CFP/Yit: An Inbred Mouse Strain with Slow Gastrointestinal Transit.

BACKGROUND: Gastrointestinal transit (GIT) is influenced by factors including diet, medications, genetics, and gut microbiota, with slow GIT potentially indicating a functional disorder linked to conditions, such as constipation. Although GIT studies have utilized various animal models, few effectively model spontaneous slow GIT. AIMS: We aimed to characterize the GIT phenotype of CFP/Yit (CFP), an inbred mouse strain with suggested slow GIT. METHODS: Female and male CFP mice were compared to Crl:CD1 (ICR) mice in GIT and assessed based on oral gavage of fluorescent-labeled 70-kDa dextran, feed intake, fecal amount, and fecal water content. Histopathological analysis of the colon and analysis of gut microbiota were conducted. RESULTS: CFP mice exhibited a shorter small intestine and a 1.4-fold longer colon compared to ICR mice. The median whole-GIT time was 6.0-fold longer in CFP mice than in ICR mice. CFP mice demonstrated slower gastric and cecal transits than ICR mice, with a median colonic transit time of 4.1 h (2.9-fold longer). CFP mice exhibited lower daily feed intakes and fecal amounts. Fecal water content was lower in CFP mice, apparently attributed to the longer colon. Histopathological analysis showed no changes in CFP mice, including tumors or inflammation. Moreover, CFP mice had a higher Firmicutes/Bacteroidota ratio and a relative abundance of Erysipelotrichaceae in cecal and fecal contents. CONCLUSIONS: This study indicates that CFP mice exhibit slow transit in the stomach, cecum, and colon. As a novel mouse model, CFP mice can contribute to the study of gastrointestinal physiology and disease.

Manipulating the growth environment through co-culture to enhance stress tolerance and viability of probiotic strains in the gastrointestinal tract.

IMPORTANCE: The viability of probiotics in the human gastrointestinal tract is important, as some reports indicate that the health benefits of live bacteria are greater than those of dead ones. Therefore, the higher the viability of the probiotic strain, the better it may be. However, probiotic strains lose their viability due to gastrointestinal stress such as gastric acid and bile. This study provides an example of the use of co-culture or pH-controlled monoculture, which uses more stringent conditions (lower pH) than normal monoculture to produce probiotic strains that are more resistant to gastrointestinal stress. In addition, co-cultured beverages showed higher viability of the probiotic strain in the human gastrointestinal tract than monocultured beverages in our human study.

Identification of genes essential for bile acid resistance in the probiotic Lacticaseibacillus paracasei strain Shirota.

Bile acid resistance is crucial to allow probiotic strains to survive in the gastrointestinal tract and exert health-promoting effects on their hosts. Our aim here was to determine the mechanism of this resistance via a genetic approach by identifying the genes essential for bile acid resistance in Lacticaseibacillus paracasei strain Shirota (LcS). We generated 4649 transposon-inserted lines of L. paracasei YIT 0291, which has the same genome sequence as LcS but lacks the pLY101 plasmid, and we screened them for bile-acid-sensitive mutants. The growth of 14 mutated strains was strongly inhibited by bile acid, and we identified 10 genes that could be involved in bile acid resistance. Expression of these genes was not markedly induced by bile acid, suggesting that their homeostatic expression is important for exerting bile acid resistance. Two mutants in which the transposon was independently inserted into cardiolipin synthase (cls) genes, showed strong growth inhibition. Disruption of the cls genes in LcS caused decreased cardiolipin (CL) production and the accumulation of the precursor phosphatidylglycerol in bacterial cells. These data suggest that LcS possesses several mechanisms for exerting bile acid resistance, and that homeostatic CL production is among the factors most essential for this resistance.

Reconstruction of the personal information from human genome reads in gut metagenome sequencing data.

Human DNA present in faecal samples can result in a small number of human reads in gut shotgun metagenomic sequencing data. However, it is presently unclear how much personal information can be reconstructed from such reads, and this has not been quantitatively evaluated. Such a quantitative evaluation is necessary to clarify the ethical concerns related to data sharing and to enable efficient use of human genetic information in stool samples, such as for research and forensics. Here we used genomic approaches to reconstruct personal information from the faecal metagenomes of 343 Japanese individuals with associated human genotype data. Genetic sex could be accurately predicted based on the sequencing depth of sex chromosomes for 97.3% of the samples. Individuals could be re-identified from the matched genotype data based on human reads recovered from the faecal metagenomic data with 93.3% sensitivity using a likelihood score-based method. This method also enabled us to predict the ancestries of 98.3% of the samples. Finally, we performed ultra-deep shotgun metagenomic sequencing of five faecal samples as well as whole-genome sequencing of blood samples. Using genotype-calling approaches, we demonstrated that the genotypes of both common and rare variants could be reconstructed from faecal samples. This included clinically relevant variants. Our approach can be used to quantify personal information contained within gut metagenome data.

Human gut microbiota influences drug-metabolizing enzyme hepatic Cyp3a: A human flora-associated mice study.

Several studies revealed that gut microbiota affects the hepatic drug-metabolizing enzyme cytochrome P450 (Cyp). We hypothesized that individual gut microbiota variations could contribute to CYP activity. Human flora-associated (HFA) mice are established from germ-free mice using human feces and are often used to determine the effect of the human gut microbiota on the host. This study generated two groups of HFA mice using feces from two healthy individuals. Then, the composition of gut microbiota and hepatic Cyp activity was compared to analyze the effects of gut microbiota in healthy individuals on hepatic Cyp activity. A principal coordinate analysis based on the UniFrac distance for the composition of the cecal and fecal microbiota revealed apparent differences between the recipient groups. Hepatic Cyp, which is a marked difference in Cyp3a activity and Cyp3a11 gene expression, was observed between the recipient groups. Cyp2c and Cyp1a activities did not differ between recipient groups, with significantly lower enzymatic activities in recipients than in germ-free mice. These results indicate that the human gut microbiota affects hepatic Cyp activity. Especially, human gut microbiota composition differences have a pronounced effect on Cyp3a activity via Cyp3a11 gene expression regulation. Therefore, human gut microbiota variations among individuals may affect numerous drug metabolism, leading to drug efficacy and toxicity.

Genomic repertoires linked with pathogenic potency of arthritogenic Prevotella copri isolated from the gut of patients with rheumatoid arthritis.

OBJECTIVES: Prevotella copri is considered to be a contributing factor in rheumatoid arthritis (RA). However, in some non-Westernised countries, healthy individuals also harbour an abundance of P. copri in the intestine. This study investigated the pathogenicity of RA patient-derived P. copri (P. copri RA) compared with healthy control-derived P. copri (P. copri HC). METHODS: We obtained 13 P. copri strains from the faeces of patients with RA and healthy controls. Following whole genome sequencing, the sequences of P. copri RA and P. copri HC were compared. To analyse the arthritis-inducing ability of P. copri, we examined two arthritis models (1) a collagen-induced arthritis model harbouring P. copri under specific-pathogen-free conditions and (2) an SKG mouse arthritis model under P. copri-monocolonised conditions. Finally, to evaluate the ability of P. copri to activate innate immune cells, we performed in vitro stimulation of bone marrow-derived dendritic cells (BMDCs) by P. copri RA and P. copri HC. RESULTS: Comparative genomic analysis revealed no apparent differences in the core gene contents between P. copri RA and P. copri HC, but pangenome analysis revealed the high genome plasticity of P. copri. We identified a P. copri RA-specific genomic region as a conjugative transposon. In both arthritis models, P. copri RA-induced more severe arthritis than P. copri HC. In vitro BMDC stimulation experiments revealed the upregulation of IL-17 and Th17-related cytokines (IL-6, IL-23) by P. copri RA. CONCLUSION: Our findings reveal the genetic diversity of P. copri, and the genomic signatures associated with strong arthritis-inducing ability of P. copri RA. Our study contributes towards elucidation of the complex pathogenesis of RA.

Normal variation of the gut microbiota affects hepatic cytochrome P450 activity in mice.

Several studies revealed that substantial artificial changes in the gut microbiota resulted in modification of hepatic cytochrome P450 3a (Cyp3a) in mice. Consequently, we hypothesized that "normal" variation of the gut microbiota might also alter hepatic Cyp activity and lead to individual differences in drug metabolism. Therefore, this study investigated the effects of normal gut microbiota variation on hepatic Cyp activity under the same genetic and environmental conditions using ex-germ-free mice. Using the feces of three breeder BALB/c mice (Jcl, Slc, and Crj), germ-free BALB/cYit mice were conventionalized (Yit-Jcl, Yit-Slc, and Yit-Crj). The gut microbiota composition and hepatic Cyp activity of these donors and recipients were evaluated. 16S rRNA sequencing revealed clear differences of the gut microbiota among donors and among recipients. Cyp3a activity was significantly higher in Slc mice than in Jcl and Crj mice. Notably, among recipients, Cyp3a activity was significantly higher in Yit-Slc and Yit-Crj mice than in Yit-Jcl mice. Cyp2b activity was significantly higher in Slc mice than in Jcl and Crj mice. Cyp2b activity was significantly higher in Yit-Slc mice than in Yit-Jcl mice. Additionally, in correlation analysis, some genera displayed significant positive or negative correlations with Cyp activity, particular the strong positive correlation between Clostridium sensu stricto 1 with Cyp3a activity. In conclusion, this study demonstrated that normal variation of the gut microbiota affected hepatic Cyp3a and Cyp2b activity, which might result in individual differences of drug metabolism.

Xylan utilisation promotes adaptation of Bifidobacterium pseudocatenulatum to the human gastrointestinal tract.

Dietary carbohydrates impact the composition of the human gut microbiota. However, the relationship between carbohydrate availability for individual bacteria and their growth in the intestinal environment remains unclear. Here, we show that the availability of long-chain xylans (LCX), one of the most abundant dietary fibres in the human diet, promotes the growth of Bifidobacterium pseudocatenulatum in the adult human gut. Genomic and phenotypic analyses revealed that the availability of LCX-derived oligosaccharides is a fundamental feature of B. pseudocatenulatum, and that some but not all strains possessing the endo-1,4-ß-xylanase (BpXyn10A) gene grow on LCX by cleaving the xylose backbone. The BpXyn10A gene, likely acquired by horizontal transfer, was incorporated into the gene cluster for LCX-derived oligosaccharide utilisation. Co-culturing with xylanolytic Bacteroides spp. demonstrated that LCX-utilising strains are more competitive than LCX non-utilising strains even when LCX-derived oligosaccharides were supplied. In LCX-rich dietary interventions in adult humans, levels of endogenous B. pseudocatenulatum increased only when BpXyn10A was detected, indicating that LCX availability is a fitness determinant in the human gut. Our findings highlight the enhanced intestinal adaptability of bifidobacteria via polysaccharide utilisation, and provide a cornerstone for systematic manipulation of the intestinal microbiota through dietary intervention using key enzymes that degrade polysaccharide as biomarkers.

Lypd8 inhibits attachment of pathogenic bacteria to colonic epithelia.

Mucosal barriers segregate commensal microbes from the intestinal epithelia to maintain gut homeostasis. Ly6/Plaur domain-containing 8 (Lypd8), a highly glycosylated glycosylphosphatidylinositol-anchored protein selectively expressed on colonic enterocytes, promotes this segregation by inhibiting bacterial invasion of the inner mucus layer and colonic epithelia. However, it remains unclear whether Lypd8 prevents infection with enteric bacterial pathogens. Here, we demonstrate that Lypd8 strongly contributes to early-phase defense against Citrobacter rodentium, which causes colitis by inducing attachment and effacement (A/E) lesions on colonic epithelia. Lypd8 inhibits C. rodentium attachment to intestinal epithelial cells by binding to intimin, thereby suppressing the interaction between intimin and translocated intimin receptor. Lypd8 deficiency leads to rapid C. rodentium colonization in the colon, resulting in severe colitis with Th17-cell and neutrophil expansion in the lamina propria. This study identifies a novel function for Lypd8 against A/E bacteria and highlights the role of enterocytes as crucial players in innate immunity for protection against enteric bacterial pathogens.

Comparison of Japanese and Indian intestinal microbiota shows diet-dependent interaction between bacteria and fungi.

The bacterial species living in the gut mediate many aspects of biological processes such as nutrition and activation of adaptive immunity. In addition, commensal fungi residing in the intestine also influence host health. Although the interaction of bacterium and fungus has been shown, its precise mechanism during colonization of the human intestine remains largely unknown. Here, we show interaction between bacterial and fungal species for utilization of dietary components driving their efficient growth in the intestine. Next generation sequencing of fecal samples from Japanese and Indian adults revealed differential patterns of bacterial and fungal composition. In particular, Indians, who consume more plant polysaccharides than Japanese, harbored increased numbers of Prevotella and Candida. Candida spp. showed strong growth responses to the plant polysaccharide arabinoxylan in vitro. Furthermore, the culture supernatants of Candida spp. grown with arabinoxylan promoted rapid proliferation of Prevotella copri. Arabinose was identified as a potential growth-inducing factor in the Candida culture supernatants. Candida spp. exhibited a growth response to xylose, but not to arabinose, whereas P. copri proliferated in response to both xylose and arabinose. Candida spp., but not P. copri, colonized the intestine of germ-free mice. However, P. copri successfully colonized mouse intestine already harboring Candida. These findings demonstrate a proof of concept that fungal members of gut microbiota can facilitate a colonization of the intestine by their bacterial counterparts, potentially mediated by a dietary metabolite.

Author Correction: Diversity in gut bacterial community of school-age children in Asia.

A correction has been published and is appended to both the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Hydrogen-Rich Saline Regulates Intestinal Barrier Dysfunction, Dysbiosis, and Bacterial Translocation in a Murine Model of Sepsis.

Bacterial translocation is a major cause of multiple organ dysfunction syndrome in critical illness, and its management is an important therapeutic strategy. In this study, we focused on the key factors responsible for bacterial translocation including the intestinal microbiome and investigated the impact of molecular hydrogen therapy as a countermeasure against bacterial translocation in a murine model of sepsis. The experimental protocols were divided into the sham, saline treatment (control), and hydrogen treatment (H2) groups. In the H2 group, 15 mL/kg of hydrogen-rich saline (7 ppm) was gavaged daily for 7 days following cecal ligation and puncture (CLP). In the control group, normal saline was gavaged in the same way. In the results, the 7-day survival rate was significantly improved in the H2 group versus the control group (69% vs. 31%, P < 0.05). The incidence of bacterial translocation at 24 h after CLP as assessed by cultivation of mesenteric lymph nodes and blood was significantly decreased in the H2 group versus the control group. Administration of hydrogen-rich saline also prevented the expansion of facultative anaerobic Enterobacteriaceae and ameliorated intestinal hyperpermeability at 24 h after CLP. Intestinal tissue levels of inflammatory mediators such as inducible nitric oxide synthases, tumor necrosis factor α, interleukin (IL)-1ß, IL-6, and oxidative stress marker malondialdehyde at 6 h after CLP were down-regulated in the H2 group. These results suggest luminal administration of hydrogen-rich saline, which prevents intestinal dysbiosis, hyperpermeability, and bacterial translocation, could potentially be a new therapeutic strategy in critical illness.

Slc3a2 Mediates Branched-Chain Amino-Acid-Dependent Maintenance of Regulatory T Cells.

Foxp3+ regulatory T (Treg) cells, which suppress immune responses, are highly proliferative in vivo. However, it remains unclear how the active replication of Treg cells is maintained in vivo. Here, we show that branched-chain amino acids (BCAAs), including isoleucine, are required for maintenance of the proliferative state of Treg cells via the amino acid transporter Slc3a2-dependent metabolic reprogramming. Mice fed BCAA-reduced diets showed decreased numbers of Foxp3+ Treg cells with defective in vivo proliferative capacity. Mice lacking Slc3a2 specifically in Foxp3+ Treg cells showed impaired in vivo replication and decreased numbers of Treg cells. Slc3a2-deficient Treg cells showed impaired isoleucine-induced activation of the mTORC1 pathway and an altered metabolic state. Slc3a2 mutant mice did not show an isoleucine-induced increase of Treg cells in vivo and exhibited multi-organ inflammation. Taken together, these findings demonstrate that BCAA controls Treg cell maintenance via Slc3a2-dependent metabolic regulation.

Evolution of gut Bifidobacterium population in healthy Japanese infants over the first three years of life: a quantitative assessment.

Bifidobacteria are important members of human gut microbiota; however, quantitative data on their early-life dynamics is limited. Here, using a sensitive reverse transcription-qPCR approach, we demonstrate the carriage of eight signature infant-associated Bifidobacterium species (B. longum, B. breve, B. bifidum, B. catenulatum group, B. infantis, B. adolescentis, B. angulatum and B. dentium) in 76 healthy full-term vaginally-born infants from first day to three years of life. About 21% babies carry bifidobacteria at first day of life (6.2 ± 1.9 log10 cells/g feces); and this carriage increases to 64% (8.0 ± 2.2), 79% (8.5 ± 2.1), 97% (9.3 ± 1.8), 99% (9.6 ± 1.6), and 100% (9.7 ± 0.9) at age 7 days, 1, 3 and 6 months, and 3 years, respectively. B. longum, B. breve, B. catenulatum group and B. bifidum are among the earliest and abundant bifidobacterial clades. Interestingly, infants starting formula-feed as early as first week of life have higher bifidobacterial carriage compared to exclusively breast-fed counterparts. Bifidobacteria demonstrate an antagonistic correlation with enterobacteria and enterococci. Further analyses also reveal a relatively lower/ delayed bifidobacterial carriage in cesarean-born babies. The study presents a quantitative perspective of the early-life gut Bifidobacterium colonization and shows how factors such as birth and feeding modes could influence this acquisition even in healthy infants.

Fungal ITS1 Deep-Sequencing Strategies to Reconstruct the Composition of a 26-Species Community and Evaluation of the Gut Mycobiota of Healthy Japanese Individuals.

The study of mycobiota remains relatively unexplored due to the lack of sufficient available reference strains and databases compared to those of bacterial microbiome studies. Deep sequencing of Internal Transcribed Spacer (ITS) regions is the de facto standard for fungal diversity analysis. However, results are often biased because of the wide variety of sequence lengths in the ITS regions and the complexity of high-throughput sequencing (HTS) technologies. In this study, a curated ITS database, ntF-ITS1, was constructed. This database can be utilized for the taxonomic assignment of fungal community members. We evaluated the efficacy of strategies for mycobiome analysis by using this database and characterizing a mock fungal community consisting of 26 species representing 15 genera using ITS1 sequencing with three HTS platforms: Illumina MiSeq (MiSeq), Ion Torrent Personal Genome Machine (IonPGM), and Pacific Biosciences (PacBio). Our evaluation demonstrated that PacBio's circular consensus sequencing with greater than 8 full-passes most accurately reconstructed the composition of the mock community. Using this strategy for deep-sequencing analysis of the gut mycobiota in healthy Japanese individuals revealed two major mycobiota types: a single-species type composed of Candida albicans or Saccharomyces cerevisiae and a multi-species type. In this study, we proposed the best possible processing strategies for the three sequencing platforms, of which, the PacBio platform allowed for the most accurate estimation of the fungal community. The database and methodology described here provide critical tools for the emerging field of mycobiome studies.

Dysbiosis Contributes to Arthritis Development via Activation of Autoreactive T Cells in the Intestine.

OBJECTIVE: The intestinal microbiota is involved in the pathogenesis of arthritis. Altered microbiota composition has been demonstrated in patients with rheumatoid arthritis (RA). However, it remains unclear how dysbiosis contributes to the development of arthritis. The aim of this study was to investigate whether altered composition of human intestinal microbiota in RA patients contributes to the development of arthritis. METHODS: We analyzed the fecal microbiota of patients with early RA and healthy controls, using 16S ribosomal RNA-based deep sequencing. We inoculated fecal samples from RA patients and healthy controls into germ-free arthritis-prone SKG mice and evaluated the immune responses. We also analyzed whether the lymphocytes of SKG mice harboring microbiota from RA patients react with the arthritis-related autoantigen 60S ribosomal protein L23a (RPL23A). RESULTS: A subpopulation of patients with early RA harbored intestinal microbiota dominated by Prevotella copri; SKG mice harboring microbiota from RA patients had an increased number of intestinal Th17 cells and developed severe arthritis when treated with zymosan. Lymphocytes in regional lymph nodes and the colon, but not the spleen, of these mice showed enhanced interleukin-17 (IL-17) responses to RPL23A. Naive SKG mouse T cells cocultured with P copri-stimulated dendritic cells produced IL-17 in response to RPL23A and rapidly induced arthritis. CONCLUSION: We demonstrated that dysbiosis increases sensitivity to arthritis via activation of autoreactive T cells in the intestine. Autoreactive SKG mouse T cells are activated by dysbiotic microbiota in the intestine, causing joint inflammation. Dysbiosis is an environmental factor that triggers arthritis development in genetically susceptible mice.

An Improved Method for High Quality Metagenomics DNA Extraction from Human and Environmental Samples.

To explore the natural microbial community of any ecosystems by high-resolution molecular approaches including next generation sequencing, it is extremely important to develop a sensitive and reproducible DNA extraction method that facilitate isolation of microbial DNA of sufficient purity and quantity from culturable and uncultured microbial species living in that environment. Proper lysis of heterogeneous community microbial cells without damaging their genomes is a major challenge. In this study, we have developed an improved method for extraction of community DNA from different environmental and human origin samples. We introduced a combination of physical, chemical and mechanical lysis methods for proper lysis of microbial inhabitants. The community microbial DNA was precipitated by using salt and organic solvent. Both the quality and quantity of isolated DNA was compared with the existing methodologies and the supremacy of our method was confirmed. Maximum recovery of genomic DNA in the absence of substantial amount of impurities made the method convenient for nucleic acid extraction. The nucleic acids obtained using this method are suitable for different downstream applications. This improved method has been named as the THSTI method to depict the Institute where the method was developed.

Lypd8 promotes the segregation of flagellated microbiota and colonic epithelia.

Colonic epithelial cells are covered by thick inner and outer mucus layers. The inner mucus layer is free of commensal microbiota, which contributes to the maintenance of gut homeostasis. In the small intestine, molecules critical for prevention of bacterial invasion into epithelia such as Paneth-cell-derived anti-microbial peptides and regenerating islet-derived 3 (RegIII) family proteins have been identified. Although there are mucus layers providing physical barriers against the large number of microbiota present in the large intestine, the mechanisms that separate bacteria and colonic epithelia are not fully elucidated. Here we show that Ly6/PLAUR domain containing 8 (Lypd8) protein prevents flagellated microbiota invading the colonic epithelia in mice. Lypd8, selectively expressed in epithelial cells at the uppermost layer of the large intestinal gland, was secreted into the lumen and bound flagellated bacteria including Proteus mirabilis. In the absence of Lypd8, bacteria were present in the inner mucus layer and many flagellated bacteria invaded epithelia. Lypd8(-/-) mice were highly sensitive to intestinal inflammation induced by dextran sulfate sodium (DSS). Antibiotic elimination of Gram-negative flagellated bacteria restored the bacterial-free state of the inner mucus layer and ameliorated DSS-induced intestinal inflammation in Lypd8(-/-) mice. Lypd8 bound to flagella and suppressed motility of flagellated bacteria. Thus, Lypd8 mediates segregation of intestinal bacteria and epithelial cells in the colon to preserve intestinal homeostasis.

Diversity of Intestinal Clostridium coccoides Group in the Japanese Population, as Demonstrated by Reverse Transcription-Quantitative PCR.

We used sensitive rRNA-targeted reverse transcription-quantitative PCR (RT-qPCR) to quantify the Clostridium coccoides group, which is a major anaerobic population in the human intestine. For this purpose, the C. coccoides group was classified into 3 subgroups and 19 species for expediency in accordance with the existing database, and specific primers were newly developed to evaluate them. Population levels of the C. coccoides group in human feces determined by RT-qPCR were equivalent to those determined by fluorescence in situ hybridization. RT-qPCR analysis of fecal samples from 96 volunteers (32 young children, 32 adults and 32 elderly) by using the 22 new primer sets together with the C. coccoides group-specific primer setm revealed that (i) total counts obtained as the sum of the 3 subgroups and 19 species were equivalent to the results obtained by using the C. coccoides group-specific primer set; (ii) total C. coccoides-group counts in the elderly were significantly lower than those in young children and adults; (iii) genus Blautia was the most common subgroup in the human intestinal C. coccoides-group populations at all age populations tested; (iv) the prevalences of Fusicatenibacter saccharivorans and genus Dorea were significantly higher in adults than in young children and the elderly; and (v) the prevalences of C. scindens and C. hylemonae, both of which produce secondary bile acid in the human intestine, were significantly higher in the elderly than in young children and adults. Hierarchical clustering and principal component analysis showed clear separation of the bacterial components between adult and elderly populations. Taken together, these data suggest that aging plays an important role in the diversity of C. coccoides-group populations in human intestinal microbiota; changes in this diversity likely influence the health of the host.