Complete genome sequence of anti-inflammatory properties Bifidobacterium animalis subsp. lactis IDCC4301 isolated from infant feces

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Korean Journal of Microbiology (2020) Vol. 56, No. 1, pp. 62-64 pISSN 0440-2413

DOI https://doi.org/10.7845/kjm.2020.9144 eISSN 2383-9902

Complete genome sequence of anti-inflammatory properties Bifidobacterium animalis subsp. lactis IDCC4301 isolated from infant feces

Tae-Yoon Kim, Min-Goo Kim, Jin Seok Moon* , Hyuk-Sang Kwon, and SungKu Choi Research Laboratories, ILDONG Pharmaceutical Co., Ltd., Hwaseong 18449, Republic of Korea

유아분변에서 분리한 항염증 효과를 보이는 Bifidobacterium animalis subsp. lactis IDCC4301의 유전체 염기서열 해독

김태윤 ･ 김민구 ･ 문진석* ･ 권혁상 ･ 최성구 일동제약 중앙연구소

(Received November 20, 2019; Revised December 16, 2019; Accepted December 16, 2019)

*For correspondence. E-mail: [email protected];

Tel.: +82-31-371-2896; Fax: +82-31-371-2900

Bifidobacterium animalis subsp. lactis IDCC4301 (= KCTC 13587BP) was isolated from infant feces, and exerted anti- inflammatory effect on dextran sodium sulfate (DSS)-induced colitis mice model as they improved clinical parameters. Here we present the complete genome sequence of strain, which consist of a circular 1,944,141 bp chromosome with GC content is 60.49%, 1,580 coding genes (CDS), 15 pseudo genes, and 67 RNA genes. Within the genes involved with were identified probiotic properties and safety assessments, respectively.

Keywords: Bifidobacterium animalis subsp. lactis, anti-inflammatory, genome sequence

In our previous study, Bifidobacterium animalis subsp.

lactis IDCC4301 (B. lactis IDCC4301) had improved the symptoms of DSS-induced colitis in mice (Je et al., 2018).

Despite the above biotechnological applications of IDCC4301, the genome sequence of this strain is unknown. To gain a better insight into the functional properties on intestinal health in the host, the complete genome of B. lactis IDCC4301 was sequenced.

For this, B. lactis IDCC4301 (isolated from infant feces) was grown in TOS-MUP (Merck) medium at 37°C for 18 h under the anaerobic condition. The culture was maintained in 30%

glycerol solution at -70°C until needed. The genomic DNA of IDCC4301 was extracted from cells after cultivation in BL medium at 37°C using a Wizard Genomic DNA Purification Kit (Promega) and the DNA concentration and quality were checked using a spectrophotometer UV-1601PC (Shimadzu).

The complete genome sequence of strain IDCC4301 was obtained from 1.5 Gb PacBio RS II platform data (Pacific Biosciences) and 1.4 Gb Illumina HiSeq platform data (Illumina) at the Macrogen Inc. The sequence reads were assembled using a de novo assembler HGAP3 (The subreads were extracted using the following parameters: min. subread length: 500 bp;

minimum polymerase read quality: 0.80; minimum read length:

100 bp), and the contigs were polished by using Pilon (v1.21)

(Walker et al., 2014). The IDCC4301 genome was annotated

by NCBI Prokaryotic Genome Annotation Pipeline with best-

placed reference protein set GeneMarkS 2 (v 4.6). Default

parameters were used for all software. Functional classification

of the coding genes was expected to use BLASTP search

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Genome sequence of B. animalis subsp. lactis IDCC4301 ∙

63 Korean Journal of Microbiology, Vol. 56, No. 1

Table 1. Genome features of Bifidobacterium animalis subsp. lactis IDCC

4301

Attribute Value

Genome size (bp) 1,944,141

GC content (%) 60.49

No. of contigs 1 (CP031703.1.)

Total genes 1,662

Protein-coding genes 1,580

Pseudogenes 15

rRNAs (5S, 16S, 23S) 12 (4, 4, 4)

tRNAs 52

ncRNAs 3

against the Clusters of Orthologous Groups (COGs) database (Tatusova et al., 2016). Virulence factors and antibiotic resistance genes were performed using against the virulence factor data- base (VFDB) (Chen et al., 2012) and the comprehensive anti- biotic resistance database (CARD) (McArthur et al., 2013).

The complete genome of the strain IDCC4301 consists of single circular chromosome that is 1,944,141 bp with 60.49%

GC-content. A total of 1,662 predicted genes, 1,580 protein coding genes, 52 transfer RNA (tRNA), 12 ribosomal RNA (rRNA), 3 ncRNA, and 15 pseudo genes were annotated (Table 1). No plasmids were present in the genome.

We compared the average nucleotide identity (ANI) value between strain IDCC4301 and B. lactis DSM 10140

^T

and B.

animalis ATCC 25527

^T

which were the most closely related species on the basis of high 16S rRNA similarities (100.00 and 98.90%, respectively). The OrthoANI value was calculated using EZbiocloud (http://www.ezbiocloud.net/tools/ani) (Yoon et al., 2017). Strain IDCC4301 was the highest OrthoANI value of 99.87% with B. lactis DSM 10140

^T

and a value of 95.96%

with B. animalis ATCC 25527

^T

(Chun et al., 2018).

The genome contained oxidative stress response genes such as Na

⁺

:H

⁺

antiporter NhaA (D0Y52_00876), choloylglycine hydrolase (D0Y52_00667), nicotinamidase (D0Y52_00091 and D0Y52_00922), thiol peroxidase (D0Y52_00669), gluta- redoxin (D0Y52_00136), and thioredoxin reductase (D0Y52_

00670, D0Y52_01181, and D0Y52_01429). In addition, the genes involved in cell adhesion, such as fibronectin binding protein (FnBP) (D0Y52_00726) and chaperonin (D0Y52_00166 and D0Y52_00462) were identified. These results elucidate the adhesive capacity of IDCC4301 to epithelial cells and its

survival strategy in the gastrointestinal environment.

The determination of the antibiotic resistant gene is con- firmed according to the reference presented by CARD. The tetW gene (D0Y52_00001) was exhibited to chromosomal DNA in this strain and thus is not likely to be transferable. Four toxic genes (cytolysin, aggregation substance, hyaluronidase, gelatinase) were detected in the genome data. Toxicity genes were extracted by comparing nucleotide sequences based on four selected virulence factors. A blastn program was used for the analysis and a specific parameter was used to search for sequences that are likely to be toxic genes. As a result, the four virulence genes were not detected in the genome data.

The genomic data onto B. lactis IDCC4301 provides a genetic basis to further elucidate its mechanism of anti-inflammatory activity and will facilitate its application in the functional food industry.

Nucleotide sequence accession number(s)

B. lactis IDCC4301 has been deposited in the korean collection for type cultures under accession number KCTC 13587BP, and its complete genome sequence has been deposited in GenBank under accession number CP031703.1.

적 요

Bifidobacterium animalis subsp. lactis IDCC4301은 항염 증 효과가 우수한 균주로서, 유아 분변에서 분리되었다. 균주 의 전체 크기는 1,944,141 bp이고 G + C 함량은 60.49%였으며 1,580개의 단백질 암호화 유전자 및 67개의 RNA 유전자를 보 유하고 있었다. 이 결과를 기초로 IDCC4301 균주 유전체의 프 로바이오틱 및 안전성 관련 유전자를 확인할 수 있었다.

Acknowledgments

This work was supported by Korea Institute of planning and

evaluation for technology in food, agriculture, forestry and

fisheries (IPET) through high value-added food technology

development program, funded by ministry of agriculture, food

and rural affairs (MAFRA) (116017032SB010).

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