search for




 

Draft genome sequence of Bacteroides sp. KGMB 02408 isolated from a healthy Korean feces
Korean J. Microbiol 2019;55(3):296-299
Published online September 30, 2019
© 2019 The Microbiological Society of Korea.

Seung Yeob Yu1, Ji-Sun Kim1, Byeong Seob Oh1, Seoung Woo Ryu1, Seung-Hwan Park1, Se Won Kang1, Jam-Eon Park1, Seung-Hyeon Choi1, Kook-Il Han1, Keun Chul Lee1, Mi Kyung Eom1, Min Kuk Suh1, Han Sol Kim1, Dong Ho Lee2, Hyuk Yoon2, Byung-Yong Kim3, Je Hee Lee3, Jung-Sook Lee1,4, and Ju Huck Lee1*

1Korean Collection for Type Cultures, Korea Research Institute of Bioscience and Biotechnology, Jeongeup 56212, Republic of Korea
2Seoul National University Bundang Hospital, Seongnam 13620, Republic of Korea
3ChunLab, Inc., Seoul 06725, Republic of Korea
4University of Science and Technology (UST), Daejeon 34113, Republic of Korea
Correspondence to: *E-mail: juhuck@kribb.re.kr; Tel.: +82-63-570-5634; Fax: +82-63-570-5609
Received August 13, 2019; Revised September 4, 2019; Accepted September 4, 2019.
Abstract
The genus of Bacteroides has been isolated from vertebrate animal feces. Bacteroides sp. KGMB 02408 was isolated from fecal samples obtained from a healthy Korean. The whole-genome sequence of Bacteroides sp. KGMB 02408 was analyzed using the PacBio Sequel platform. The genome comprises a 5,771,427 bp chromosome with a G + C content of 39.50%, 5,005 total genes, 18 rRNA genes, and 74 tRNA genes. Furthermore, we found that strain KGMB 02408 had some genes for oxidoreductases and menaquinone biosynthesis in its genome based on the result of genome analysis.
Keywords : Bacteroides sp. KGMB 02408, human feces, menaquinone
Body

Various symbiotic microorganisms exist in the human intestine. Gram-negative Bacteriodetes accounts for a large proportion of intestinal microorganisms in healthy adults, most of which are Bacteroides, Alistipes, and Prevotella (The Human Microbiome Project Consortium, 2012). The genus Bacteroides was first proposed by Castellani and Chalmers (1919). Members of the genus Bacteroides are Gram-negative, non-spore-forming, rod-shaped, non-motile and strictly anaerobic bacteria, and comprise more than 92 species (Paster et al., 1994; Smith et al., 2006). As a remarkable genus within the Bacteroidetes phylum, Bacteroides species are one of the most numerous groups of microbiota found in gastrointestinal tract and feces (109–1011 cells/g) of humans and animals, suggesting their strong adaptation in the gut (Finegold et al., 1983). They have various biochemical mechanisms to adopt and survive in the intestinal environment; for example, reducing oxygen level in the gut, degrading and fermenting organic substances present in the colon, and causing host to produce certain food source such as fucosylated glycoprotein consumed by other microorganisms (Wexler et al., 2007; Xu et al., 2007). Furthermore, they can metabolize starch, pullulan and glycogen in the gastrointestinal tract due to their polysaccharide utilization gene cluster (Koropatkin et al., 2012). Since the Bacteroides species possess the abilities and characters mentioned above, they can provide favorable gut environment for other microorganism and maintain host’s gut health and homeostasis. Recently, a novel bacterial strain designated KGMB 02408 was isolated from a healthy Korean feces. On the basis of the phylogenetic, phenotypic and chemotaxonomic characteristics, strains KGMB 02408 (= KCTC 15687T = SSUB 10523T) was found to belong to a novel species as a member of the genus Bacteroides within the family Bacteroidaceae of Bacteroidia. Here, we described the draft genome sequence and annotation of Bacteroides sp. KGMB 02408 isolated from a healthy Korean feces.

The Bacteroides sp. KGMB 02408 was grown in Tryptic Soy Agar (BD) supplemented with 5% sheep blood (TSAB) in anaerobic chamber (Coy Laboratory Products) containing 90% N2, 5% H2, and 5% CO2. To obtain the genomic DNA of strain KGMB 02408, the grown cells on TSAB plates were collected by the loop and then a Wizard genomic DNA purification kit (Promega) was used to extract the genomic DNA. The purified genomic DNA shearing to a size of 10 kb was done by using a g-TUBE™ device according to the manufacturer’s instructions (Covaris). Fragmented DNA size was measured by the Agilent 2100 Bioanalyzer with the DNA 12000 assay (Agilent) and quantity was analyzed by a Qubit 2.0 fluorometer with a Qubit dsDNA HS Assay Kit (Invitrogen). Single-Molecule Real-Time (SMRT) bell library was prepared according to the manufacturer’s instructions (Pacific Biosciences) without a non-size selection. Genome sequencing was performed using a Pacific Biosciences Sequel (Pacific Biosciences) with 2.0 sequencing chemistry and 600 min movies.

The De novo genome assembly was performed with the Hierarchical Genome Assembly Process (HGAP4) pipeline in the SMRT Analysis version 4.0 using default parameters. Potential contamination in genome assembles were checked by the Contamination Estimator by 16S (ContEst16S) and CheckM tools (Li et al., 2015; Parks et al., 2015). The gene prediction algorithm called Prodigal and tRNAscan-SE were used to search coding DNA sequences (CDSs) and tRNAs, respectively. The CRISPRs were found using PILER-CR and CRISPR Recognition Tool (CRT), and rRNAs and other non-coding RNAs were searched by covariance model search with inference of Rfam 12.0. Each of the CDSs was annotated by homology search against Swiss-prot, SEED, EggNOG 4.5, and KEGG databases.

The Table 1 shows the genome statistics; the draft genome of Bacteroides sp. KGMB 02408 is composed of a 5,771,427 bp chromosome with a G + C content of 39.5%. The genome is showed to contain 5,005 CDSs, 18 rRNAs (5S, 16S, 23S), and 74 tRNAs (Fig. 1). A total of 4,162 genes were functionally assigned to categories based on clusters of orthologous group (COG) assignments. The majority of the genes are related to recombination and repair [348 genes (8.36%)] and cell wall/membrane/envelope biogenesis [319 genes (7.66%)].

General features of Bacteroides sp. KGMB 02408

Property Value
Genome assembly
 Assemble method SMRT Analysis version 4.0
 Genome coverage 85.22X
Genome features
 Genome size (bp) 5,771,427
 G + C content (%) 39.50
 No. of contigs 53
 rRNA genes (5S, 16S, 23S) 18 (6, 5, 7)
 tRNA genes 74
 Open reading frame 5,005
 CDS assigned by COG 4,162
 GenBank Accession No. BHWB01000000

Fig. 1.

Graphical circular map of Bacteroides sp. KGMB 02408. From the center to the outside: GC skew (red and green), G + C content (yellow and blue), CDSs on the reverse strand (colored by COG categories), CDS on the forward strand (colored by COG categories), and RNA genes (rRNAs-red and tRNAs-blue)



We have identified a variety of genes involved in oxidative-reductive reaction and menaquinone biosynthesis in the genome. Interestingly, our previous data showed that strain KGMB 02408 was capable of producing acids from D-xylose by dehydrogenase. Moreover, the menaquinone 8 and menaquinone 10 were detected as the major respiratory quinones in KGMB 02408. The genome contains family of the oxidoreductases such as D-xylose 1-dehydrogenase (NADP) HDHD and xdh genes. These genes are involved in D-xylose metabolism, producing D-xylono-1,5-lactone, NADPH and H+ from D-xylose and NADP+. The genome sequence also revealed the genes for quinone biosynthesis such as demethylmenaquinone methyltransferase ubiE and 1,4-dihydroxy-2-naphthoate polyprenyltransferase menA genes. In particular, the ubiE gene catalyzes a chemical reaction, generating menaquinol and S-adenosyl-L-homocysteine from demethylmenaquinol and S-adenosyl-L-methionine in the last step of menaquinone biosynthesis. In addition, the genome has isoprenoid synthesis gene such as isopentenyl-diphosphate Delta-isomerase IdI. This isomerization is a key step in the biosynthesis of isoprenoids through the mevalonate pathway and the MEP pathway, hence strain KGMB 02408 is expected to produce long-chain menaquinones such as menaquinone 8. The draft genome sequence of Bacteroides sp. KGMB 02408 will contribute to understanding the physiological functions of Bacteroides sp. KGMB 02408 in the gut.

Based on the 16S rRNA gene sequence similarity and average nucleotide identity, the strain KGMB 02408 is most closely related to Bacteroides faecichinchillae KCTC 15666T with the values of 96.5%.

Nucleotide sequence accession number

Bacteroides sp. KGMB 02408 has been deposited in the Korean Collection for Type Cultures under accession number KCTC 15687. The GenBank/EMBL/DDBJ accession number for the genome sequence of Bacteroides sp. KGMB 02408 is BHWB01000000.

적 요

Bacteroides 속 균주들은 척추동물의 분변 등에서 분리된 것으로 알려져 있다. 본 연구에서는 건강한 한국인 분변으로부터 Bacteroides sp. KGMB 02408 균주를 분리하였으며 PacBio Sequel 플랫폼을 이용하여 Bacteroides sp. KGMB 02408 균주의 유전체서열을 분석하였다. 유전체는 G + C 구성 비율이 39.5%이고, 5,005개의 유전자와 rRNA 18개 tRNA 74개로 구성되었으며, 염색체의 크기는 5,771,427 bp였다. 또한, 유전체 분석 결과를 통해 산화-환원 효소와 메나퀴논 생합성 및 그와 관련된 다양한 유전자를 발견하였다.

Acknowledgements

This work was supported by the Bio & Medical Technology Development program (Project No. NRF-2016M3A9F3947962) of the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT (MSIT) of the Republic of Korea and a grant from the Korea Research Institute of Bioscience & Biotechnology (KRIBB) Research initiative program.

References
  1. Castellani A, and Chalmers AJ. 1919. Manual of tropical medicine, pp. 959–960. 3rd ed., Williams Wood & Co., New York.
    Pubmed CrossRef
  2. Finegold SM, Sutter VL, and Mathisen GE. 1983. Normal indigenous intestinal flora (pp. 3-31) in Human intestinal microflora in health and disease. Academic Press ISBN 0-12-341280-3.
  3. Koropatkin NM, Cameron EA, and Martens EC. 2012. How glycan metabolism shapes the human gut microbiota. Nat. Rev. Microbiol 10, 323-335.
    Pubmed CrossRef
  4. Li X, Jensen RL, Højberg O, Canibe N, and Jensen BB. 2015. Olsenella scatoligenes sp. nov., a 3-methylindole-(skatole) and 4-methylphenol-(p-cresol) producing bacterium isolated from pig faeces. Int. J. Syst. Evol. Microbiol 65, 1227-1233.
    Pubmed CrossRef
  5. Parks DH, Imelfort M, Skennerton CT, Hugenholtz P, and Tyson GW. 2015. CheckM:assessing the quality of microbial genomes recovered from isolates, single cells, metagenomes. Genome Res 25, 1043-1055.
    CrossRef
  6. Paster BJ, Dewhirst FE, Olsen I, and Fraser GJ. 1994. Phylogeny of Bacteroides, Prevotella and Porphyromonas spp. and related bacteria. J. Bacteriol 176, 725-732.
    Pubmed CrossRef
  7. Smith CJ, Rocha ER, and Paster BJ. 2006. The medically important Bacteroides spp. in health and disease. In Dworkin M, Falkow S, Rosenberg E, Schleifer KH, and Stackebrandt E, et al (eds.) The Prokaryotes: A Handbook on the Biology of Bacteria, 3rd ed., Vol. 7, pp. 381–427. Springer, New York, USA.
  8. The Human Microbiome Project Consortium. 2012. Structure, function and diversity of the healthy human microbiome. Nature 486, 207-214.
    Pubmed CrossRef
  9. Wexler HM. 2007. Bacteroides:the Good, the Bad, the Nitty-Gritty. Clin. Microbiol. Rev 20, 593-621.
    Pubmed CrossRef
  10. Xu J, Mahowald MA, Ley RE, Lozupone CA, Hamady M, Martens EC, Henrissat B, Coutinho PM, Minx P, and Latereille P, et al. 2007. Evolution of symbiotic bacteria in the distal human intestine. PLoS Biol 5, e156.
    Pubmed CrossRef


September 2019, 55 (3)