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Draft genome sequence of Olsenella sp. KGMB 04489 isolated from healthy Korean human feces
Korean J. Microbiol 2018;54(4):456-459
Published online December 31, 2018
© 2018 The Microbiological Society of Korea.

Kook-Il Han1, Se Won Kang1, Ji-Sun Kim1, Keun Chul Lee1, Mi Kyung Eom1, Min Kuk Suh1, Seung-Hwan Park1, Ju Huck Lee1, Jam-Eon Park1, Byeong Seob Oh1, Seung Yeob Yu1, Seung-Hyeon Choi1, Dong Ho Lee2, Yoon Hyuk2, Byung-Yong Kim3, Seung-Jo Yang3, and Jung-Sook Lee1,4,*

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: jslee@kribb.re.kr; Tel.: +82-63-570-5618; Fax: +82-63-570-5609
Received September 19, 2018; Revised October 30, 2018; Accepted November 7, 2018.
Abstract

The genus of Olsenella has been isolated from vertebrate animal mouth, rumen, and feces. Olsenella sp. KGMB 04489 was isolated from fecal samples obtained from a healthy Korean. The whole-genome sequence of Olsenella sp. KGMB 04489 was analyzed using the PacBio Sequel platform. The genome comprises a 2,108,034 bp chromosome with a G + C content of 65.50%, 1,838 total genes, 13 rRNA genes, and 52 tRNA genes. Also, we found that strain KGMB 04489 had some genes for hydrolysis enzymes, and antibiotic biosynthesis and resistance in its genome based on the result of genome analysis.

Keywords : Olsenella sp. KGMB 04489, feces, PacBio Sequel
Body
he human gut microbiome plays a very important role in health by helping control digestion and benefiting immune system (Kau et al., 2011). The collection and storage of gut microbes along with their genetic and metabolic profiles are becoming increasingly important for therapeutic approaches to a wide range of diseases (Bolan et al., 2016). Therefore, the Korean gut microbiome bank is launched in 2016. Recently, a novel bacterial strain designated KGMB 04489 was isolated from fecal samples obtained from a healthy korean. On the basis of the phylogenetic, phenotypic and chemotaxonomic characteristics, strains KGMB 04489T (= KCTC 15699T = CCUG 72345T) was found to belong to a novel species as a member of the genus Olsenella within the family Atopobiaceae of actinobacteria.

The genus Olsenella was first proposed by Dewhirst et al. (2001). The reclassification of Atopobiaceae family (www.bacterio.net/atopobiaceae.html) includes the genus Atopobium and Olsenella (Gupta et al., 2013). Members of the genus Olsenella are Gram-positive, non-spore-forming, obligate anaerobic, and non-motile bacteria (Dewhirst et al., 2001; Dewhirst and Wade, 2015; Li et al., 2015). The human oral cavity, bovine rumen, sheep rumen, pig jejunum, and pig feces of homoeothermic vertebrates are indicated as the habitats of Olsenella (Olsen et al., 1991; Kraatz et al., 2011; Li et al., 2015). Here, we describe the draft genome sequence and annotation of Olsenella sp. KGMB 04489 isolated from healthy Korean human feces.

The Olsenella sp. KGMB 04489 was grown in mod-PYG (modified Peptone-Yeast extract with glucose) medium in anaerobic chamber (Coy Laboratory Products) containing 90% N2, 5% H2, and 5% CO2. The composition of mod-PYG agar was as follows: 5.0 g trypticase peptone, 5.0 g peptone, 10.0 g yeast extract, 5.0 g beef extract, 5.0 g glucose, 2.0 g K2HPO4, 1.0 ml Tween 80, 0.5 g cysteine-HCl × H2O, 1.0 mg resazurin, 950.0 ml DW, 40.0 ml salt solution (per L: 0.25 g CaCl2 × 2H2O, 0.5 g MgSO4 × 7H2O, 1.0 g K2HPO4, 1.0 g KH2PO4, 10.0 g NaHCO3, 2.0 g NaCl), 10.0 ml haemin solution (dissolve 50 mg haemin in 1 ml 1 N NaOH; make up to 100 ml with DW), 0.2 ml vitamin K1 solution (dissolve 0.1 ml of vitamin K1 in 20 ml 95% ethanol and filter sterilize), 20 g agar at pH 7.2.

The genomic DNA was extracted using a Wizard genomic DNA purification kit (Promega). Purified genomic DNA sheared to a size of 10 kb using a g-TUBE™ device according to the manufacturer’s instructions (Covaris). Fragmented DNA quantity was analyzed by a Qubit 2.0 fluorometer with a Qubit dsDNA HS Assay Kit (Invitrogen) and size was measured by the Agilent 2100 Bioanalyzer with the DNA 12000 assay (Agilent). 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.

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 was checked by the Contamination Estimator by 16S (ContEst16S) and CheckM tools (Parks et al., 2015; Lee et al., 2017). The coding DNA sequences (CDSs) and tRNAs were predicted using prodigal and tRNAscan-SE, respectively. The CRISPRs were searched using PILER-CR, CRISPR Recognition Tool (CRT), rRNAs, and other non-coding RNAs were searched by covariance model search with inference of Rfam 12.0. The annotation of each CDS was made by homology search against Swiss-prot, EggNOG 4.5, SEED, and KEGG databases.

The genome statistics are showed in Table 1. The draft genome of Olsenella sp. KGMB 04489 is composed of a 2,108,034 bp chromosome with a G + C content of 65.5%. The genome contains 1,838 CDSs, 13 rRNAs (5S, 16S, 23S), and 52 tRNAs (Fig. 1). A total of 1,704 genes were functionally assigned to categories based on clusters of orthologous group (COG) assignments.

General features of Olsenella sp. KGMB 04489

 PropertyValue
Genome assembly
 Assemble methodSMRT Analysis version 4.0
 Genome coverage357×
Genome features
 Genome size (bp)2,108,034
 G + C content (%)65.50
 No. of contigs6
 rRNA genes (5S, 16S, 23S)13 (4, 4, 5)
 tRNA genes52
 Open reading frame1,838
 CDS assigned by COG1,704
 GenBank Accession No.QSNG00000000

Fig. 1.

Graphical circular map of Olsenella sp. KGMB 04489. Marked characteristics are shown from outside to the center; coding DNA sequences (CDS) on forward strand, CDS on reverse strand, tRNA, rRNA, GC content, and GC skew.


We found that various genes involving in hydrolysis, and antibiotic biosynthesis and resistance were identified in the genome. The genome revealed the presence of β-glucosidases and chitinase, which involved in the degradation of cellulose and chitin, respectively. The genome sequence contained genes for antibiotic biosynthesis such as tetracenomycin polyketide synthesis O-methyltransferase TcmP tcmP/elmP and erythromycin 3’’-O-methyltransferase eryG gene. Additionally, the genome had several antibiotic resistance genes, such as undecaprenyl-diphosphate phosphatase bacA, penicillin-binding protein 1A mrcA, macrolide export ATP-binding/permease protein MacB macB, dihydrofolate reductase folA, transcriptional regulatory protein, tetracycline resistance protein TetM from transposon TnFO1, penicillin-binding protein, and macrolide efflux protein. The draft genome sequence of Olsenella sp. KGMB 04489 will contribute to understanding the physiological functions of Olsenella sp. KGMB 04489 in the gut.

Based on the 16S rRNA gene sequence similarity and average nucleotide identity, the strain KGMB 04489 is most closely related to Olsenella scatoligenes SK9K4T with the values of 94.3%.

Nucleotide sequence accession number

Olsenella sp. KGMB 04489 has been deposited in the Korean Collection for Type Cultures under accession number KCTC 15699. The GenBank/EMBL/DDBJ accession number for the genome sequence of Olsenella sp. KGMB 04489 is QSNG00000000.

적 요

Olsenella 속 균주들은 척추동물의 구강, 반추위 및 분변 등에서 분리된 것으로 알려져 있다. 본 연구에서는 건강한 한국인 분변으로부터 Olsenella sp. KGMB 04489 균주를 분리하였으며 PacBio Sequel 플랫폼을 이용하여 Olsenella sp. KGMB 04489 균주의 유전체서열을 분석하였다. 유전체는 G + C 구성 비율이 65.5%이고, 1,838개의 유전자와 rRNA 13개, tRNA 52개로 구성되었으며, 염색체의 크기는 2,108,034 bp였다. 또한, 유전체 분석 결과를 통해 가수분해효소와 항생제 합성 및 내성과 관련된 다양한 유전자를 발견하였다.

Acknowledgements

This work was supported by the Bio & Medical Technology Development program 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.

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December 2018, 54 (4)