Abstract
Strains of Amycolatopsis orientalis produce vancomycin or other related glycopeptide antibiotic compounds. Here we report the draft genome sequences of glycopeptide nonproducers Amycolatopsis orientalis subsp. orientalis DSM 43388 and DSM 46075. Their genome information will provide insights into the acquisition and regulation of glycopeptide antibiotic resistance genes.
GENOME ANNOUNCEMENT
Amycolatopsis orientalis is known to produce glycopteptide antibiotics, including vancomycin (1) and other related antibiotic compounds such as orienticins (2), eremomycin (3), and chloroeremomycin (also known as A82846B and LY264826) (4). We previously reported the genome sequence of vancomycin-producing Amycolatopsis orientalis strain KCTC 9412T (5). In this study, the strains DSM 43388 and DSM 46075 (ATCC 35164), which do not produce vancomycin and were obtained from the public culture collection, were chosen for sequencing with a view to understanding the diversification of glycopeptide antibiotic biosynthesis and resistance genes.
Cells were grown in tryptic soy broth containing wheat starch at 30°C. Cells were harvested and resuspended with 50 mM EDTA (pH 8.0) and were subsequently treated with achromopeptidase (5 mg/ml) and lysozyme (10 mg/ml) for cell disruption (6). Genomic DNA (gDNA) was purified by use of a gDNA isolation kit (Promega). Illumina library construction and genome sequencing were performed by the Human Derived Material Center in KRIBB, Daejeon, Republic of Korea, using an Illumina HiSeq 2000 system. A total of 2 × 101-nucleotide (nt) paired reads (47,213,684 and 49,370,206 bp) were produced for a total of 4.77 and 4.99 gb for DSM 43388 and DSM 46075, respectively.
Quality trimming of reads and de novo assembly were done using the CLC Genomics Workbench ver. 6.0.1 (CLC Bio). For DSM 43388, 197 contigs with a G+C content of 70.0% were obtained (total contig length, 8,920,384 bp; N50, 114,355 bp). The DSM 46075 assembly resulted in 182 contigs with a G+C content of 69.4% (total contig length, 9,445,329 bp; N50, 123,184 bp). The assembled sequences were automatically annotated using the RAST server (7). The numbers of protein-coding genes were 8,636 and 9,082.
Unexpectedly, the average nucleotide identities (ANI) of these two strains with the type strain KCTC 9412T, calculated by using the JSpecies program (8), were too low (76.6% and 76.5% for DSM 43388 and DSM 46075, respectively), suggesting these two strains should be reclassified to other species. The two strains are rather close to each other, with a high ANI value (>91%). Vancomycin biosynthetic gene clusters were not identified in the assemblies of DSM 43388 and DSM 46075. Vancomycin resistance genes vanH, vanA, and vanX (9) were located in the front of the vancomycin synthetic modules in the genome of KCTC9412T (5). However, the vanHAX genes in DSM 46075 were located next to genes encoding a VanS/VanR two-component system, which might regulate the expression of glycopeptide antibiotic resistance genes.
Nucleotide sequence accession numbers.
These whole-genome shotgun projects have been deposited at DDBJ/EMBL/GenBank under the accession numbers ASXG00000000 and ASXH00000000 for DSM 43388 and DSM 46075, respectively. The versions described in this paper are ASXG01000000 and ASXH01000000.
ACKNOWLEDGMENTS
This study was supported by the Small and Medium Business Administration (S2060609), Ministry of Trade, Industry and Energy, and the KRIBB Research Initiative Program, Ministry of Science, ICT, and Future Planning, Republic of Korea.
Footnotes
Citation Jeong H, Sim YM, Kim HJ, Lee Y-J, Lee D-W, Lim S-K, Lee SJ. 2013. Genome sequences of Amycolatopsis orientalis subsp. orientalis strains DSM 43388 and DSM 46075. Genome Announc. 1(4):e00545-13. doi:10.1128/genomeA.00545-13.
REFERENCES
- 1. Levine DP. 2006. Vancomycin: a history. Clin. Infect. Dis. 42(Suppl 1):S5–S12 [DOI] [PubMed] [Google Scholar]
- 2. Tsuji N, Kobayashi M, Kamigauchi T, Yoshimura Y, Terui Y. 1988. New glycopeptide antibiotics. I. The structures of orienticins. J. Antibiot. 41:819–822 [DOI] [PubMed] [Google Scholar]
- 3. Berdnikova TF, Tokareva NL, Abramova EA, Dokshina NIu, Potapova NP. 1988. The structure of the aglycon of eremomycin—a new antibiotic of the polycyclic glycopeptide group. Antibiot. Khimioter. 33:566–570 (In Russian.) [PubMed] [Google Scholar]
- 4. Nagaraja R, Berry DM, Hunt AH, Occolowitz JL, Scahbel AA. 1989. Conversion of antibiotic A82846B to Orienticin A and structural relationships of related antibiotics. J. Org. Chem. 54:983–886 [Google Scholar]
- 5. Jeong H, Sim YM, Kim HJ, Lee D-W, Lim S-K, Lee SJ. 2013. Genome sequence of the vancomycin-producing Amycolatopsis orientalis subsp. orientalis strain KCTC 9412T. Genome Announc. 1(3):e00408-13. 10.1128/genomeA.00408-13 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6. Nikodinovic J, Barrow KD, Chuck JA. 2003. High yield preparation of genomic DNA from Streptomyces. BioTechniques 35:932–936 [DOI] [PubMed] [Google Scholar]
- 7. Aziz RK, Bartels D, Best AA, DeJongh M, Disz T, Edwards RA, Formsma K, Gerdes S, Glass EM, Kubal M, Meyer F, Olsen GJ, Olson R, Osterman AL, Overbeek RA, McNeil LK, Paarmann D, Paczian T, Parrello B, Pusch GD, Reich C, Stevens R, Vassieva O, Vonstein V, Wilke A, Zagnitko O. 2008. The RAST server: rapid annotations using subsystems technology. BMC Genomics 9:75 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8. Richter M, Rosselló-Móra R. 2009. Shifting the genomic gold standard for the prokaryotic species definition. Proc. Natl. Acad. Sci. U. S. A. 106:19126–19131 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 9. Marshall CG, Lessard IA, Park I, Wright GD. 1998. Glycopeptide antibiotic resistance genes in glycopeptide-producing organisms. Antimicrob. Agents Chemother. 42:2215–2220 [DOI] [PMC free article] [PubMed] [Google Scholar]