Journal of Microbiology

Volume 56(10); October 2018

Review

[MINIREVIEW] Progress of analytical tools and techniques for human gut microbiome research: Eun-Ji Song , Eun-Sook Lee , Young-Do Nam; J. Microbiol. 2018;56(10):693-705. Published online September 28, 2018; DOI: https://doi.org/10.1007/s12275-018-8238-5

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Massive DNA sequencing studies have expanded our insights and understanding of the ecological and functional characteristics of the gut microbiome. Advanced sequencing technologies allow us to understand the close association of the gut microbiome with human health and critical illnesses. In the future, analyses of the gut microbiome will provide key information associating with human individual health, which will help provide personalized health care for diseases. Numerous molecular biological analysis tools have been rapidly developed and employed for the gut microbiome researches; however, methodological differences among researchers lead to inconsistent data, limiting extensive share of data. It is therefore very essential to standardize the current
method
ologies and establish appropriate pipelines for human gut microbiome research. Herein, we review the methods and procedures currently available for studying the human gut microbiome, including fecal sample collection, metagenomic DNA extraction, massive DNA sequencing, and data analyses with bioinformatics. We believe that this review will contribute to the progress of gut microbiome research in the clinical and practical aspects of human health.

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Journal Articles

Brevibacterium anseongense sp. nov., isolated from soil of ginseng field: Mi-Seon Jung , Xiao-Tian Quan , Muhammad Zubair Siddiqi , Qingzhen Liu , Sang Yong Kim , Ji-Hyang Wee , Wan Taek Im; J. Microbiol. 2018;56(10):706-712. Published online August 22, 2018; DOI: https://doi.org/10.1007/s12275-018-8181-5

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Abstract

A Gram-positive, aerobic, non-motile, pale-yellow, and rodshaped bacterium, designated as Gsoil 188T, was isolated from the soil of a ginseng field in Pocheon, South Korea. A phylogenetic analysis based on 16S rRNA gene sequence comparison revealed that the strain formed a distinct lineage within the genus Brevibacterium and was most closely related to B. epidermidis NBRC 14811T (98.4%), B. sediminis FXJ8.269T (98.2%), B. avium NCFB 3055T (98.1%), and B. oceani BBH7T (98.1%), while it shared less than 98.1% identity with the other species of this genus. The DNA G + C content was 68.1 mol%. The predominant quinone was MK-8(H2). The major fatty acids were anteiso-C15:0 and anteiso-C17:0. The cell wall peptidoglycan of strain Gsoil 188T contained meso-diaminopimelic acid. The major polar lipids were phosphatidylglycerol, diphosphatidylglycerol, and an unidentified aminolipid. The physiological and biochemical characteristics, low DNA-DNA relatedness values, and taxonomic analysis allowed the differentiation of strain Gsoil 188T from the other recognized species of the genus Brevibacterium. Therefore, strain Gsoil 188T represents a novel species of the genus Brevibacterium, for which the name Brevibacterium anseongense sp. nov. is proposed, with the type strain Gsoil 188T (= KACC 19439T = LMG 30331T).

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The threonine-tRNA ligase gene region is applicable in classification, typing, and phylogenetic analysis of bifidobacteria: Ji&# , Chahrazed Mekadim , Radko Pechar , V&# , Eva Vlková; J. Microbiol. 2018;56(10):713-721. Published online September 28, 2018; DOI: https://doi.org/10.1007/s12275-018-8167-3

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Abstract

In the modern era, molecular genetic techniques are crucial in ecological studies, as well as in the classification, typing, and phylogenetic analysis of prokaryotes. These techniques are mainly aimed at whole genome comparisons and PCRderived experiments, including amplifying the 16S rRNA and other various housekeeping genes used in taxonomy, as well as MLST (multilocus sequence typing) and MLSA (multilocus sequence analysis) of different taxonomic bacterial groups. The gene encoding threonine-tRNA ligase (thrS) is a gene potentially applicable as an identification and phylogenetic marker in bacteria. It is widely distributed in bacterial genomes and is subject to evolutionary selection pressure due to its important function in protein synthesis. In this study, specific primers were used to amplify a thrS gene fragment (~740 bp) in 36 type and 30 wild strains classified under family Bifidobacteriaceae. The full-length gene has not yet been considered as a possible identification, classification, and phylogenetic marker in bifidobacteria. The thrS sequences revealed higher sequence variability (82.7% of pairwise identities) among members of the family than that shown by 16S rRNA gene sequences (96.0%). Although discrepancies were found between the thrS-derived and previously reported whole genome phylogenetic analyses, the main phylogenetic groups of bifidobacteria were properly assigned. Most wild strains of bifidobacteria were better differentiated based on their thrS sequences than on their 16S rRNA gene identities. Phylogenetic confidence of the evaluated gene with respect to other alternative genetic markers widely used in taxonomy of bifidobacteria (fusA, GroELhsp60, pyrG, and rplB genes) was confirmed using the localized incongruence difference - Templeton analysis.

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Abstract

An obligate anaerobic, Gram-reaction-negative, non-sporeforming, non-motile, rod shaped bacterium designated YMC B3181T was isolated from the blood of a patient with peritonitis. Strain B3181T grew at 20 to 40°C with optimum growth at 37°C. 16S rRNA gene sequence similarity showed strain B3181T belongs to the genus Parabacteroides and is closely related to Parabacteroides faecis 157T (97.3%), Parabacteroides gordonii MS-1T (96.6%), and Parabacteroides goldsteinii DSM 19448T (95.7%). The G + C content of the genomic DNA was 42.3 mol%. The major cellular fatty acids were anteiso- C15:0 and iso-C17:0 3-OH, and the predominant respiratory quinones were MK-9 and MK-10 menaquinones. Genomic and chemotaxonomic data supported affiliation of B3181T with the genus Parabacteroides. Strain B3181T was phylogenetically and phenotypically different from recognized species of the genus Parabacteroides. Accordingly, this isolate belongs to a novel species for which the name Parabacteroides chongii sp. nov. (type strain YMC B3181T = LMG 30065T = KACC 19034T) is proposed.

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Identification of trehalose as a compatible solute in different species of acidophilic bacteria: Pedro A. Galleguillos , Barry M. Grail , Kevin B. Hallberg , Cecilia S. Demergasso , D. Barrie Johnson; J. Microbiol. 2018;56(10):727-733. Published online September 28, 2018; DOI: https://doi.org/10.1007/s12275-018-8176-2

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Abstract

The major industrial heap bioleaching processes are located in desert regions (mainly Chile and Australia) where fresh water is scarce and the use of resources with low water activity becomes an attractive alternative. However, in spite of the importance of the microbial populations involved in these processes, little is known about their response or adaptation to osmotic stress. In order to investigate the response to osmotic stress in these microorganisms, six species of acidophilic bacteria were grown at elevated osmotic strength in liquid media, and the compatible solutes synthesised were identified using ion chromatography and MALDI-TOF mass spectrometry. Trehalose was identified as one of, or the sole, compatible solute in all species and strains, apart from Acidithiobacillus thiooxidans where glucose and proline levels increased at elevated osmotic potentials. Several other potential compatible solutes were tentatively identified by MALDITOF analysis. The same compatible solutes were produced by these bacteria regardless of the salt used to produce the osmotic stress. The results correlate with data from sequenced genomes which confirm that many chemolithotrophic and heterotrophic acidophiles possess genes for trehalose synthesis. This is the first report to identify and quantify compatible solutes in acidophilic bacteria that have important roles in biomining technologies.

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Microbial diversity in the rumen, reticulum, omasum, and abomasum of yak on a rapid fattening regime in an agro-pastoral transition zone: Dan Xue , Huai Chen , Xiaolin Luo , Jiuqiang Guan , Yixin He , Xinquan Zhao; J. Microbiol. 2018;56(10):734-743. Published online August 22, 2018; DOI: https://doi.org/10.1007/s12275-018-8133-0

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Abstract

The ruminant digestive system harbors a complex gut microbiome, which is poorly understood in the case of the four stomach compartments of yak. High-throughput sequencing and quantitative PCR were used to analyse microbial communities in the rumen, reticulum, omasum, and abomasum of six domesticated yak. The diversity of prokaryotes was higher in reticulum and omasum than in rumen and abomasum. Bacteroidetes predominated in the four stomach compartments, with abundance gradually decreasing in the trend rumen > reticulum > omasum > abomasum. Microorganism composition was different among the four compartments, all of which contained high levels of bacteria, methanogens, protozoa and anaerobic fungi. Some prokaryotic genera were associated with volatile fatty acids and pH. This study provides the first insights into the microorganism composition of four stomach compartments in yak, and may provide a foundation for future studies in this area.

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Roles of eIF4E-binding protein Caf20 in Ste12 translation and P-body formation in yeast: Kiyoung Park , Yu-Seon Lee , Daehee Jung , Jinmi Kim; J. Microbiol. 2018;56(10):744-747. Published online August 22, 2018; DOI: https://doi.org/10.1007/s12275-018-8230-0

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Abstract

Translation initiation factor eIF4E forms eIF4E-eIF4G complex at the 5’ cap of mRNA. This interaction can be inhibited by the family of 4E-binding proteins (4E-BP). In yeast Saccharomyces cerevisiae, two 4E-BPs, Caf20 and Eap1, compete with eIF4G for binding to eIF4E via the shared conserved interaction motif. In order to investigate the roles of Caf20 in gene-specific translational regulation and the formation of mRNA granules (P-bodies), we introduced substitution mutations, caf20-Y4A or caf20-L9A, in the eIF4E-binding motif for CAF20. Overexpression of the wild-type CAF20 showed an increased protein level of Ste12 transcription factor as well as highly developed P-body formation. However, 4E-binding site mutations of CAF20 led to a reduced number of P-body foci and decreased levels of Ste12 protein. The phenotypes of the caf20 deletion mutation were also analyzed, and we suggest that Caf20 plays a critical role in Ste12 protein expression and in the control of P-body formation.

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Lysobacter panacihumi sp. nov., isolated from ginseng cultivated soil: Yue Huo , Jong-Pyo Kang , Joon Hurh , Yaxi Han , Jong-Chan Ahn , Ramya Mathiyalagan , Chunhong Piao , Deok-Chun Yang; J. Microbiol. 2018;56(10):748-752. Published online September 28, 2018; DOI: https://doi.org/10.1007/s12275-018-8202-4

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Abstract

A Gram-negative, non-motile, aerobic, catalase-, and oxidasepositive bacterial strain, designated DCY117T, was isolated from ginseng cultivated soil in Gochang-gun, Republic of Korea, and was characterized taxonomically using a multifaceted approach. 16S rRNA gene sequence analysis revealed that strain DCY117T showed highest similarity to Lysobacter ruishenii CTN-1T (95.3%). Phylogenetic analysis revealed that closely related relatives of strain DCY117T were L. aestuarii S2-CT (95.1%), L. daejeonensis GH1-9T (95.0%), and L. caeni BUT-8T (94.9%). Diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), and phosphatidylethanolamine (PE) were the major polar lipids of strain DCY117T. The major isoprenoid quinone was Q-8. The major cellular fatty acids of strain DCY117T were iso-C15:0, iso-C16:0, and summed feature 9 (comprising iso-C17:1 ω9c and/or 10-methyl-C16:0). Genomic DNA G + C content was 61.8 mol%. On the basis of our findings, strain DCY117T is a novel species in the genus Lysobacter. We propose the name Lysobacter panacihumi sp. nov., and the type strain is DCY117T (= KCTC 62019T = JCM 32168T).

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Streptomyces sp. strain SK68, isolated from peanut rhizosphere, promotes growth and alleviates salt stress in tomato (Solanum lycopersicum cv. Micro-Tom): Karthiyaini Damodharan , Sasikumar Arunachalam Palaniyandi , Bao Le , Joo-Won Suh , Seung Hwan Yang; J. Microbiol. 2018;56(10):753-759. Published online September 28, 2018; DOI: https://doi.org/10.1007/s12275-018-8120-5

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Abstract

A novel actinobacterium, strain SK68, was isolated from the rhizosphere of peanut plant and its salinity stress alleviation ability was studied using tomato (Solanum lycopersicum cv. Micro-Tom) plants. Based on 16S rDNA based phylogenetic analysis, strain SK68 has been identified as a Streptomyces sp. Strain SK68 had branched substrate mycelium bearing smooth surfaced spores and the spore colour is brownish grey on ISP4 medium. It exhibited enzyme activities such as xylanase, cellulase, amylase, and pectinase and degraded hypoxanthine, casein, and L-tyrosine. The strain SK68 differed in its banding pattern in BOX-PCR and RAPD fingerprinting compared to the closely matching type strains Streptomyces erythrochromogenes NBRC 3304T (AB184746), S. flavotricini NBRC 12770T (AB184132), S. racemochromogenes NBRC 12906T (AB184235), and S. polychromogenes NBRC 13072T (NR041109). Strain SK68 was evaluated for its salinity stress-alleviating activity in tomato plants with 180 mmol/L NaCl under gnotobiotic condition. A significant increase in plant biomass was observed in strain SK68-inoculated tomato plants under salt stress compared to control and salt-stressed non-inoculated plants.

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Cultivable butyrate-producing bacteria of elderly Japanese diagnosed with Alzheimer’s disease: Thi Thuy Tien Nguyen , Yuta Fujimura , Iyo Mimura , Yusuke Fujii , Ngoc Luong Nguyen , Kensuke Arakawa , Hidetoshi Morita; J. Microbiol. 2018;56(10):760-771. Published online August 22, 2018; DOI: https://doi.org/10.1007/s12275-018-8297-7

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Abstract

The group of butyrate-producing bacteria within the human gut microbiome may be associated with positive effects on memory improvement, according to previous studies on dementia- associated diseases. Here, fecal samples of four elderly Japanese diagnosed with Alzheimer’s disease (AD) were used to isolate butyrate-producing bacteria. 226 isolates were randomly picked, their 16S rRNA genes were sequenced, and assigned into sixty OTUs (operational taxonomic units) based on BLASTn results. Four isolates with less than 97% homology to known sequences were considered as unique OTUs of potentially butyrate-producing bacteria. In addition, 12 potential butyrate-producing isolates were selected from the remaining 56 OTUs based on scan-searching against the PubMed and the ScienceDirect databases. Those belonged to the phylum Bacteroidetes and to the clostridial clusters I, IV, XI, XV, XIVa within the phylum Firmicutes. 15 out of the 16 isolates were indeed able to produce butyrate in culture as determined by high-performance liquid chromatography with UV detection. Furthermore, encoding genes for butyrate formation in these bacteria were identified by sequencing of degenerately primed PCR products and included the genes for butyrate kinase (buk), butyryl-CoA: acetate CoAtransferase (but), CoA-transferase-related, and propionate CoA-transferase. The results showed that eight isolates possessed buk, while five isolates possessed but. The CoA-transfer- related gene was identified as butyryl-CoA:4-hydroxybutyrate CoA transferase (4-hbt) in four strains. No strains contained the propionate CoA-transferase gene. The biochemical and butyrate-producing pathways analyses of butyrate producers presented in this study may help to characterize the butyrate-producing bacterial community in the gut of AD patients.

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