search for




 

Complete genome sequence of Kitasatospora sp. MMS16-CNU292, an acidophilic actinobacterium
Korean J. Microbiol. 2019;55(4):405-407
Published online December 31, 2019
© 2019 The Microbiological Society of Korea.

Min Ji Kim and Seung Bum Kim*

충남대학교 생명시스템과학대학 미생물 · 분자생명과학과
Correspondence to: *E-mail: sbk01@cnu.ac.kr;
Tel.: +82-42-821-6412; Fax: +82-42-822-7367
Received September 11, 2019; Revised September 23, 2019; Accepted September 25, 2091.
Abstract

Kitasatospora sp. MMS16-CNU292 is an acidophilic actinobacterium showing antibacterial activity, and its complete genome was analyzed. The genome consisted of three contigs and a total length of 8,587,212 bp. The G + C content was 71.5 mol%, and the annotation results revealed presence of 6,934 protein coding genes, 30 rRNAs, 79 tRNAs, and 4 ncRNA genes. Based on 16S rRNA gene sequence, the strain was closest to Kitasatospora azatica KCTC 9699T, showing 98.75% similarity. The analysis of biosynthetic gene clusters for secondary metabolites indicated presence of 30 clusters including those for non-ribosomal peptide synthetases, polypeptide synthases, NRPS-PKS hybrids, lanthipeptides, terpenes, and lassopeptide.

Keywords : Kitasatospora, acidophilc actinobacterium, biosynthetic gene cluster, complete genome
Body

The genus Kitasatospora, first proposed by Omura et al. (1982), belongs to the family Streptomycetaceae (Waksman and Henrici, 1943) together with the genera Streptacidiphilus and Streptomyces. Kitasatospora and Streptomyces are sister taxa and share a number of phenotypic properties, but the former contains major amounts of meso-diaminopimelic acid and galactose in their cell walls (Kämpfer, 2012). Kitasatospora is an aerobic, Gram-stain-positive, non-acid-alcohol-fast actinobacterial group which forms intensively branched, non- fragmenting mycelia (Takahashi et al., 1999). Kitasatospora species were found to produce a large number of secondary metabolites with diverse structures and interesting activities, in particular those active against eukaryotes (Takahashi et al., 2017).

In this study, an acidophilic actinobacterial strain designated Kitasatospora sp. MMS16-CNU292 was isolated from a pine grove soil in Daejeon, Korea (36°22'13.3''N, 127°21'02.6''E). The strain was isolated on acidified starch casein agar (SCA, BD) supplemented with antifungal agents cycloheximidine and nystatin, and also gentamicin, an antibacterial agent active mainly against Gram-negative bacteria (each at 50 µg/ml). The strain was deposited in the Korean Collection for Type Cultures (KCTC) under the accession number KCTC 49011.

Extraction of the genomic DNA for the whole genome sequence of strain MMS16-CNU292 was performed using commercial genomic DNA extraction kit (Solgent). The analysis of whole genome shotgun sequencing for MMS16- CNU292 was carried out using single molecule realtime sequencing technology (SMRT) with the PacBio RS II system (Pacific Biosciences). Generation of the long and highly accurate sequences, contigs and final assembly were conducted by SMRT analysis HGAP 3.0. The NCBI Prokaryotic Genome Annotation Pipeline (PGAP) 4.5 was used for the annotation of genes.

The whole genome sequencing by the PacBio platform produced a total length of 8,587,212 bp composed 3 contigs with the DNA G + C content of 71.5 mol%. The annotation results indicated the presence of the total 7,664 coding sequences (CDS) of which 6,934 were protein-coding genes and 730 were pseudogenes, and 30 rRNAs, 79 tRNAs, and 4 ncRNAs were also identified (Table 1).

The features of MMS16-CNU292 genome

FeatureValue
Genome size8,587,212 bp
G + C content71.5 mol%
Contigs3
Total CDS7,664
Protein coding CDS6,934
Pseudogenes730
rRNA genes (5S, 16S, 23S)30 (10, 10, 10)
tRNA genes79
ncRNAs4

On the basis of 16S rRNA gene sequence similarity, the strain formed a novel evolutionary lineage within Kitasatospora and shared highest similarities with Kitasatospora azatica KCTC 9699T (Nakagaito et al., 1992) (98.75%), Kitasatospora kifunensis IFO 15206T (Groth et al., 2003) (98.74%), Kitasatospora purpeofusca NRRL B-1817T (Labeda et al., 2017) (98.61%), Kitasatospora nipponensis HKI 0315T Groth et al. (2004) (98.42%), and Streptomyces novaecaesareae NRRL B-1267T (Waksman et al., 1948) (98.41%).

The orthoANI values between strain MMS16-CNU292 and the two mostly related type strains K. azatica and K. purpeofusca were calculated as 83.87 and 78.19% respectively, which are clearly below the suggested cutoff for species boundary of 95% (Chun et al., 2018).

The analysis of biosynthetic gene clusters for secondary metabolites showed that the MMS16-CNU292 strain contains 30 biosynthetic clusters in total. These include 5 clusters for non-ribosomal peptide synthetases (NRPS), 2 clusters for polyketide synthase (PKS), 9 clusters for NRPS-PKS hybrids, 5 clusters for lanthipeptides, 3 clusters for terpenes, and 1 cluster for lasso peptide.

Nucleotide sequence accession number

The BioProject number for Kitasatospora sp. MMS16- CNU292 is PRJNA532311, and the sequence accession number for the genome is NZ_VIGB00000000.

적 요

소나무 숲에서 분리한 호산성 방선균 Kitasatosproa sp. MMS16-CNU292 균주는 다수의 세균에 대한 항균 활성을 나타냈다. 본 연구에서는 MMS16-CNU292 분리주의 유전체 분석을 실시하였고, 그 결과 3개의 콘티그로 구성된 총 연장 8,587,212 bp의 유전체 정보를 확보하였다. DNA G + C 함량은 71.5 mol% 였으며, 6,934개의 단백질 지정 유전자, 30개의 rRNA, 79개의 tRNA 및 4개의 ncRNA 유전자를 확인하였다. 16S rRNA 유전자 염기서열 분석 결과 MMS16-CNU292는 Kitasatospora azatica KCTC 9699T와 가장 유사했고, 98.75%의 유사도를 보였다. 이차대사산물에 대한 생합성 유전자 클러스터 분석 결과 non-ribosomal peptide synthetase (NRPS), polyketide synthase (PKS), NRPS-PKS 하이브리드, lanthipeptide, terpene 및 lassopeptide 생합성 유전자 클러스터 등 총 30개 유전자 클러스터를 보유하는 것으로 나타났다.

Acknowledgements

This work was supported by the 2019 CNU Research Grant of Chungnam National University (grant no. 2019-0853-01).

References
  1. Chun J, Oren A, Ventosa A, Christensen H, Arahal DR, da Costa MS, Rooney AP, Yi H, Xu XW, and De Meyer S, et al. 2018. Proposed minimal standards for the use of genome data for the taxonomy of prokaryotes. Int. J. Syst. Evol. Microbiol. 68, 461-466.
    Pubmed CrossRef
  2. Groth I, Rodríguez C, Schütze B, Schmitz P, Leistner E, and Goodfellow M. 2004. Five novel Kitasatospora species from soil: Kitasatospora arboriphila sp. nov., K. gansuensis sp. nov., K. nipponensis sp. nov., K. paranensis sp. nov. and K. terrestris sp. nov. Int. J. Syst. Evol. Microbiol. 54, 2121-2129.
    Pubmed CrossRef
  3. Groth I, Schütze B, Boettcher T, Pullen CB, Rodriguez C, Leistner E, and Goodfellow M. 2003. Kitasatospora putterlickiae sp. nov., isolated from rhizosphere soil, transfer of Streptomyces kifunensis to the genus Kitasatospora as Kitasatospora kifunensis comb. nov., and emended description of Streptomyces aureofaciens Duggar 1948. Int. J. Syst. Evol. Microbiol. 53, 2033-2040.
    Pubmed CrossRef
  4. Kämpfer P. 2012. Genus Incertae sedis I. Kitasatospora, pp. 1768-1777. In Whitman WB, Goodfellow M, Kämpfer P, Busse HJ, Trujillo ME, Ludwig W, Suzuki KI, and Parte AE.. (eds.), Bergey's Manual of Systematic Bacteriology, 2nd edn.. Springer, New York, USA 5.
  5. Labeda DP, Dunlap CA, Rong X, Huang Y, Doroghazi JR, Ju KS, and Metcalf WW. 2017. Phylogenetic relationships in the family Streptomycetaceae using multi-locus sequence analysis. Antonie van Leeuwenhoek. 110, 563-583.
    Pubmed CrossRef
  6. Nakagaito Y, Yokota A, and Hasegawa T. 1992. Three new species of the genus Streptomyces: Streptomyces cochleatus sp. nov., Streptomyces paracochleatus sp. nov., and Streptomyces azaticus sp. nov. J. Gen. Appl. Microbiol. 38, 105-120.
    CrossRef
  7. Omura S, Takahashi Y, Iwai Y, and Tanaka H. 1982. Kitasatospora, a new genus of the order Actinomycetales. J. Antibiot. 35, 1013-1019.
    Pubmed CrossRef
  8. Takahashi Y. 2017. Genus Kitasatospora, taxonomic features and diversity of secondary metabolites. J. Antibiot. 70, 506-513.
    Pubmed CrossRef
  9. Takahashi Y, Seino A, Iwai Y, and Omura S. 1999. Taxonomic study and morphological differentiation of an actinomycete genus, Kitasatospora. Zentralbl. Bakteriol. 289, 265-284.
    Pubmed CrossRef
  10. Waksman SA, and Henrici AT. 1943. The nomenclature and classification of the actinomycetes. J. Bacteriol. 46, 337-341.
    Pubmed KoreaMed CrossRef
  11. Waksman SA, and Henrici AT. 1948. Family Ⅲ. Streptomycetaceae Waksman and Henrici, pp. 929-980. In Breed M. (ed.), Bergey's Manual of Determinative Bacteriology, 6th edn. .


December 2019, 55 (4)