healthy subjects. After 1 week of critical illness, coupled with intensive care treatment, including antibiotics, a very definite alteration in the overall microbiota composition was evident, as revealed by a reduction in the number of DGGE bands. Further pronounced changes to the DGGE banding profiles could be observed in patients remaining in the ICU for 2 weeks. Moreover, a dominant band, identified by sequencing as highly related to Enterococcus, was detected in the DGGE profile of some of our patient subjects. We also performed real-time PCR and obtained results that were in agreement with our qualitative evaluations using DGGE. The degree of organ failure and ICU mortality was significantly higher in patients for whom a high reduction in microbiota biodiversity was coupled with a massive presence of enterococci. A statistically significant link between these two ecological traits and the use of clindamycin was also found.

Impact of cystic fibrosis disease on archaea and bacteria composition of gut microbiota.

Cystic fibrosis is often associated with intestinal inflammation due to several factors, including altered gut microbiota composition. In this study, we analyzed the fecal microbiota among patients with cystic fibrosis of 10-22 years of age, and compared the findings with age-matched healthy subjects. The participating patients included 14 homozygotes and 14 heterozygotes with the delF508 mutation, and 2 heterozygotes presenting non-delF508 mutations. We used PCR-DGGE and qPCR to analyze the presence of bacteria, archaea and sulfate-reducing bacteria. Overall, our findings confirmed disruption of the cystic fibrosis gut microbiota. Principal component analysis of the qPCR data revealed no differences between homozygotes and heterozygotes, while both groups were distinct from healthy subjects who showed higher biodiversity. Archaea were under the detection limit in all homozygotes subjects, whereas methanogens were detected in 62% of both cystic fibrosis heterozygotes and healthy subjects. Our qPCR results revealed a low frequency of sulfate-reducing bacteria in the homozygote (13%) and heterozygote (13%) patients with cystic fibrosis compared with healthy subjects (87.5%). This is a pioneer study showing that patients with cystic fibrosis exhibit significant reduction of H2-consuming microorganisms, which could increase hydrogen accumulation in the colon and the expulsion of this gas through non-microbial routes.

Beneficial effect of auto-aggregating Lactobacillus crispatus on experimentally induced colitis in mice.

We tested the therapeutic relevance of auto aggregation in lactobacilli by comparing the effect on DSS induced colitis of viable Lactobacillus crispatus M247, isolated from healthy humans, to L. crispatus MU5, an isogenic spontaneous mutants of M247, the latter lacking the auto aggregation phenotype which allows the adhesion to human mucus. Aggregating L. crispatus M247, but not the non-aggregating MU5, was retrievable from mice feces and adherent to the colonic mucosa. Daily administration of L. crispatus M247, but not heat killed L. crispatus M247 or aggregation deficient L. crispatus MU5, dose-dependently reduced the severity of DSS colitis. Indeed, L. crispatus MU5 administered in a 30% sucrose solution, known to restore the aggregation phenotype, had a protective effect comparable to mice receiving L. crispatus M247. These results indicate that a surface-mediated property such as aggregation may play a pivotal role in the protective effects obtained by dietary supplementation with L. crispatus M247 during colitis.

Enteral nutrition and microflora in pediatric Crohn's disease.

BACKGROUND: Exclusive enteral nutrition (EN) is an established primary therapy for pediatric Crohn's disease (CD). The mechanism of action of such treatment is still conjectural. The aim of the present study was to investigate if EN-induced remission is associated with modification of the fecal microflora in CD. METHODS: Stool samples were collected from 5 healthy children and adolescents over a period of 3 months, and from 9 children and adolescents with active CD. To induce disease remission, children with CD received a course of exclusive EN for 8 weeks with a polymeric formula (Modulen IBD, Nestlè). At the end of the course of exclusive EN, children returned to a free diet but continued to take 40% of the daily caloric intake as polymeric formula. Fecal microflora was analyzed by 16S ribosomal DNA polymerase chain reaction and temperature gradient gel electrophoresis (TGGE) with direct visual comparison of band profiles of PCR products. RESULTS: In 8 of 9 children, the exclusive EN alone induced disease remission. In 1 child, it was necessary to add steroids to the exclusive EN course to achieve remission. In all children with CD, analysis of gel band distribution revealed profound modification of the fecal microflora after exclusive EN. Variations of band distribution corresponding to different bacterial species were observed also in children on partial EN and required time to achieve stability of the band profile. In contrast, control healthy children showed a host-specific and stable TGGE profile over time. CONCLUSION: These data suggest that a possible mechanism of action of EN in inducing disease remission in CD is the capacity of modification of gut microflora. Possible explanations of such capacity are both low residue and prebiotic properties of the polymeric liquid formula.

Bacterial diversity in a contaminated Alpine glacier as determined by culture-based and molecular approaches.

Glaciers are important ecosystems, hosting bacterial communities that are adapted to cold conditions and scarcity of available nutrients. Several works focused on the composition of bacterial communities in glaciers and on the long-range atmospheric deposition of pollutants in glaciers, but it is not clear yet if ski resorts can represent a source of point pollution in near-by glaciers, and if these pollutants can influence the residing bacterial communities. To test these hypotheses, 12 samples were analyzed in Madaccio Glacier, in a 3200 ma.s.l. from two areas, one undisturbed and one close to a summer ski resort that is active since the 1930s. Chemical analyses found concentrations up to 43 ng L(-1) for PCBs and up to 168 µg L(-1) for PAHs in the contaminated area: these values are significantly higher than the ones found in undisturbed glaciers because of long-range atmospheric deposition events, and can be explained as being related to the near-by ski resort activities. Isolation of strains on rich medium plates and PCR-DGGE analyses followed by sequencing of bands allowed the identification of a bacterial community with phylogenetic patterns close to other glacier environments, with Proteobacteria and Actinobacteria the mostly abundant phyla, with Acidobacteria, Firmicutes and Cyanobacteria also represented in the culture-independent analyses. A number of isolates were identified by molecular and biochemical methods as phylogenetic related to known xenobiotic-degrading strains: glaciers subjected to chemical contamination can be important reservoirs of bacterial strains with potential applications in bioremediation.

Disrupted intestinal microbiota and intestinal inflammation in children with cystic fibrosis and its restoration with Lactobacillus GG: a randomised clinical trial.

BACKGROUND & AIMS: Intestinal inflammation is a hallmark of cystic fibrosis (CF). Administration of probiotics can reduce intestinal inflammation and the incidence of pulmonary exacerbations. We investigated the composition of intestinal microbiota in children with CF and analyzed its relationship with intestinal inflammation. We also investigated the microflora structure before and after Lactobacillus GG (LGG) administration in children with CF with and without antibiotic treatment. METHODS: The intestinal microbiota were analyzed by denaturing gradient gel electrophoresis (DGGE), real-time polymerase chain reaction (RT-PCR), and fluorescence in situ hybridization (FISH). Intestinal inflammation was assessed by measuring fecal calprotectin (CLP) and rectal nitric oxide (rNO) production in children with CF as compared with healthy controls. We then carried out a small double-blind randomized clinical trial with LGG. RESULTS: Twenty-two children with CF children were enrolled in the study (median age, 7 years; range, 2-9 years). Fecal CLP and rNO levels were higher in children with CF than in healthy controls (184±146 µg/g vs. 52±46 µg/g; 18±15 vs. 2.6±1.2 µmol/L NO2 (-), respectively; P<0.01). Compared with healthy controls, children with CF had significantly different intestinal microbial core structures. The levels of Eubacterium rectale, Bacteroides uniformis, Bacteroides vulgatus, Bifidobacterium adolescentis, Bifidobacterium catenulatum, and Faecalibacterium prausnitzii were reduced in children with CF. A similar but more extreme pattern was observed in children with CF who were taking antibiotics. LGG administration reduced fecal CLP and partially restored intestinal microbiota. There was a significant correlation between reduced microbial richness and intestinal inflammation. CONCLUSIONS: CF causes qualitative and quantitative changes in intestinal microbiota, which may represent a novel therapeutic target in the treatment of CF. Administration of probiotics restored gut microbiota, supporting the efficacy of probiotics in reducing intestinal inflammation and pulmonary exacerbations. TRIAL REGISTRATION: ClinicalTrials.gov NCT 01961661.

Survival of yogurt bacteria in the human gut.

Whether Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus can be recovered after passage through the human gut was tested by feeding 20 healthy volunteers commercial yogurt. Yogurt bacteria were found in human feces, suggesting that they can survive transit in the gastrointestinal tract.