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Background: The intestinal microbial communities and their temporal dynamics are gaining increasing interest due to the significant implications for human health. Recent studies have shown the dynamic behavior of the gut microbiota in free-living, healthy persons. To date, it is not known whether these dynamics are applicable during prolonged life sharing in a confined and controlled environment., Results: The MARS500 project, the longest ground-based space simulation ever, provided us with a unique opportunity to trace the crew microbiota over 520 days of isolated confinement, such as that faced by astronauts in real long-term interplanetary space flights, and after returning to regular life, for a total of 2 years. According to our data, even under the strictly controlled conditions of an enclosed environment, the human gut microbiota is inherently dynamic, capable of shifting between different steady states, typically with rearrangements of autochthonous members. Notwithstanding a strong individuality in the overall gut microbiota trajectory, some key microbial components showed conserved temporal dynamics, with potential implications for the maintenance of a health-promoting, mutualistic microbiota configuration., Conclusions: Sharing life in a confined habitat does not affect the resilience of the individual gut microbial ecosystem, even in the long term. However, the temporal dynamics of certain microbiota components should be monitored when programming future mission simulations and real space flights, to prevent breakdowns in the metabolic and immunological homeostasis of the crewmembers.

Background: Bioinformatics tools available for metagenomic sequencing analysis are principally devoted to the identification of microorganisms populating an ecological niche, but they usually do not consider viruses. Only some software have been designed to profile the viral sequences, however they are not efficient in the characterization of viruses in the context of complex communities, like the intestinal microbiota, containing bacteria, archeabacteria, eukaryotic microorganisms and viruses. In any case, a comprehensive description of the host-microbiota interactions can not ignore the profile of eukaryotic viruses within the virome, as viruses are definitely critical for the regulation of the host immunophenotype., Results: ViromeScan is an innovative metagenomic analysis tool that characterizes the taxonomy of the virome directly from raw data of next-generation sequencing. The tool uses hierarchical databases for eukaryotic viruses to unambiguously assign reads to viral species more accurately and >1000 fold faster than other existing approaches. We validated ViromeScan on synthetic microbial communities and applied it on metagenomic samples of the Human Microbiome Project, providing a sensitive eukaryotic virome profiling of different human body sites., Conclusions: ViromeScan allows the user to explore and taxonomically characterize the virome from metagenomic reads, efficiently denoising samples from reads of other microorganisms. This implies that users can fully characterize the microbiome, including bacteria and viruses, by shotgun metagenomic sequencing followed by different bioinformatic pipelines.

Background: Lactobacillus rhamnosus is a non-starter lactic acid bacterium that plays a significant role during cheese ripening, leading to the formation of flavor. In long-ripened cheeses it persists throughout the whole time of ripening due to its capacity to adapt to changing environmental conditions. The versatile adaptability of L. rhamnosus to different ecosystems has been associated with the capacity to use non-conventional energy sources, regulating different metabolic pathways. However, the molecular mechanisms allowing the growth of L. rhamnosus in the cheese dairy environment are still poorly understood. The aim of the present study was to identify genes potentially contributing to the growth ability of L. rhamnosus PR1019 in cheese-like medium (CB) using a transcriptomic approach, based on cDNA-amplified fragment length polymorphism (cDNA-AFLP) and quantitative real-time reverse transcription-PCR (qPCR)., Results: Using three primer combinations, a total of 89 and 98 transcript-derived fragments were obtained for L. rhamnosus PR1019 grown in commercial MRS medium and CB, respectively. The cDNA-AFLP results were validated on selected regulated genes by qPCR. In order to investigate the main adaptations to growth in a cheese-mimicking system, we focused on 20 transcripts over-expressed in CB with respect to MRS. It is worth noting the presence of transcripts involved in the degradation of pyruvate and ribose. Pyruvate is a intracellular metabolite that can be produced through different metabolic routes starting from the carbon sources present in cheese, and can be released in the cheese matrix with the starter lysis. Similarly the ribonucleosides released with starter lysis could deliver ribose that represents a fermentable carbohydrate in environments, such as cheese, where free carbohydrates are lacking.Both pyruvate degradation and ribose catabolism induce a metabolite flux toward acetate, coupled with ATP production via acetate kinase. Taking into account these considerations, we suggest that the energy produced through these pathways may concur to explain the great ability of L. rhamnosus PR1019 to grow on CB., Conclusions: By a transcriptomic approach we identified a set of genes involved in alternative metabolic pathways in L. rhamnosus that could be responsible for L. rhamnosus growth in cheese during ripening.

Background: The vaginal microbiota of healthy women consists of a wide variety of anaerobic and aerobic bacterial genera and species dominated by the genus Lactobacillus. The activity of lactobacilli helps to maintain the natural healthy balance of the vaginal microbiota. This role is particularly important during pregnancy because vaginal dismicrobism is one of the most important mechanisms for preterm birth and perinatal complications. In the present study, we characterized the impact of a dietary supplementation with the probiotic VSL#3, a mixture of Lactobacillus, Bifidobacterium and Streptococcus strains, on the vaginal microbiota and immunological profiles of healthy women during late pregnancy., Results: An association between the oral intake of the probiotic VSL#3 and changes in the composition of the vaginal microbiota of pregnant women was revealed by PCR-DGGE population profiling. Despite no significant changes were found in the amounts of the principal vaginal bacterial populations in women administered with VSL#3, qPCR results suggested a potential role of the probiotic product in counteracting the decrease of Bifidobacterium and the increase of Atopobium, that occurred in control women during late pregnancy. The modulation of the vaginal microbiota was associated with significant changes in some vaginal cytokines. In particular, the decrease of the anti-inflammatory cytokines IL-4 and IL-10 was observed only in control women but not in women supplemented with VSL#3. In addition, the probiotic consumption induced the decrease of the pro-inflammatory chemokine Eotaxin, suggesting a potential anti-inflammatory effect on the vaginal immunity., Conclusion: Dietary supplementation with the probiotic VSL#3 during the last trimester of pregnancy was associated to a modulation of the vaginal microbiota and cytokine secretion, with potential implications in preventing preterm birth., Trial Registration: ClinicalTrials.gov NCT01367470.

Background: Playing a strategic role in the host immune function, the intestinal microbiota has been recently hypothesized to be involved in the etiology of atopy. In order to investigate the gastrointestinal microbial ecology of atopic disease, here we performed a pilot comparative molecular analysis of the faecal microbiota in atopic children and healthy controls., Results: Nineteen atopic children and 12 healthy controls aged 4-14 years were enrolled. Stools were collected and the faecal microbiota was characterized by means of the already developed phylogenetic microarray platform, HTF-Microbi.Array, and quantitative PCR. The intestinal microbiota of atopic children showed a significant depletion in members of the Clostridium cluster IV, Faecalibacterium prausnitzii, Akkermansia muciniphila and a corresponding increase of the relative abundance of Enterobacteriaceae., Conclusion: Depleted in key immunomodulatory symbionts, the atopy-associated microbiota can represent an inflammogenic microbial consortium which can contribute to the severity of the disease. Our data open the way to the therapeutic manipulation of the intestinal microbiota in the treatment of atopy by means of pharmaceutical probiotics.

Background: Affecting the core functional microbiome, peculiar high level taxonomic unbalances of the human intestinal microbiota have been recently associated with specific diseases, such as obesity, inflammatory bowel diseases, and intestinal inflammation., Results: In order to specifically monitor microbiota unbalances that impact human physiology, here we develop and validate an original DNA-microarray (HTF-Microbi.Array) for the high taxonomic level fingerprint of the human intestinal microbiota. Based on the Ligase Detection Reaction-Universal Array (LDR-UA) approach, the HTF-Microbi.Array enables specific detection and approximate relative quantification of 16S rRNAs from 30 phylogenetically related groups of the human intestinal microbiota. The HTF-Microbi.Array was used in a pilot study of the faecal microbiota of eight young adults. Cluster analysis revealed the good reproducibility of the high level taxonomic microbiota fingerprint obtained for each of the subject., Conclusion: The HTF-Microbi.Array is a fast and sensitive tool for the high taxonomic level fingerprint of the human intestinal microbiota in terms of presence/absence of the principal groups. Moreover, analysis of the relative fluorescence intensity for each probe pair of our LDR-UA platform can provide estimation of the relative abundance of the microbial target groups within each samples. Focusing the phylogenetic resolution at division, order and cluster levels, the HTF-Microbi.Array is blind with respect to the inter-individual variability at the species level.

Background: The human gut harbors a diverse community of microorganisms which serve numerous important functions for the host wellbeing. Functional foods are commonly used to modulate the composition of the gut microbiota contributing to the maintenance of the host health or prevention of disease. In the present study, we characterized the impact of one month intake of a synbiotic food, containing fructooligosaccharides and the probiotic strains Lactobacillus helveticus Bar13 and Bifidobacterium longum Bar33, on the gut microbiota composition and metabolic profiles of 20 healthy subjects., Results: The synbiotic food did not modify the overall structure of the gut microbiome, as indicated by Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE). The ability of the probiotic L. helveticus and B. longum strains to pass through the gastrointestinal tract was hypothesized on the basis of real-time PCR data. In spite of a stable microbiota, the intake of the synbiotic food resulted in a shift of the fecal metabolic profiles, highlighted by the Gas Chromatography Mass Spectrometry Solid Phase Micro-Extraction (GC-MS/SPME) analysis. The extent of short chain fatty acids (SCFA), ketones, carbon disulfide and methyl acetate was significantly affected by the synbiotic food consumption. Furthermore, the Canonical discriminant Analysis of Principal coordinates (CAP) of GC-MS/SPME profiles allowed a separation of the stool samples recovered before and after the consumption of the functional food., Conclusion: In this study we investigated the global impact of a dietary intervention on the gut ecology and metabolism in healthy humans. We demonstrated that the intake of a synbiotic food leads to a modulation of the gut metabolic activities with a maintenance of the gut biostructure. In particular, the significant increase of SCFA, ketones, carbon disulfide and methyl acetate following the feeding period suggests potential health promoting effects of the synbiotic food.