Lactobacillus ruminis strains cluster according to their mammalian gut source

peer-reviewedBackground Lactobacillus ruminis is a motile Lactobacillus that is autochthonous to the human gut, and which may also be isolated from other mammals. Detailed characterization of L. ruminis has previously been restricted to strains of human and bovine origin. We therefore sought to expand our bio-bank of strains to identify and characterise isolates of porcine and equine origin by comparative genomics. Results We isolated five strains from the faeces of horses and two strains from pigs, and compared their motility, biochemistry and genetic relatedness to six human isolates and three bovine isolates including the type strain 27780T. Multilocus sequence typing analysis based on concatenated sequence data for six individual loci separated the 16 L. ruminis strains into three clades concordant with human, bovine or porcine, and equine sources. Sequencing the genomes of four additional strains of human, bovine, equine and porcine origin revealed a high level of genome synteny, independent of the source animal. Analysis of carbohydrate utilization, stress survival and technological robustness in a combined panel of sixteen L. ruminis isolates identified strains with optimal survival characteristics suitable for future investigation as candidate probiotics. Under laboratory conditions, six human isolates of L. ruminis tested were aflagellate and non-motile, whereas all 10 strains of bovine, equine and porcine origin were motile. Interestingly the equine and porcine strains were hyper-flagellated compared to bovine isolates, and this hyper-flagellate phenotype correlated with the ability to swarm on solid medium containing up to 1.8% agar. Analysis by RNA sequencing and qRT-PCR identified genes for the biosynthesis of flagella, genes for carbohydrate metabolism and genes of unknown function that were differentially expressed in swarming cells of an equine isolate of L. ruminis. Conclusions We suggest that Lactobacillus ruminis isolates have potential to be used in the functional food industry. We have also identified a MLST scheme able to distinguish between strains of L. ruminis of different origin. Genes for non-digestible oligosaccharide metabolism were identified with a putative role in swarming behaviour.This work was supported by a Principal Investigator Award (07/IN.1/B1780) from Science Foundation Ireland to P.W. O’Toole

Composition and temporal stability of the gut microbiota in older persons

The composition and function of the human gut microbiota has been linked to health and disease. We previously identified correlations between habitual diet, microbiota composition gradients and health gradients in an unstratified cohort of 178 elderly subjects. To refine our understanding of diet–microbiota associations and differential taxon abundance, we adapted an iterative bi-clustering algorithm (iterative binary bclustering of gene sets (iBBiG)) and applied it to microbiota composition data from 732 faecal samples from 371 ELDERMET cohort subjects, including longitudinal samples. We thus identified distinctive microbiota configurations associated with ageing in both community and long-stay residential care elderly subjects. Mixed-taxa populations were identified that had clinically distinct associations. Microbiota temporal instability was observed in both community-dwelling and long-term care subjects, particularly in those with low initial microbiota diversity. However, the stability of the microbiota of subjects had little impact on the directional change of the microbiota as observed for long-stay subjects who display a gradual shift away from their initial microbiota. This was not observed in community-dwelling subjects. This directional change was associated with duration in long-stay. Changes in these bacterial populations represent the loss of the health-associated and youth-associated microbiota components and gain of an elderly associated microbiota. Interestingly, community-associated microbiota configurations were impacted more by the use of antibiotics than the microbiota of individuals in long-term care, as the community-associated microbiota showed more loss but also more recovery following antibiotic treatment. This improved definition of gut microbiota composition patterns in the elderly will better inform the design of dietary or antibiotic interventions targeting the gut microbiota

Improved Food Safety through Sterility of Air in Food Processing and Packaging.

End of Project ReportThe aims of this research were to develop reliable methods for evaluating the level of airborne micro-organisms in food processing facilities and to study the viability and behaviour of micro-organisms in air filtration systems. The main conclusions were as follows: · Assessment of air quality involves monitoring both particulate and microbial levels as there is no simple relationship between these phenomena in a food process environment. · The large fluctuations in microbial levels found in air in these studies underline the necessity of frequent and regular air sampling in processing facilities. · It was established that micro-organisms can grow inside an air filter under certain environmental conditions and give rise to intermittent microbial germ contamination of the cleaned air. · It was demonstrated that flowing air affects the survival of microorganisms and the survival rate is dependent on filter class. Hence more emphasis on filter design aimed at effective microbial control is advised. · A combined system for filtering and sterilisation by ozone was demonstrated to be an efficient technique for extending the microbial separation efficiency of air filters.European Unio

Chromosomal G + C Content Evolution in Yeasts: Systematic Interspecies Differences, and GC-Poor Troughs at Centromeres

The G + C content at synonymous codon positions (GC3s) in genes varies along chromosomes in most eukaryotes. In Saccharomyces cerevisiae, regions of high GC3s are correlated with recombination hot spots, probably due to biased gene conversion. Here we examined how GC3s differs among groups of related yeast species in the Saccharomyces and Candida clades. The chromosomal locations of GC3s peaks and troughs are conserved among four Saccharomyces species, but we find that there have been highly consistent small shifts in their GC3s values. For instance, 84% of all S. cerevisiae genes have a lower GC3s value than their S. bayanus orthologs. There are extensive interspecies differences in the Candida clade both in the median value of GC3s (ranging from 22% to 49%) and in the variance of GC3s among genes. In three species—Candida lusitaniae, Pichia stipitis, and Yarrowia lipolytica—there is one region on each chromosome in which GC3s is markedly reduced. We propose that these GC-poor troughs indicate the positions of centromeres because in Y. lipolytica they coincide with the five experimentally identified centromeres. In P. stipitis, the troughs contain clusters of the retrotransposon Tps5. Likewise, in Debaryomyces hansenii, there is one cluster of the retrotransposon Tdh5 per chromosome, and all these clusters are located in GC-poor troughs. Locally reduced G + C content around centromeres is consistent with a model in which G + C content correlates with recombination rate, and recombination is suppressed around centromeres, although the troughs are unexpectedly wide (100–300 kb)

Effect of room temperature transport vials on DNA quality and phylogenetic composition of faecal microbiota of elderly adults and infants

Background: Alterations in intestinal microbiota have been correlated with a growing number of diseases. Investigating the faecal microbiota is widely used as a non-invasive and ethically simple proxy for intestinal biopsies. There is an urgent need for collection and transport media that would allow faecal sampling at distance from the processing laboratory, obviating the need for same-day DNA extraction recommended by previous studies of freezing and processing methods for stool. We compared the faecal bacterial DNA quality and apparent phylogenetic composition derived using a commercial kit for stool storage and transport (DNA Genotek OMNIgene GUT) with that of freshly extracted samples, 22 from infants and 20 from older adults. Results: Use of the storage vials increased the quality of extracted bacterial DNA by reduction of DNA shearing. When infant and elderly datasets were examined separately, no differences in microbiota composition were observed due to storage. When the two datasets were combined, there was a difference according to a Wilcoxon test in the relative proportions of Faecalibacterium, Sporobacter, Clostridium XVIII, and Clostridium XlVa after 1 week's storage compared to immediately extracted samples. After 2 weeks' storage, Bacteroides abundance was also significantly different, showing an apparent increase from week 1 to week 2. The microbiota composition of infant samples was more affected than that of elderly samples by storage, with significantly higher Spearman distances between paired freshly extracted and stored samples (

A long and abundant non-coding RNA in Lactobacillus salivarius

Lactobacillus salivarius, found in the intestinal microbiota of humans and animals, is studied as an example of the sub-dominant intestinal commensals that may impart benefits upon their host. Strains typically harbour at least one megaplasmid that encodes functions contributing to contingency metabolism and environmental adaptation. RNA sequencing (RNA-seq) transcriptomic analysis of L. salivarius strain UCC118 identified the presence of a novel unusually abundant long non-coding RNA (lncRNA) encoded by the megaplasmid, and which represented more than 75 % of the total RNA-seq reads after depletion of rRNA species. The expression level of this 520 nt lncRNA in L. salivarius UCC118 exceeded that of the 16S rRNA, it accumulated during growth, was very stable over time and was also expressed during intestinal transit in a mouse. This lncRNA sequence is specific to the L. salivarius species; however, among 45 L. salivarius genomes analysed, not all (only 34) harboured the sequence for the lncRNA. This lncRNA was produced in 27 tested L. salivarius strains, but at strain-specific expression levels. High-level lncRNA expression correlated with high megaplasmid copy number. Transcriptome analysis of a deletion mutant lacking this lncRNA identified altered expression levels of genes in a number of pathways, but a definitive function of this new lncRNA was not identified. This lncRNA presents distinctive and unique properties, and suggests potential basic and applied scientific developments of this phenomenon

Tumour-associated and non-tumour-associated microbiota in colorectal cancer

Objective: A signature that unifies the colorectal cancer (CRC) microbiota across multiple studies has not been identified. In addition to methodological variance, heterogeneity may be caused by both microbial and host response differences, which was addressed in this study. Design: We prospectively studied the colonic microbiota and the expression of specific host response genes using faecal and mucosal samples (‘ON’ and ‘OFF’ the tumour, proximal and distal) from 59 patients undergoing surgery for CRC, 21 individuals with polyps and 56 healthy controls. Microbiota composition was determined by 16S rRNA amplicon sequencing; expression of host genes involved in CRC progression and immune response was quantified by real-time quantitative PCR. Results: The microbiota of patients with CRC differed from that of controls, but alterations were not restricted to the cancerous tissue. Differences between distal and proximal cancers were detected and faecal microbiota only partially reflected mucosal microbiota in CRC. Patients with CRC can be stratified based on higher level structures of mucosal-associated bacterial co-abundance groups (CAGs) that resemble the previously formulated concept of enterotypes. Of these, Bacteroidetes Cluster 1 and Firmicutes Cluster 1 were in decreased abundance in CRC mucosa, whereas Bacteroidetes Cluster 2, Firmicutes Cluster 2, Pathogen Cluster and Prevotella Cluster showed increased abundance in CRC mucosa. CRC-associated CAGs were differentially correlated with the expression of host immunoinflammatory response genes. Conclusions: CRC-associated microbiota profiles differ from those in healthy subjects and are linked with distinct mucosal gene-expression profiles. Compositional alterations in the microbiota are not restricted to cancerous tissue and differ between distal and proximal cancers

Generating a taxonomy for genetic conditions relevant to reproductive planning

As genome or exome sequencing (hereafter genome-scale sequencing) becomes more integrated into standard care, carrier testing is an important possible application. Carrier testing using genome-scale sequencing can identify a large number of conditions, but choosing which conditions/genes to evaluate as well as which results to disclose can be complicated. Carrier testing generally occurs in the context of reproductive decision-making and involves patient values in a way that other types of genetic testing may not. The Kaiser Permanente Clinical Sequencing Exploratory Research program is conducting a randomized clinical trial of preconception carrier testing that allows participants to select their preferences for results from among broad descriptive categories rather than selecting individual conditions. This paper describes 1) the criteria developed by the research team, the return of results committee (RORC), and stakeholders for defining the categories; 2) the process of refining the categories based on input from patient focus groups and validation through a patient survey; and, 3) how the RORC then assigned specific gene-condition pairs to taxonomy categories being piloted in the trial. The development of four categories (serious, moderate/mild, unpredictable, late onset) for sharing results allows patients to select results based on their values without separately deciding their interest in knowing their carrier status for hundreds of conditions. A fifth category, lifespan limiting, was always shared. The lessons learned may be applicable in other results disclosure situations, such as incidental findings