Dimension reduction of microbiome data linked Bifidobacterium and Prevotella to allergic rhinitis.

Dimension reduction has been used to visualise the distribution of multidimensional microbiome data, but the composite variables calculated by the dimension reduction methods have not been widely used to investigate the relationship of the human gut microbiome with lifestyle and disease. In the present study, we applied several dimension reduction methods, including principal component analysis, principal coordinate analysis (PCoA), non-metric multidimensional scaling (NMDS), and non-negative matrix factorization, to a microbiome dataset from 186 subjects with symptoms of  allergic rhinitis (AR) and 106 controls. All the dimension reduction methods supported that the distribution of microbial data points appeared to be continuous rather than discrete. Comparison of the composite variables calculated from the different dimension reduction methods showed that the characteristics of the composite variables differed depending on the distance matrices and the dimension reduction methods. The first composite variables calculated from PCoA and NMDS with the UniFrac distance were strongly associated with AR (FDR adjusted P = 2.4 × 10-4 for PCoA and P = 2.8 × 10-4 for NMDS), and also with the relative abundance of Bifidobacterium and Prevotella. The abundance of Bifidobacterium was also linked to intake of several nutrients, including carbohydrate, saturated fat, and alcohol via composite variables. Notably, the association between the composite variables and AR was much stronger than the association between the relative abundance of individual genera and AR. Our results highlight the usefulness of the dimension reduction methods for investigating the association of microbial composition with lifestyle and disease in clinical research.

Asymptomatic Enteric Virus Infections and Association with the Gut Microbiome in Rural Residents of Northern Laos.

Viral gastrointestinal infections are an important public health concern, and the occurrence of asymptomatic enteric virus infections makes it difficult to prevent and control their spread. This study aimed to determine the prevalence of and factors associated with asymptomatic enteric virus infection in adults in northern Laos. Fecal samples were collected from apparently healthy participants who did not report diarrhea or high fever at the time of the survey in northern Laos, and enteric viruses were detected using polymerase chain reaction (PCR) and reverse transcription (RT)-PCR. Individual characteristics, including the gut microbiome, were compared between asymptomatic carriers and noncarriers of each enteric virus. Of the participants (N = 255), 12 (4.7%) were positive for norovirus genogroup I (GI), 8 (3.1%) for human adenovirus, and 1 (0.4%) for norovirus GII; prevalence tended to be higher in less-modernized villages. Gut microbial diversity (evaluated by the number of operational taxonomic units) was higher in asymptomatic carriers of norovirus GI or human adenovirus than in their noncarriers. Gut microbiome compositions differed significantly between asymptomatic carriers and noncarriers of norovirus GI or human adenovirus (permutational analysis of variance, P <0.05). These findings imply an association between asymptomatic enteric virus infection and modernization and/or the gut microbiome in northern Laos.

Metagenomic gut microbiome analysis of Japanese patients with multiple chemical sensitivity/idiopathic environmental intolerance.

BACKGROUND: Although the pathology of multiple chemical sensitivity (MCS) is unknown, the central nervous system is reportedly involved. The gut microbiota is important in modifying central nervous system diseases. However, the relationship　between the gut microbiota and MCS remains unclear. This study aimed to identify gut microbiota variations associated with MCS using shotgun metagenomic sequencing of fecal samples. METHODS: We prospectively recruited 30 consecutive Japanese female patients with MCS and analyzed their gut microbiomes using shotgun metagenomic sequencing. The data were compared with metagenomic data obtained from 24 age- and sex-matched Japanese healthy controls (HC). RESULTS: We observed no significant difference in alpha and beta diversity of the gut microbiota between the MCS patients and HC. Focusing on the important changes in the literatures, at the genus level, Streptococcus, Veillonella, and Akkermansia were significantly more abundant in MCS patients than in HC (p < 0.01, p < 0.01, p = 0.01, respectively, fold change = 4.03, 1.53, 2.86, respectively). At the species level, Akkermansia muciniphila was significantly more abundant (p = 0.02, fold change = 3.3) and Faecalibacterium prausnitzii significantly less abundant in MCS patients than in HC (p = 0.03, fold change = 0.53). Functional analysis revealed that xylene and dioxin degradation pathways were significantly enriched (p < 0.01, p = 0.01, respectively, fold change = 1.54, 1.46, respectively), whereas pathways involved in amino acid metabolism and synthesis were significantly depleted in MCS (p < 0.01, fold change = 0.96). Pathways related to antimicrobial resistance, including the two-component system and cationic antimicrobial peptide resistance, were also significantly enriched in MCS (p < 0.01, p < 0.01, respectively, fold change = 1.1, 1.2, respectively). CONCLUSIONS: The gut microbiota of patients with MCS shows dysbiosis and alterations in bacterial functions related to exogenous chemicals and amino acid metabolism and synthesis. These findings may contribute to the further development of treatment for MCS. TRIAL REGISTRATION: This study was registered with the University Hospital Medical Information Clinical Trials Registry as UMIN000031031. The date of first trial registration: 28/01/2018.

Uncovering microbiomes of the rice phyllosphere using long-read metagenomic sequencing.

The plant microbiome is crucial for plant growth, yet many important questions remain, such as the identification of specific bacterial species in plants, their genetic content, and location of these genes on chromosomes or plasmids. To gain insights into the genetic makeup of the rice-phyllosphere, we perform a metagenomic analysis using long-read sequences. Here, 1.8 Gb reads are assembled into 26,067 contigs including 142 circular sequences. Within these contigs, 669 complete 16S rRNA genes are clustered into 166 bacterial species, 121 of which show low identity (<97%) to defined sequences, suggesting novel species. The circular contigs contain novel chromosomes and a megaplasmid, and most of the smaller circular contigs are defined as novel plasmids or bacteriophages. One circular contig represents the complete chromosome of a difficult-to-culture bacterium Candidatus Saccharibacteria. Our findings demonstrate the efficacy of long-read-based metagenomics for profiling microbial communities and discovering novel sequences in plant-microbiome studies.

Alteration of shoaling behavior and dysbiosis in the gut of medaka (Oryzias latipes) exposed to 2-µm polystyrene microplastics.

There is global concern that microplastics may harm aquatic life. Here, we examined the effects of fine polystyrene microplastics (PS-MPs, 2-µm diameter, 0.1 mg/L, 2.5 × 107 particles/L) on the behavior and the microbiome (linked to brain-gut interaction) of a fish model using medaka, Oryzias latipes. We found that shoaling behavior was reduced in PS-MP-exposed medaka compared with control fish during the exposure period, but it recovered during a depuration period. There was no difference in swimming speed between the PS-MP-exposed and control groups during the exposure period. Analysis of the dominant bacterial population (those comprising ≥1% of the total bacterial population) in the gut of fish showed that exposure to PS-MPs tended to increase the relative abundance of the phylum Fusobacteria and the genus Vibrio. Furthermore, structural-equation modeling of gut bacteria on the basis of machine-learning data estimated strong relationship involved in the reduction of the functional bacterial species of minority (<1% of the total bacterial population) such as the genera Muribaculum (an undefined role), Aquaspirillum (a candidate for nitrate metabolism and magnetotactics), and Clostridium and Phascolarctobacterium (potential producers of short-chain fatty acids, influencing behavior by affecting levels of neurotransmitters) as a group of gut bacteria in association with PS-MP exposure. Our results suggest that fish exposure to fine microplastics may cause dysbiosis and ultimately cause social behavior disorders linked to brain-gut interactions. This effect could be connected to reduction of fish fitness in the ecosystem and reduced fish survival.

Gut microbiota and fecal metabolites in sustained unresponsiveness by oral immunotherapy in school-age children with cow's milk allergy.

BACKGROUND: Oral immunotherapy (OIT) can ameliorate cow's milk allergy (CMA); however, the achievement of sustained unresponsiveness (SU) is challenging. Regarding the pathogenesis of CMA, recent studies have shown the importance of gut microbiota (Mb) and fecal water-soluble metabolites (WSMs), which prompted us to determine the change in clinical and gut environmental factors important for acquiring SU after OIT for CMA. METHODS: We conducted an ancillary cohort study of a multicenter randomized, parallel-group, delayed-start design study on 32 school-age children with IgE-mediated CMA who underwent OIT for 13 months. We defined SU as the ability to consume cow's milk exceeding the target dose in a double-blind placebo-controlled food challenge after OIT followed by a 2-week-avoidance. We longitudinally collected 175 fecal specimens and clustered the microbiome and metabolome data into 29 Mb- and 12 WSM-modules. RESULTS: During OIT, immunological factors improved in all participants. However, of the 32 participants, 4 withdrew because of adverse events, and only 7 were judged SU. Gut environmental factors shifted during OIT, but only in the beginning, and returned to the baseline at the end. Of these factors, milk- and casein-specific IgE and the Bifidobacterium-dominant module were associated with SU (milk- and casein-specific IgE; OR for 10 kUA/L increments, 0.67 and 0.66; 95%CI, 0.41-0.93 and 0.42-0.90; Bifidobacterium-dominant module; OR for 0.01 increments, 1.40; 95%CI, 1.10-2.03), and these associations were observed until the end of OIT. CONCLUSIONS: In this study, we identified the clinical and gut environmental factors associated with SU acquisition in CM-OIT.

Complete genome sequences of Bifidobacterium faecale strain JCM19861T, isolated from human feces.

Here, we report the complete genome sequence of the Bifidobacterium faecale strain JCM 19861T (= CU 3-7T = KACC 17904T), isolated from infant feces by Jung-Hye Choi's group in 2014. The B. faecale JCM 19861T genome comprised a circular chromosome of 2,213,206 bp, with a G + C content of 59.0%.

Recovery of microbial DNA by agar-containing solution from extremely low-biomass specimens including skin.

Recovering a sufficient amount of microbial DNA from extremely low-biomass specimens, such as human skin, to investigate the community structure of the microbiome remains challenging. We developed a sampling solution containing agar to increase the abundance of recovered microbial DNA. Quantitative PCR targeting the 16S rRNA gene revealed a significant increase in the amount of microbial DNA recovered from the developed sampling solution compared with conventional solutions from extremely low-biomass skin sites such as the volar forearm and antecubital fossa. In addition, we confirmed that the developed sampling solution reduces the contamination rate of probable non-skin microbes compared to the conventional solutions, indicating that the enhanced recovery of microbial DNA was accompanied by a reduced relative abundance of contaminating microbes in the 16S rRNA gene amplicon sequencing data. In addition, agar was added to each step of the DNA extraction process, which improved the DNA extraction efficiency as a co-precipitant. Enzymatic lysis with agar yielded more microbial DNA than conventional kits, indicating that this method is effective for analyzing microbiomes of low-biomass specimens.

Rationally-defined microbial consortia suppress multidrug-resistant proinflammatory Enterobacteriaceae via ecological control.

Persistent colonization and outgrowth of pathogenic organisms in the intestine may occur due to long-term antibiotic usage or inflammatory conditions, which perpetuate dysregulated immunity and tissue damage1,2. Gram-negative Enterobacteriaceae gut pathobionts are particularly recalcitrant to conventional antibiotic treatment3,4, though an emerging body of evidence suggests that manipulation of the commensal microbiota may be a practical alternative therapeutic strategy5-7. In this study, we rationally isolated and down-selected commensal bacterial consortia from healthy human stool samples capable of strongly and specifically suppressing intestinal Enterobacteriaceae. One of the elaborated consortia, consisting of 18 commensal strains, effectively controlled ecological niches by regulating gluconate availability, thereby reestablishing colonization resistance and alleviating antibiotic-resistant Klebsiella-driven intestinal inflammation in mice. Harnessing these microbial activities in the form of live bacterial therapeutics may represent a promising solution to combat the growing threat of proinflammatory, antimicrobial-resistant bacterial infection.

Gut insulin action protects from hepatocarcinogenesis in diabetic mice comorbid with nonalcoholic steatohepatitis.

Diabetes is known to increase the risk of nonalcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC). Here we treat male STAM (STelic Animal Model) mice, which develop diabetes, NASH and HCC associated with dysbiosis upon low-dose streptozotocin and high-fat diet (HFD), with insulin or phlorizin. Although both treatments ameliorate hyperglycemia and NASH, insulin treatment alone lead to suppression of HCC accompanied by improvement of dysbiosis and restoration of antimicrobial peptide production. There are some similarities in changes of microflora from insulin-treated patients comorbid with diabetes and NASH. Insulin treatment, however, fails to suppress HCC in the male STAM mice lacking insulin receptor specifically in intestinal epithelial cells (ieIRKO), which show dysbiosis and impaired gut barrier function. Furthermore, male ieIRKO mice are prone to develop HCC merely on HFD. These data suggest that impaired gut insulin signaling increases the risk of HCC, which can be countered by restoration of insulin action in diabetes.

Strain-level detection of Fusobacterium nucleatum in colorectal cancer specimens by targeting the CRISPR-Cas region.

IMPORTANCE: Fusobacterium nucleatum is one of the predominant oral bacteria in humans. However, this bacterium is enriched in colorectal cancer (CRC) tissues and may be involved in CRC development. Our previous research suggested that F. nucleatum is present in CRC tissues originating from the oral cavity using a traditional strain-typing method [arbitrarily primed polymerase chain reaction (AP-PCR)]. First, using whole-genome sequencing, this study confirmed an exemplary similarity between the oral and tumoral strains derived from each patient with CRC. Second, we successfully developed a method to genotype this bacterium at the strain level, targeting the clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated system, which is hypervariable (defined as F. nucleatum-strain genotyping PCR). This method can identify F. nucleatum strains in cryopreserved samples and is significantly superior to traditional AP-PCR, which can only be performed on isolates. The new methods have great potential for application in etiological studies of F. nucleatum in CRC.

Tomato root-associated Sphingobium harbors genes for catabolizing toxic steroidal glycoalkaloids.

IMPORTANCE: Saponins are a group of plant specialized metabolites with various bioactive properties, both for human health and soil microorganisms. Our previous works demonstrated that Sphingobium is enriched in both soils treated with a steroid-type saponin, such as tomatine, and in the tomato rhizosphere. Despite the importance of saponins in plant-microbe interactions in the rhizosphere, the genes involved in the catabolism of saponins and their aglycones (sapogenins) remain largely unknown. Here we identified several enzymes that catalyzed the degradation of steroid-type saponins in a Sphingobium isolate from tomato roots, RC1. A comparative genomic analysis of Sphingobium revealed the limited distribution of genes for saponin degradation in our saponin-degrading isolates and several other isolates, suggesting the possible involvement of the saponin degradation pathway in the root colonization of Sphingobium spp. The genes that participate in the catabolism of sapogenins could be applied to the development of new industrially valuable sapogenin molecules.

Gut microbial carbohydrate metabolism contributes to insulin resistance.

Insulin resistance is the primary pathophysiology underlying metabolic syndrome and type 2 diabetes1,2. Previous metagenomic studies have described the characteristics of gut microbiota and their roles in metabolizing major nutrients in insulin resistance3-9. In particular, carbohydrate metabolism of commensals has been proposed to contribute up to 10% of the host's overall energy extraction10, thereby playing a role in the pathogenesis of obesity and prediabetes3,4,6. Nevertheless, the underlying mechanism remains unclear. Here we investigate this relationship using a comprehensive multi-omics strategy in humans. We combine unbiased faecal metabolomics with metagenomics, host metabolomics and transcriptomics data to profile the involvement of the microbiome in insulin resistance. These data reveal that faecal carbohydrates, particularly host-accessible monosaccharides, are increased in individuals with insulin resistance and are associated with microbial carbohydrate metabolisms and host inflammatory cytokines. We identify gut bacteria associated with insulin resistance and insulin sensitivity that show a distinct pattern of carbohydrate metabolism, and demonstrate that insulin-sensitivity-associated bacteria ameliorate host phenotypes of insulin resistance in a mouse model. Our study, which provides a comprehensive view of the host-microorganism relationships in insulin resistance, reveals the impact of carbohydrate metabolism by microbiota, suggesting a potential therapeutic target for ameliorating insulin resistance.

Bacterial induction of B cell senescence promotes age-related changes in the gut microbiota.

The elucidation of the mechanisms of ageing and the identification of methods to control it have long been anticipated. Recently, two factors associated with ageing-the accumulation of senescent cells and the change in the composition of gut microbiota-have been shown to play key roles in ageing. However, little is known about how these phenomena occur and are related during ageing. Here we show that the persistent presence of commensal bacteria gradually induces cellular senescence in gut germinal centre B cells. Importantly, this reduces both the production and diversity of immunoglobulin A (IgA) antibodies that target gut bacteria, thereby changing the composition of gut microbiota in aged mice. These results have revealed the existence of IgA-mediated crosstalk between the gut microbiota and cellular senescence and thus extend our understanding of the mechanism of gut microbiota changes with age, opening up possibilities for their control.

The propionate-GPR41 axis in infancy protects from subsequent bronchial asthma onset.

Evidence has accumulated that gut microbiota and its metabolites, in particular the short-chain fatty acid propionate, are significant contributors to the pathogenesis of a variety of diseases. However, little is known regarding its impact on pediatric bronchial asthma, one of the most common allergic diseases in childhood. This study aimed to elucidate whether, and if so how, intestinal propionate during lactation is involved in the development of bronchial asthma. We found that propionate intake through breast milk during the lactation period resulted in a significant reduction of airway inflammation in the offspring in a murine house dust mite-induced asthma model. Moreover, GPR41 was the propionate receptor involved in suppressing this asthmatic phenotype, likely through the upregulation of Toll-like receptors. In translational studies in a human birth cohort, we found that fecal propionate was decreased one month after birth in the group that later developed bronchial asthma. These findings indicate an important role for propionate in regulating immune function to prevent the pathogenesis of bronchial asthma in childhood.

A comprehensive microbial analysis of pediatric patients with acute appendicitis.

BACKGROUND: Pathogenesis of pediatric acute appendicitis (AA) is yet to be elucidated. Therefore, we performed a comprehensive microbial analysis of saliva, feces, and appendiceal lumen of AA patients using 16S ribosomal RNA (rRNA) gene amplicon sequencing to elucidate the pathogenesis of pediatric AA. METHODS: This study included 33 AA patients and 17 healthy controls (HCs) aged <15 y. Among the AA patients, 18 had simple appendicitis, and 15 had complicated appendicitis. Salivary and fecal samples were obtained from both groups. The contents of the appendiceal lumen were collected from the AA group. All samples were analyzed using 16S rRNA gene amplicon sequencing. RESULTS: The relative abundance of Fusobacterium was significantly higher in the saliva of AA patients as compared to that in HCs (P = 0.011). Bacteroides, Escherichia, Fusobacterium, Coprobacillus, and Flavonifractor were significantly increased in the feces of AA patients, as compared to that in HCs (P = 0.020, 0.010, 0.029, 0.031, and 0.002, respectively). In the appendiceal lumen, Bacteroides, Parvimonas, Fusobacterium, and Alloprevotella were the top bacterial genera with an average relative abundance >5% (16.0%, 9.1%, 7.9%, and 6.0%, respectively). CONCLUSIONS: The relative abundance of Fusobacterium was high in the appendiceal lumen of pediatric AA patients. Moreover, the relative abundance of Fusobacterium was significantly higher in the saliva and feces of pediatric AA patients than in those of healthy children. These results suggest that ectopic colonization of oral Fusobacterium in the appendix might play an important role in the pathogenesis of pediatric AA.

Complete Genome Sequences of Polynucleobacter sp. Subcluster PnecA Strains SHI2 and SHI8, Isolated from an Oligotrophic-Dystrophic Lake in Japan.

Members of the genus Polynucleobacter belonging to the subcluster PnecA comprise freshwater bacterioplankton with worldwide distribution. Here, we report the complete genome sequences of two Polynucleobacter sp. strains (PnecA), SHI2 and SHI8, isolated from the surface water of an oligotrophic-dystrophic lake in a humid continental climate in Japan.

An agroecological structure model of compost-soil-plant interactions for sustainable organic farming.

Compost is used worldwide as a soil conditioner for crops, but its functions have still been explored. Here, the omics profiles of carrots were investigated, as a root vegetable plant model, in a field amended with compost fermented with thermophilic Bacillaceae for growth and quality indices. Exposure to compost significantly increased the productivity, antioxidant activity, color, and taste of the carrot root and altered the soil bacterial composition with the levels of characteristic metabolites of the leaf, root, and soil. Based on the data, structural equation modeling (SEM) estimated that amino acids, antioxidant activity, flavonoids and/or carotenoids in plants were optimally linked by exposure to compost. The SEM of the soil estimated that the genus Paenibacillus and nitrogen compounds were optimally involved during exposure. These estimates did not show a contradiction between the whole genomic analysis of compost-derived Paenibacillus isolates and the bioactivity data, inferring the presence of a complex cascade of plant growth-promoting effects and modulation of the nitrogen cycle by the compost itself. These observations have provided information on the qualitative indicators of compost in complex soil-plant interactions and offer a new perspective for chemically independent sustainable agriculture through the efficient use of natural nitrogen.

Complete Genome Sequences of Three Polynucleobacter sp. Subcluster PnecC Strains, KF022, KF023, and KF032, Isolated from a Shallow Eutrophic Lake and a River in Japan.

The genus Polynucleobacter subcluster PnecC consists of bacteria representing the ubiquitous taxon of freshwater bacterioplankton. Here, we report the complete genome sequences of three Polynucleobacter sp. (PnecC) strains, namely, KF022, KF023, and KF032, which were isolated from surface water of a temperate shallow eutrophic lake and its inflow river in Japan.

Complete Genome Sequences of Three Limnohabitans sp. (Lhab-A3) Strains, INBF002, TEGF004, and MORI2, Isolated from Two Lakes and a River in Japan.

Freshwater bacterioplankton of the genus Limnohabitans represent a dominant group that has worldwide distribution. Here, we report the complete genome sequences of three Limnohabitans sp. (Lhab-A3 tribe) strains, i.e., INBF002, TEGF004, and MORI2, which were isolated from surface water samples from two shallow eutrophic lakes and a river in Japan.