Blautia parvula sp. nov., isolated from Japanese faecal samples.

Two Gram-positive, anaerobic, non-spore-forming and coccoid or oval-shaped bacterial strains, namely, DN0138T and DN0266, were isolated from faecal samples of healthy Japanese people. Phylogenetic analyses based on 16S rRNA gene sequences revealed that strain DN0138T clustered with a species of the genus Blautia and was closely related to Blautia producta JCM 1471T, Blautia coccoides JCM 1395T, Blautia hominis KB1T and 'Blautia marasmi' Marseille-P2377, with sequence similarities of 98.6, 98.5, 98.8 and 98.2 %, respectively. The average nucleotide identity values were 85.3 % for B. producta JCM 1471T, 85.0 % for B. coccoides NCTC 11035T, 84.3 % for B. hominis KB1T and 84.3 % for 'B. marasmi' Marseille-P2377. The major end products of glucose metabolism were acetic acid, lactic acid and succinic acid. The genome length of strain DN0138T was 6 247 046 bp with 46.7 mol% G+C content of genome sequence. Based on their phenotypic, cellular fatty acid and phylogenetic characteristics, the three isolates represent a novel species within the genus Blautia, for which the name Blautia parvula sp. nov. is proposed. The type strain is DN0138T (=NBRC 113351T=BCRC 81349T).

Draft Genome Sequence of the Marine-Derived, Anticancer Compound-Producing Bacterium Streptomyces sp. Strain GMY01.

The marine Streptomyces sp. strain GMY01 was isolated from Indonesian marine sediment. Genome mining analysis revealed that GMY01 has 28 biosynthetic gene clusters, dominated by genes encoding nonribosomal peptide synthetase and polyketide synthase.

Nitrosopumilus zosterae sp. nov., an autotrophic ammonia-oxidizing archaeon of phylum Thaumarchaeota isolated from coastal eelgrass sediments of Japan.

A novel mesophilic and aerobic ammonia-oxidizing archaeon of the phylum Thaumarchaeota, strain NM25T, was isolated from coastal eelgrass zone sediment sampled in Shimoda (Japan). The cells were rod-shaped with an S-layer cell wall. The temperature range for growth was 20-37 °C, with an optimum at 30 °C. The pH range for growth was pH 6.1-7.7, with an optimum at pH 7.1. The salinity range for growth was 5-40 %, with an optimum range of 15-32 %. Cells obtained energy from ammonia oxidation and used bicarbonate as a carbon source. Utilization of urea was not observed for energy generation and growth. Strain NM25T required a hydrogen peroxide scavenger, such as α-ketoglutarate, pyruvate or catalase, for sustained growth on ammonia. Growth of strain NM25T was inhibited by addition of low concentrations of some organic compounds and organic mixtures, including complete inhibition by glycerol, peptone and yeast extract. Phylogenetic analysis of four concatenated housekeeping genes (16S rRNA, rpoB, rpsI and atpD) and concatenated AmoA, AmoB, AmoC amino acid sequences indicated that the isolate is similar to members of the genus Nitrosopumilus. The closest relative is Nitrosopumilus ureiphilus PS0T with sequence similarities of 99.5 % for the 16S rRNA gene and 97.2 % for the amoA gene. Genome relatedness between strain NM25T and N. ureiphilus PS0T was assessed by average nucleotide identity and digital DNA-DNA hybridization, giving results of 85.4 and 40.2 %, respectively. On the basis of phenotypic, genotypic and phylogenetic data, strain NM25T represents a novel species of the genus Nitrosopumilus, for which the name sp. nov, is proposed. The type strain is NM25T (=NBRC 111181T=ATCC TSD-147T).

Validation and standardization of DNA extraction and library construction methods for metagenomics-based human fecal microbiome measurements.

BACKGROUND: Validation and standardization of methodologies for microbial community measurements by high-throughput sequencing are needed to support human microbiome research and its industrialization. This study set out to establish standards-based solutions to improve the accuracy and reproducibility of metagenomics-based microbiome profiling of human fecal samples. RESULTS: In the first phase, we performed a head-to-head comparison of a wide range of protocols for DNA extraction and sequencing library construction using defined mock communities, to identify performant protocols and pinpoint sources of inaccuracy in quantification. In the second phase, we validated performant protocols with respect to their variability of measurement results within a single laboratory (that is, intermediate precision) as well as interlaboratory transferability and reproducibility through an industry-based collaborative study. We further ascertained the performance of our recommended protocols in the context of a community-wide interlaboratory study (that is, the MOSAIC Standards Challenge). Finally, we defined performance metrics to provide best practice guidance for improving measurement consistency across methods and laboratories. CONCLUSIONS: The validated protocols and methodological guidance for DNA extraction and library construction provided in this study expand current best practices for metagenomic analyses of human fecal microbiota. Uptake of our protocols and guidelines will improve the accuracy and comparability of metagenomics-based studies of the human microbiome, thereby facilitating development and commercialization of human microbiome-based products. Video Abstract.

Complete Genome Sequence of Flavonifractor plautii JCM 32125T.

We report the complete genome sequence of Flavonifractor plautii JCM 32125T (=VPI 0310T). The genome consists of a single circular chromosome of 3,985,392 bp (G+C content, 60.9%) and was predicted to contain 3 complete sets of rRNA genes, 63 tRNA genes, and 3,764 protein-coding sequences.

Complete Genome Sequence of Blautia producta JCM 1471T.

We report a complete genome sequence of Blautia producta JCM 1471T The genome consists of a single circular chromosome of 6,197,116 bp with a G+C content of 45.7%. The genome was annotated as containing 5 complete sets of rRNA genes, 70 tRNA genes, and 5,516 protein-coding sequences.

Complete Genome Sequence of Collinsella aerofaciens JCM 10188T.

We report a complete genome sequence of Collinsella aerofaciens JCM 10188T (=VPI 1003T). The genome consists of a circular chromosome (2,428,218 bp with 60.6% G+C content) and two extrachromosomal elements. The genome was predicted to contain 5 sets of rRNA genes, 58 tRNA genes, and 2,079 protein-encoding sequences.

Complete Genome Sequence of Megamonas funiformis JCM 14723T.

We announce the complete genome sequence of Megamonas funiformis JCM 14723T (YIT 11815T). The genome consists of a circular chromosome (2,522,577 bp, 31.5% G+C content) and a plasmid of 46,189 bp (29.4% G+C content). The genome was predicted to contain 6 rRNA operons, 53 tRNA genes, and 2,440 protein-coding sequences.

Complete genome sequence of natural rubber-degrading, gram-negative bacterium, Rhizobacter gummiphilus strain NS21T.

Gram-negative natural rubber-degrader, Rhizobacter gummiphilus NS21T, which was isolated from soil in the botanical garden in Japan, is a newly proposed species of genus of Rhizobacter. It has been reported that the latA1 gene is involved in the natural rubber degradation in this strain. To gain novel insights into natural rubber degradation pathway, the complete genome sequence of this strain was determined. The genome of strain NS21T consists of 6,398,096 bp of circular chromosome (GenBank accession number CP015118.1) with G + C content of 69.72%. The genome contains 5687 protein-coding and 68 RNA genes. Among the predicted genes, 4810 genes were categorized as functional COGs. Homology search revealed that existence of latA1 homologous gene (latA2) in this genome. Quantitative reverse-transcription-PCR and deletion analyses indicated that natural rubber degradation of this strain requires latA2 as well as latA1.

Ecological impact assessment of a bioaugmentation site on remediation of chlorinated ethylenes by multi-omics analysis.

Bioremediation may affect the ecological system around bioremediation sites. However, little is known about how microbial community structures change over time after the initial injection of degraders. In this study, we have assessed the ecological impact of bioaugmentation using metagenomic and metatranscriptomic approaches to remove trichlorinated ethylene/cis-dichloroethylene (TCE/cDCE) by Rhodococcus jostii strain RHA1 as an aerobic chemical compound degrader. Metagenomic analysis showed that the number of organisms belonging to the genus Rhodococcus, including strain RHA1, increased from 0.1% to 76.6% of the total microbial community on day 0 at the injection site. Subsequently, the populations of strain RHA1 and other TCE/cDCE-degrading bacteria gradually decreased over time, whereas the populations of the anaerobic dechlorinators Geobacter and Dehalococcoides increased at later stages. Metatranscriptomic analysis revealed a high expression of aromatic compound-degrading genes (bphA1-A4) in strain RHA1 after RHA1 injection. From these results, we concluded that the key dechlorinators of TCE/cDCE were mainly aerobic bacteria, such as RHA1, until day 1, after which the key dechlorinators changed to anaerobic bacteria, such as Geobacter and Dehalococcocides, after day 6 at the injection well. Based on the α-diversity, the richness levels of the microbial community were increased after injection of strain RHA1, and the microbial community composition had not been restored to that of the original composition during the 19 days after treatment. These results provide insights into the assessment of the ecological impact and bioaugmentation process of RHA1 at bioremediation sites.

Reclassification of Nocardia species based on whole genome sequence and associated phenotypic data.

Type strains of 72 validated Nocardia species were phylogenetically analyzed based on the multilocus sequence analysis (MLSA) concatenated atpD-groL1-groL2-recA-rpoA-secY-sodA-ychF. Furthermore, their similarity based on digital DNA-DNA hybridization (dDDH) was calculated. Nocardia soli, Nocardia cummidelens and Nocardia salmonicida, Nocardia nova and Nocardia elegans, Nocardia exalbida and Nocardia gamkensis, and Nocardia coubleae and Nocardia ignorata formed coherent clades, respectively. Moreover, each set showed over 70% relatedness by dDDH and shared common phenotypic characteristics. Therefore, we propose a reclassification of Nocardia soli and Nocardia cummidelens as a later heterotypic synonym of Nocardia salmonicida, Nocardia elegans as a later heterotypic synonym of Nocardia nova, Nocardia gamkensis as a later heterotypic synonym of Nocardia exalbida, and Nocardia coubleae as a later heterotypic synonym of Nocardia ignorata.

Proteobacteria and Bacteroidetes are major phyla of filterable bacteria passing through 0.22 µm pore size membrane filter, in Lake Sanaru, Hamamatsu, Japan.

141 filterable bacteria that passed through a 0.22 µm pore size filter were isolated from Lake Sanaru in Hamamatsu, Japan. These belonged to Proteobacteria, Bacteroidetes, Firmicutes, or Actinobacteria among which the first two phyla comprised the majority of the isolates. 48 isolates (12 taxa) are candidates assignable to new bacterial species or genera of Proteobacteria or Bacteroidetes.

Complete Genome Sequence of the Polychlorinated Biphenyl-Degrading Bacterium Pseudomonas putida KF715 (NBRC 110667) Isolated from Biphenyl-Contaminated Soil.

Pseudomonas putida KF715 (NBRC 110667) utilizes biphenyl as a sole source of carbon and degrades polychlorinated biphenyls (PCBs). Here, we report a complete genome sequence of the KF715 strain, which comprises a circular chromosome and four plasmids. Biphenyl catabolic genes were located on the largest plasmid, pKF715A.

Phylogenetic analysis reveals the taxonomically diverse distribution of the Pseudomonas putida group.

Pseudomonas putida is well-known for degradation activities for a variety of compounds and its infections have been reported. Thus, P. putida includes both clinical and nonclinical isolates. To date, no reports have examined the phylogenetic relationship between clinical and nonclinical isolates of the P. putida group. In this study, fifty-nine strains of P. putida group containing twenty-six clinical, and thirty-three nonclinical, isolates, were subjected to phylogenetic and taxonomic analyses based on 16S rRNA gene sequences and nine housekeeping gene sequences, including argS, dnaN, dnaQ, era, gltA, gyrB, ppnK, rpoB, and rpoD, to obtain insights into the diversity of species in this group. More than 97.6% similarity was observed among the 16S rRNA gene sequences of all the strains examined, indicating that the resolution of 16S rRNA gene sequences is inadequate. Phylogenetic analysis based on the individual housekeeping genes listed above improved the resolution of the phylogenetic trees, which are different from each other. Multilocus sequence analysis (MLSA) based on the concatenated sequences of the nine genes significantly improved the resolution of the phylogenetic tree, and yielded approximately the same results as average nucleotide identity (ANI) analysis, suggesting its high reliability. ANI analysis classified the fifty-nine strains into twenty-six species containing seventeen singletons and nine strain clusters based on the 95% threshold. It also indicated the mixed distribution of clinical and nonclinical isolates in the six clusters, suggesting that the genomic difference between clinical and nonclinical isolates of the P. putida group is subtle. The P. putida type strain NBRC 14164T is a singleton that is independently located from the P. putida strains distributed among the six clusters, suggesting that the classification of these strains and the differentiation of species in the P. putida group should be re-examined. This study greatly expands insights into the phylogenetic diversity of the P. putida group.

Biodegradation of high concentrations of mixed polycyclic aromatic hydrocarbons by indigenous bacteria from a river sediment: a microcosm study and bacterial community analysis.

This study assessed the biodegradation of mixtures of polycyclic aromatic hydrocarbons (PAHs) by indigenous bacteria in river sediment. Microcosms were constructed from sediment from the Chao Phraya River (the main river in Thailand) by supplementation with high concentrations of fluorene, phenanthrene, pyrene (300 mg kg-1 of each PAH), and acenaphthene (600 mg kg-1). Fluorene and phenanthrene were completely degraded, whereas 50% of the pyrene and acenaphthene were removed at the end of the incubation period (70 days). Community analyses revealed the dynamics of the bacterial profiles in the PAH-degrading microcosms after PAH exposure. Actinobacteria predominated and became significantly more abundant in the microcosms after 14 days of incubation at room temperature under aerobic conditions. Furthermore, the remaining PAHs and alpha diversity were positively correlated. The sequencing of clone libraries of the PAH-RHDα genes also revealed that the dioxygenase genes of Mycobacterium sp. comprised 100% of the PAH-RHDα library at the end of the microcosm setup. Moreover, two PAH-degrading Actinobacteria (Arthrobacter sp. and Rhodococcus ruber) were isolated from the original sediment sample and showed high activity in the degradation of phenanthrene and fluorene in liquid cultivation. This study reveals that indigenous bacteria had the ability to degrade high concentrations of mixed PAHs and provide clear evidence that Actinobacteria may be potential candidates to play a major role in PAH degradation in the river sediment.

Culture-independent study of bacterial communities in tropical river sediment.

Ubiquitous microbial communities in river sediments actively govern organic matter decomposition, nutrient recycling, and remediation of toxic compounds. In this study, prokaryotic diversity in two major rivers in central Thailand, the Chao Phraya (CP) and the Tha Chin (TC) distributary was investigated. Significant differences in sediment physicochemical properties, particularly silt content, were noted between the two rivers. Tagged 16S rRNA sequencing on a 454 platform showed that the sediment microbiomes were dominated by Gammaproteobacteria and sulfur/sulfate reducing Deltaproteobacteria, represented by orders Desulfobacteriales and Desulfluromonadales together with organic degraders Betaproteobacteria (orders Burkholderiales and Rhodocyclales) together with the co-existence of Bacteroidetes predominated by Sphingobacteriales. Enrichment of specific bacterial orders was found in the clayey CP and silt-rich TC sediments, including various genera with known metabolic capability on decomposition of organic matter and xenobiotic compounds. The data represent one of the pioneered works revealing heterogeneity of bacteria in river sediments in the tropics.

Complete Genome Sequence of Sphingopyxis macrogoltabida Strain 203N (NBRC 111659), a Polyethylene Glycol Degrader.

We determined the complete genome sequence of Sphingopyxis macrogoltabida strain 203N, a polyethylene glycol degrader. Because the PacBio assembly (285× coverage) seemed to be full of nucleotide-level mismatches, the Newbler assembly of MiSeq mate-pair and paired-end data was used for finishing and the PacBio assembly was used as a reference. The PacBio assembly carried 414 nucleotide mismatches over 5,953,153 bases of the 203N genome.

Complete Genome Sequence of Sphingopyxis terrae Strain 203-1 (NBRC 111660), a Polyethylene Glycol Degrader.

The complete genome sequence of Sphingopyxis terrae strain 203-1, which is capable of growing on polyethylene glycol, was determined. The genome consisted of a chromosome with a size of 3.98 Mb and a plasmid with a size of 4,328 bp. The strain was deposited to the National Institute of Technology and Evaluation (Tokyo, Japan) under the number NBRC 111660.

Draft Genome Sequence of Comamonas thiooxydans Strain PHE2-6 (NBRC 110656), a Chlorinated-Ethene-Degrading Bacterium.

Comamonas thiooxydans strain PHE2-6 (NBRC 110656), which was isolated from a trichloroethene-contaminated site in Japan, utilizes phenol as a sole source of carbon and cometabolizes cis- and trans-dichloroethenes. We report here the draft genome sequence of this strain, containing 5,309,680 bp, with 60.6% G+C content.

Isolation and characterization of a bacterial strain that degrades cis-dichloroethenein the absence of aromatic inducers.

Bacteria capable of degrading cis-dichloroethene (cDCE) were screened from cDCE-contaminated soil, and YKD221, a bacterial strain that exhibited a higher growth on minimal salt agar plates in the presence of cDCE than in the absence of cDCE, were isolated. Phylogenetic studies of the 16S rRNA as well as gyrB, rpoD, and recA in YKD221 indicated that this strain is closely related to the type strains of Pseudomonas plecoglossicida, monteilii, and putida. An average nucleotide identity analysis indicated that YKD221 is most closely related to P. putida strains, including the type strain, which suggests that YKD221 belongs to P. putida. Although the genome of YKD221 was very similar to that of P. putida F1, a toluene-degrading strain, the YKD221 genome has 15 single-nucleotide polymorphisms and 4 insertions compared with the F1 genome. YKD221 caused the release of sufficient chloride ions from cDCE to suggest that the strain is able to completely dechlorinate and degrade cDCE. YKD221 also degraded trichloroethene but was unable to degrade trans-dichloroethene and tetrachloroethene. The degradation activity of YKD221 was elevated after growth on toluene. Inactivation of todC1, which encodes for a large subunit of the catalytic terminal component in toluene dioxygenase, resulted in a complete loss of growth on toluene and cDCE degradation activity. This is the first evidence of the involvement of todC1C2BA-coded toluene dioxygenase in cDCE degradation. YKD221 did not appear to grow on cDCE in a minimal salt liquid medium. However, YKD221 did exhibit an enhanced increase in cell concentration and volume of cells during growth on minimal salt agar plates with cDCE when first grown in LB medium. This behavior appears to have led us to misinterpret our initial results on YKD221 as an indication of improved growth in the presence of cDCE.