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There are many differences in diet and lifestyle across Europe that may influence the development of the infant gut microbiota. This work aimed to assess the impact of geographic area, mode of delivery, feeding method, and antibiotic treatment on the fecal microbiota of infants from 5 European countries with different lifestyle characteristics Sweden, Scotland, Germany, Italy, and Spain.

There are many differences in diet and lifestyle across Europe that may influence the development of the infant gut microbiota. This work aimed to assess the impact of geographic area, mode of delivery, feeding method, and antibiotic treatment on the fecal microbiota of infants from 5 European countries with different lifestyle characteristics: Sweden, Scotland, Germany, Italy, and Spain. Fecal samples from 606 infants (age 6 weeks) recruited within the European project INFABIO were analyzed by fluorescent in situ hybridization combined with flow cytometry using a panel of 10 rRNA targeted group- and species-specific oligonucleotide probes. Information on factors potentially affecting gut microbiota composition was collected with questionnaires and associations were evaluated with multivariate analyses. The Bifidobacteriumgenus was predominant (40% average proportion of total detectable bacteria), followed by Bacteroides(11.4%) and enterobacteria (7.5%). Northern European countries were associated with higher proportions of bifidobacteria in infant feces, whereas a more diverse microbiota with more bacteroides characterized southern countries. Bifidobacteria dominated the microbiota of breast-fed infants, whereas formula-fed babies had significantly higher proportions of Bacteroidesand members of the Clostridium coccoidesand Lactobacillusgroups. Newborns delivered by cesarean section or from mothers treated with antibiotics perinatally had lower proportions of Bacteroidesand members of the Atopobiumcluster. Delivery mode and feeding method influenced the fecal microbiota of European infants at 6 weeks, as expected, but the effect of country of birth was more pronounced, with dominant bifidobacteria in northern countries and greater early diversification in southern European countries.

Growth temperature was found to control the expression of toxins A and B in Clostridium difficile VPI 10463, with a maximum at 37 degrees C and low levels at 22 and 42 degrees C in both peptone yeast (PY) and defined media. The up-regulation of toxin A and B mRNA and protein levels upon temperature upshift from 22 to 37 degrees C followed the same kinetics, showing that temperature control occurred at the level of transcription. Experiments with Clostridium perfringens using gusA as a reporter gene demonstrated that both toxin gene promoters were temperature controlled and that their high activity at 37 degrees C was dependent on the alternative sigma factor TcdD. Furthermore, tcdD was found to be autoinduced at 37 degrees C. Glucose down-regulated all these responses in the C. perfringens constructs, similar to its impact on toxin production in C. difficile PY broth cultures. C. difficile proteins induced at 37 degrees C and thus coregulated with the toxins by temperature were demonstrated by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and identified as enzymes involved in butyric acid production and as electron carriers in oxidation-reduction reactions. The regulation of toxin production in C. difficile by temperature is a novel finding apparently reflecting an adaptation of the expression of its virulence to mammalian hosts.

ABSTRACTGrowth temperature was found to control the expression of toxins A and B in Clostridium difficileVPI 10463,with a maximum at 37°C and low levels at 22 and 42°C in both peptone yeast (PY) and defined media. The up-regulation of toxin A and B mRNA and protein levels upon temperature upshift from 22 to 37°C followed the same kinetics, showing that temperature control occurred at the level of transcription. Experiments with Clostridium perfringensusing gusAas a reporter gene demonstrated that both toxin gene promoters were temperature controlled and that their high activity at 37°C was dependent on the alternative sigma factor TcdD. Furthermore, tcdDwas found to be autoinduced at 37°C. Glucose down-regulated all these responses in the C. perfringensconstructs, similar to its impact on toxin production in C. difficilePY broth cultures. C. difficileproteins induced at 37°C and thus coregulated with the toxins by temperature were demonstrated by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and identified as enzymes involved in butyric acid production and as electron carriers in oxidation-reduction reactions. The regulation of toxin production in C. difficileby temperature is a novel finding apparently reflecting an adaptation of the expression of its virulence to mammalian hosts.

AbstractBackground: Several bacterial strains, mainly those belonging to the genera Lactobacillus, Enterococcus, Streptococcus and Bifidobacterium, are currently used as probiotics.Objective: The aim of this study was to investigate the influence of probiotics on . ve biochemical microbial-related functions in vitro and in vivo, in gnotobiotic mice, by applying the GAC:MAC (germ-free animal characteristic:micro oraassociated characteristic) concept.Design: Sixteen probiotics were monoinoculated in vitro and:or monoassociated to germ-free mice to investigate the following biochemical parameters in large intestinal samples: inactivation of tryptic activity, degradation of b-aspartylglycine and of mucin, conversion of bilirubin to urobilinogen and β-glucuronidase activity.Results: Lactobacillus reuteri 2010, L. rhamnosus strain 271, L. rhamnosus ATCC7469 and L. rhamnosus GG ATCC53103 expressed β-glucuronidase activity. The other parameters tested were not in uenced by any of the probiotics.Conclusions: Bacterial β-glucuronidases have an important intestinal function. As many microbial—host and microbe—diet interactions occur in the gastrointestinal tract of mammals, microbial-related functions should be studied in greater detail in probiotics.

ABSTRACTIt was recently found that a mixture of nine amino acids down-regulate Clostridium difficiletoxin production when added to peptone yeast extract (PY) cultures of strain VPI 10463 (S. Karlsson, L. G. Burman, and T. Åkerlund, Microbiology 145:1683–1693, 1999). In the present study, seven of these amino acids were found to exhibit a moderate suppression of toxin production, whereas proline and particularly cysteine had the greatest impact, on both reference strains (n= 6) and clinical isolates (n= 28) of C. difficile(>99% suppression by cysteine in the highest toxin-producing strain). Also, cysteine derivatives such as acetylcysteine, glutathione, and cystine effectively down-regulated toxin expression. An impact of both cysteine and cystine but not of thioglycolate on toxin yield indicated that toxin expression was not regulated by the oxidation-reduction potential. Several metabolic pathways, including butyric acid and butanol production, were coinduced with the toxins in PY and down-regulated by cysteine. The enzyme 3-hydroxybutyryl coenzyme A dehydrogenase, a key enzyme in solventogenesis in Clostridium acetobutylicum, was among the most up-regulated proteins during high toxin production. The addition of butyric acid to various growth media induced toxin production, whereas the addition of butanol had the opposite effect. The results indicate a coupling between specific metabolic processes and toxin expression in C. difficileand that certain amino acids can alter these pathways coordinately. We speculate that down-regulation of toxin production by the administration of such amino acids to the colon may become a novel approach to prophylaxis and therapy for C. difficile-associated diarrhea.

To evaluate the influence of clindamycin, given alone or with Lactobacillus delbrueckiiand Streptococcus thermophilusto conventional rats, on 7α-dehydroxylation of cholic acid to deoxycholic acid.

Objectives: To evaluate the influence of clindamycin, given alone or with Lactobacillus delbrueckiiand Streptococcus thermophilusto conventional rats, on 7α‐dehydroxylation of cholic acid to deoxycholic acid.