Draft Genome Sequence of Olsenella Sp. KGMB 04489 Isolated from Healthy Korean Human Feces 건강한 한국인 분변으로부

Draft genome sequence of Olsenella sp. KGMB 04489 isolated from healthy Korean human feces

1 1 1 1 1 1 1 Kook-Il Han , Se Won Kang , Ji-Sun Kim , Keun Chul Lee , Mi Kyung Eom , Min Kuk Suh , Seung-Hwan Park , 1 1 1 1 1 2 2 Ju Huck Lee , Jam-Eon Park , Byeong Seob Oh , Seung Yeob Yu , Seung-Hyeon Choi , Dong Ho Lee , Hyuk Yoon , 3 3 1,4 Byung-Yong Kim , Seung-Jo Yang , and Jung-Sook Lee * 1 Korean Collection for Type Cultures, Korea Research Institute of Bioscience and Biotechnology, Jeongeup 56212, Republic of Korea 2 Seoul National University Bundang Hospital, Seongnam 13620, Republic of Korea 3 ChunLab, Inc. Seoul 06725, Republic of Korea 4 University of Science and Technology (UST), Daejeon 34113, Republic of Korea

건강한 한국인 분변으로부터 분리된 Olsenella sp. KGMB 04489 균주의 유전체 염기서열 초안

한국일1 ･ 강세원1 ･ 김지선1 ･ 이근철1 ･ 엄미경1 ･ 서민국1 ･ 박승환1 ･ 이주혁1 ･ 박잠언1 ･ 오병섭1 ･ 유승엽1 ･ 최승현1 ･ 이동호2 ･ 윤 혁2 ･ 김병용3 ･ 양승조3 ･ 이정숙1,4* 1 2 3 4 한국생명공학연구원 생물자원센터, 분당서울대학교병원, 천랩, 과학기술연합대학원대학교

(Received September 19, 2018; Revised October 30, 2018; Accepted November 7, 2018)

The genus of Olsenella has been isolated from vertebrate system (Kau et al., 2011). The collection and storage of gut animal mouth, rumen, and feces. Olsenella sp. KGMB 04489 microbes along with their genetic and metabolic profiles are was isolated from fecal samples obtained from a healthy becoming increasingly important for therapeutic approaches to Korean. The whole-genome sequence of Olsenella sp. KGMB a wide range of diseases (Bolan et al., 2016). Therefore, the 04489 was analyzed using the PacBio Sequel platform. The Korean gut microbiome bank is launched in 2016. Recently, a 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 novel bacterial strain designated KGMB 04489 was isolated tRNA genes. Also, we found that strain KGMB 04489 had some from fecal samples obtained from a healthy korean. On the genes for hydrolysis enzymes, and antibiotic biosynthesis and basis of the phylogenetic, phenotypic and chemotaxonomic T T resistance in its genome based on the result of genome analysis. characteristics, strains KGMB 04489 (= KCTC 15699 = CCUG T Keywords: Olsenella sp. KGMB 04489, feces, PacBio Sequel 72345 ) 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. The human gut microbiome plays a very important role in (2001). The reclassification of Atopobiaceae family (www.bacterio. health by helping control digestion and benefiting immune net/atopobiaceae.html) includes the genus Atopobium and Olsenella (Gupta et al., 2013). Members of the genus Olsenella *For correspondence. E-mail: [email protected]; are Gram-positive, non-spore-forming, obligate anaerobic, and Tel.: +82-63-570-5618; Fax: +82-63-570-5609 Draft genome sequence of Olsenella sp. KGMB 04489 ∙ 457

non-motile bacteria (Dewhirst et al., 2001; Dewhirst and Wade, search with inference of Rfam 12.0. The annotation of each 2015; Li et al., 2015). The human oral cavity, bovine rumen, sheep CDS was made by homology search against Swiss-prot, rumen, pig jejunum, and pig feces of homoeothermic vertebrates EggNOG 4.5, SEED, and KEGG databases. are indicated as the habitats of Olsenella (Olsen et al., 1991; The genome statistics are showed in Table 1. The draft Kraatz et al., 2011; Li et al., 2015). Here, we describe the draft genome of Olsenella sp. KGMB 04489 is composed of a genome sequence and annotation of Olsenella sp. KGMB 04489 2,108,034 bp chromosome with a G + C content of 65.5%. The isolated from healthy Korean human feces. genome contains 1,838 CDSs, 13 rRNAs (5S, 16S, 23S), and The Olsenella sp. KGMB 04489 was grown in mod-PYG 52 tRNAs (Fig. 1). A total of 1,704 genes were functionally (modified Peptone-Yeast extract with glucose) medium in assigned to categories based on clusters of orthologous group anaerobic chamber (Coy Laboratory Products) containing 90% (COG) assignments.

N2, 5% H2, and 5% CO2. The composition of mod-PYG agar We found that various genes involving in hydrolysis, and was as follows: 5.0 g trypticase peptone, 5.0 g peptone, 10.0 g antibiotic biosynthesis and resistance were identified in the yeast extract, 5.0 g beef extract, 5.0 g glucose, 2.0 g K2HPO4, genome. The genome revealed the presence of β-glucosidases

1.0 ml Tween 80, 0.5 g cysteine-HCl × H2O, 1.0 mg resazurin, and chitinase, which involved in the degradation of cellulose

950.0 ml DW, 40.0 ml salt solution (per L: 0.25 g CaCl2 × and chitin, respectively. The genome sequence contained genes

2H2O, 0.5 g MgSO4 × 7H2O, 1.0 g K2HPO4, 1.0 g KH2PO4, 10.0 for antibiotic biosynthesis such as tetracenomycin polyketide g NaHCO3, 2.0 g NaCl), 10.0 ml haemin solution (dissolve 50 synthesis O-methyltransferase TcmP tcmP/elmP and eryth- mg haemin in 1 ml 1 N NaOH; make up to 100 ml with DW), romycin 3''-O-methyltransferase eryG gene. Additionally, the

0.2 ml vitamin K1 solution (dissolve 0.1 ml of vitamin K1 in 20 genome had several antibiotic resistance genes, such as ml 95% ethanol and filter sterilize), 20 g agar at pH 7.2. undecaprenyl-diphosphate phosphatase bacA, penicillin-binding The genomic DNA was extracted using a Wizard genomic protein 1A mrcA, macrolide export ATP-binding/permease DNA purification kit (Promega). Purified genomic DNA sheared protein MacB macB, dihydrofolate reductase folA, transcrip- TM to a size of 10 kb using a g-TUBE device according to the tional regulatory protein, tetracycline resistance protein TetM manufacturer’s instructions (Covaris). Fragmented DNA quantity from transposon TnFO1, penicillin-binding protein, and macro- was analyzed by a Qubit 2.0 fluorometer with a Qubit dsDNA lide efflux protein. The draft genome sequence of Olsenella sp. HS Assay Kit (Invitrogen) and size was measured by the Agilent KGMB 04489 will contribute to understanding the physiolo- 2100 Bioanalyzer with the DNA 12000 assay (Agilent). Single- gical functions of Olsenella sp. KGMB 04489 in the gut. Molecule Real-Time (SMRT) bell library was prepared according Based on the 16S rRNA gene sequence similarity and average to the manufacturer’s instructions (Pacific Biosciences) without a non-size selection. Genome sequencing was performed using Table 1. General features of Olsenella sp. KGMB 04489 a Pacific Biosciences Sequel (Pacific Biosciences) with 2.0 Property Value sequencing chemistry and 600 min movies. Genome assembly De novo genome assembly was performed with the Hierarchical Assemble method SMRT Analysis version 4.0 Genome Assembly Process (HGAP4) pipeline in the SMRT Genome coverage 357× Analysis version 4.0 using default parameters. Potential con- Genome features tamination in genome assembles was checked by the Con- Genome size (bp) 2,108,034 tamination Estimator by 16S (ContEst16S) and CheckM tools G + C content (%) 65.50 (Parks et al., 2015; Lee et al., 2017). The coding DNA No. of contigs 6 rRNA genes (5S, 16S, 23S) 13 (4, 4, 5) sequences (CDSs) and tRNAs were predicted using prodigal tRNA genes 52 and tRNAscan-SE, respectively. The CRISPRs were searched Open reading frame 1,838 using PILER-CR, CRISPR Recognition Tool (CRT), rRNAs, CDS assigned by COG 1,704 and other non-coding RNAs were searched by covariance model GenBank Accession No. QSNG00000000

Korean Journal of Microbiology, Vol. 54, No. 4 458 ∙ Han et al.

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. nucleotide identity, the strain KGMB 04489 is most closely 04489 균주의 유전체서열을 분석하였다. 유전체는 G + C 구 T related to Olsenella scatoligenes SK9K4 with the values of 성 비율이 65.5%이고, 1,838개의 유전자와 rRNA 13개, tRNA 94.3%. 52개로 구성되었으며, 염색체의 크기는 2,108,034 bp였다. 또 한, 유전체 분석 결과를 통해 가수분해효소와 항생제 합성 및 Nucleotide sequence accession number 내성과 관련된 다양한 유전자를 발견하였다.

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 Acknowledgements for the genome sequence of Olsenella sp. KGMB 04489 is This work was supported by the Bio & Medical Technology QSNG00000000. Development program of the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT 적 요 (MSIT) of the Republic of Korea and a grant from the Korea Research Institute of Bioscience & Biotechnology (KRIBB) Olsenella 속 균주들은 척추동물의 구강, 반추위 및 분변 등 Research initiative program. 에서 분리된 것으로 알려져 있다. 본 연구에서는 건강한 한국 인 분변으로부터 Olsenella sp. KGMB 04489 균주를 분리하 였으며 PacBio Sequel 플랫폼을 이용하여 Olsenella sp. KGMB

미생물학회지 제54권 제4호 Draft genome sequence of Olsenella sp. KGMB 04489 ∙ 459

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