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Draft Genome of a Heavy-Metal-Resistant Bacterium, Cupriavidus Sp

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Korean Journal of Microbiology (2020) Vol. 56, No. 3, pp. 343-346 pISSN 0440-2413 DOI https://doi.org/10.7845/kjm.2020.0061 eISSN 2383-9902 Copyright ⓒ 2020, The Microbiological Society of Korea Draft genome of a heavy-metal-resistant bacterium, Cupriavidus sp. strain SW-Y-13, isolated from river water in Korea Kiwoon Baek , Young Ho Nam , Eu Jin Chung , and Ahyoung Choi* Nakdonggang National Institute of Biological Resources (NNIBR), Sangju 37242, Republic of Korea 강물에서 분리한 중금속 내성 세균 Cupriavidus sp. SW-Y-13 균주의 유전체 해독 백기운 ･ 남영호 ･ 정유진 ･ 최아영* 국립낙동강생물자원관 담수생물연구본부 (Received July 6, 2020; Revised September 18, 2020; Accepted September 18, 2020) Cupriavidus sp. strain SW-Y-13 is an aerobic, Gram-negative, found to survive in close association with pollution-causing rod-shaped bacterium isolated from river water in South Korea, heavy metals, for example, Cupriavidus metallidurans, which in 2019. Its draft genome was produced using the PacBio RS II successfully grows in the presence of Cu, Hg, Ni, Ag, Cd, Co, platform and is thought to consist of five circular chromosomes Zn, and As (Goris et al., 2001; Vandamme and Coenye, 2004; with a total of 7,307,793 bp. The genome has a G + C content Janssen et al., 2010). Several bacteria found in polluted of 63.1%. Based on 16S rRNA sequence similarity, strain SW-Y-13 is most closely related to Cupriavidus metallidurans environments have been shown to adapt to the presence of toxic (98.4%). Genome annotation revealed that the genome is heavy metals. Identification of novel bacterial mechanisms comprised of 6,613 genes, 6,536 CDSs, 12 rRNAs, 61 tRNAs, facilitating growth in heavy-metal-polluted environments and 4 ncRNAs. Resistance to Co2+ is primarily mediated by the provides useful information for the development of novel efflux system encoded by the SW-Y-13 genome, which includes technologies (Ayangbenro and Babalola, 2017). In the present the czcCBA operons, czcD genes, and czcN genes, among others. study, we describe the draft genome sequence of a heavy- This study may provide useful information on the heavy-metal metal-resistant bacterium, Cupriavidus sp. strain SW-Y-13. resistance mechanisms of strain SW-Y-13. Cupriavidus sp. strain SW-Y-13 is a novel organism and was Keywords: Cupriavidus sp., draft genome, heavy-metal-resistant isolated from the surface of the Nakdong River in South Korea bacterium, PacBio RS II (36.432178 N, 128.250770 E) using a standard dilution plating method on YPD agar (BD Difco). This collection was screened 2+ 2+ Cupriavidus belongs to the family Burkholderiaceae, from for heavy metal (Co and Cu ) resistance. Genomic DNA was which the type species Cupriavidus necator was isolated from extracted using the DNeasy Blood and Tissue kit (Qiagen) soil (Makkar and Casida, 1987). Currently, the genus comprises according to the manufacturer’s instructions. Phylogenetic 16 species that inhabit diverse environments (http://www. analysis based on 16S rRNA gene sequences revealed that bacterio.net/cupriavidus). Some Cupriavidus species have been strain SW-Y-13 is closely related to Cupriavidus metallidurans strain CH34T with 98.4% sequence similarity (Fig. 1). *For correspondence. E-mail: [email protected]; To produce sequencing data for the generation of a high- Tel.: +82-54-530-0712; Fax: +82-54-530-0719 344 ∙ Baek et al. Fig. 1. Neighbor-joining phylogenetic tree based on 16S rRNA sequences showing the relationship among isolates belonging to the Cupriavidus sp. SW-Y-13 and related taxa. Numbers at each node indicate bootstrap values (above 50%) based on 1,000 resampled data sets. Scale Bar represents 0.005 substitutions per nucleotide position. quality genome, the genomic DNA from strain SW-Y-13 was (PGAP v4.8) (Tatusova et al., 2016). Potential coding sequences extracted and further purified from a pure culture isolate grown were evaluated using the Basic Local Alignment Search Tool on YPD agar at 25°C, using the DNeasy Blood and Tissue kit (BLAST) and the UniProt (Wu et al., 2006), Pfam (Punta et al., and Wizard Genomic DNA Purification Kit (Promega), 2012), and COG (Tatusov et al., 2003) databases. rRNA and respectively. Whole-genome sequencing was performed on the tRNA genes as well as other miscellaneous features, were PacBio RS II platform (Menlo Park). Single-molecule real-time predicted using the RNAmmer 1.2 (Lagesen et al., 2007) and sequencing (SMRT) yielded a total of 136,681 subreads (1,4 tRNAscan-SE 1.21 (Lowe and Eddy, 1997) servers and the Gb, mean subread length: 10,333 bp, N50 value: 14,631) and Rfam v. 12.0 database (Nawrocki et al., 2015). de novo assembly of the SW-Y-13 genome was completed The basic genome statistics are provided in Table 1. The using the Hierarchical Genome Assembly Process 3 (HGAP3) draft genome of strain SW-Y-13 comprised 5 contigs with a G within the PacBio SMRT analysis 2.3.0 software (Chin et al., + C content of 63.1% and a genome size of 7,307,793 bp (N50, 2013). Contigs were annotated via Rapid Annotation using the 2,677,351 bp; sequencing depth of coverage, 68.7×). The Subsystem Technology (RAST v2.0) server (Aziz et al., 2008) genome contained 6,536 CDSs, 12 rRNAs (5S, 16S, 23S), 61 and the NCBI Prokaryotic Genome Annotation Pipeline tRNAs, and 4 ncRNAs genes (Fig. 2). A total of 4,088 genes 미생물학회지 제56권 제3호 Draft genome sequence of Cupriavidus sp. SW-Y-13 ∙ 345 Table 1. General genomic features of Cupriavidus sp. SW-Y-13 06057), cobalt efflux system protein (czcA; CUSW19178_ Features Value 04268, czcD; CUSW19178_03032, and CUSW19178_06063), Genome size (bp) 7,307,793 copper resistance protein (copD; CUSW19178_06088), nickel G + C content (mol%) 63.1 –cobalt–cadmium resistance protein (nccB; CUSW19178_ No. of contigs 5 04822), nickel and cobalt resistance protein (cnrR; CUSW Genes (total) 6,613 19178_04825 and cnrY; CUSW19178_04826), and arsenic CDSs (total) 6,536 resistance protein (arsB; CUSW19178_04162). The draft Genes (coding) 6,409 genome sequence of SW-Y-13 will contribute to our under- CDSs (with protein) 6,409 standing of the underlying mechanism of heavy metal resistance No. of rRNA genes (5S, 16S, 23S) 12 (4, 4, 4) in Cupriavidus species. No. of tRNA genes 61 were functionally assigned to COG categories. Nucleotide sequence accession number The genome annotation analysis revealed several genes The whole genome sequence of Cupriavidus sp. strain known to be involved with metal resistance, including cobalt– SW-Y-13 was deposited in DDBJ/ENA/GenBank under the zinc–cadmium resistance protein (czcB; CUSW19178_03307, accession number WHLX00000000. The strain was deposited czcC; CUSW19178_03308, CUSW19178_03498, CUSW19 at the Korean Culture Center for Microorganisms under the 178_04266, CUSW19178_06060, and czcN; CUSW19178_ accession number KCCM 43367. Fig. 2. Circular map of the Cupriavidus sp. strain SW-Y-13 genome. From outside to the center; the colored bands in ring 1 represent contigs; ring 2 represents the annotated genes on the forward strand (color determined by COG category); ring 3 shows the annotated genes on the reverse strand (color determined by COG category); ring 4 displays the RNA genes (rRNAs are displayed in red and tRNAs are displayed in purple); ring 5 shows the GC skew (higher-than-average values are displayed in green, while lower-than-average values are displayed in red) and ring 6 shows the GC ratio (higher-than-average values in blue and lower-than-average values in yellow). Korean Journal of Microbiology, Vol. 56, No. 3 346 ∙ Baek et al. 적 요 Clum A, Copeland A, Huddleston J, Eichler EE, et al. 2013. Nonhybrid, finished microbial genome assemblies from long- Cupriavidus sp. SW-Y-13 균주는 2019년 한국의 강물에서 read SMRT sequencing data. Nat. Methods 10, 563–569. Goris J, De Vos P, Coenye T, Hoste B, Janssens D, Brim H, Diels L, 분리된 호기성 그람 음성 막대 모양의 세균이다. PacBio RS II 플 Mergeay M, Kersters K, and Vandamme P. 2001. Classification 랫폼을 사용하여 초안 유전체를 얻었으며, 5개의 원형 염색체로 of metal-resistant bacteria from industrial biotopes as Ralstonia 구성되어있다. SW-Y-13 균주의 유전체 크기는 총 7,307,793 bp, campinensis sp. nov., Ralstonia metallidurans sp. nov. and G + C 함량은 63.1%이다. 16S rRNA 유사도에 따르면 SW-Y- Ralstonia basilensis Steinle et al. 1998 emend. Int. J. Syst. Evol. 13 균주는 Cupriavidus metallidurans (98.4%)와 가장 밀접한 Microbiol. 51, 1773–1782. Janssen PJ, Van Houdt R, Moors H, Monsieurs P, Morin N, Michaux 관련이 있다. 유전체 분석을 통해 총 6,613개의 유전자와 6,539 A, Benotmane MA, Leys N, Vallaeys T, Lapidus A, et al. 2010. 개의 CDS, 12개 rRNA, 61개 tRNA, 4개 ncRNA를 포함하였 The complete genome sequence of Cupriavidus metallidurans 다. CO2+에 대한 내성은 주로 czcCBA 오페론, czcD 유전자 및 strain CH34, a master survivalist in harsh and anthropogenic czcN 유전자를 포함하는 SW-Y-13 게놈에 의해 인코딩된 유 environments. PLoS ONE 5, e10433. Lagesen K, Hallin P, Rødland EA, Staerfeldt HH, Rognes T, and 출 시스템에 의해 매개된다. 이 연구는 SW-Y-13 균주의 중금 Ussery DW. 2007. RNAmmer: consistent and rapid annotation 속 저항 메커니즘에 대한 유용한 정보를 제공할 수 있다. of ribosomal RNA genes. Nucleic Acids Res. 35, 3100–3108. Lowe TM and Eddy SR. 1997. tRNAscan-SE: a program for improved detection of transfer RNA genes in genomic sequence. Nucleic Acids Res. 25, 955–964. Acknowledgments Makkar NS and Casida LE, Jr. 1987. Cupriavidus necator gen. nov., sp. nov.: a nonobligate bacterial predator of bacteria in soil. Int. This work was supported by a grant from the Nakdonggang J. Syst. Bacteriol. 37, 323–326. National Institute of Biological Resources (NNIBR), funded Nawrocki EP, Burge SW, Bateman A, Daub J, Eberhardt RY, Eddy by the Ministry of Environment (MOE) of the Republic of SR, Floden EW, Gardner PP, Jones TA, Tate J, et al.
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    TAXONOMIC DESCRIPTION Kweon et al., Int. J. Syst. Evol. Microbiol. 2021;71:004759 DOI 10.1099/ijsem.0.004759 Cupriavidus cauae sp. nov., isolated from blood of an immunocompromised patient Oh Joo Kweon1†, Wenting Ruan2†, Shehzad Abid Khan2, Yong Kwan Lim1, Hye Ryoun Kim1, Che Ok Jeon2,* and Mi- Kyung Lee1,* Abstract A novel Gram- stain- negative, facultative aerobic and rod- shaped bacterium, designated as MKL-01T and isolated from the blood of immunocompromised patient, was genotypically and phenotypically characterized. The colonies were found to be creamy yellow and convex. Phylogenetic analysis based on 16S rRNA gene and whole-genome sequences revealed that strain MKL-01T was most closely related to Cupriavidus gilardii LMG 5886T, present within a large cluster in the genus Cupriavidus. The genome sequence of strain MKL-01T showed the highest average nucleotide identity value of 92.1 % and digital DNA–DNA hybridization value of 44.8 % with the closely related species C. gilardii LMG 5886T. The genome size of the isolate was 5 750 268 bp, with a G+C content of 67.87 mol%. The strain could grow at 10–45 °C (optimum, 37–40 °C), in the presence of 0–10 % (w/v) NaCl (optimum, 0.5%) and at pH 6.0–10.0 (optimum, pH 7.0). Strain MKL-01T was positive for catalase and negative for oxidase. The major fatty acids were C16 : 0, summed feature 3 (C16 : 1 ω7c/C16 : 1 ω6c and/or C16 : 1 ω6c/C16 : 1 ω7c) and summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c).
  • Host–Symbiont Specificity Determined by Microbe–Microbe Competition in an Insect

    Host–Symbiont Specificity Determined by Microbe–Microbe Competition in an Insect

    Host–symbiont specificity determined by microbe– microbe competition in an insect gut Hideomi Itoha,1, Seonghan Jangb,1, Kazutaka Takeshitac, Tsubasa Ohbayashid, Naomi Ohnishie,2, Xian-Ying Mengf, Yasuo Mitania, and Yoshitomo Kikuchia,b,g,3 aBioproduction Research Institute, National Institute of Advanced Industrial Science and Technology, Hokkaido Center, 062-8517 Sapporo, Japan; bGraduate School of Agriculture, Hokkaido University, 060-8589 Sapporo, Japan; cFaculty of Bioresource Sciences, Akita Prefectural University, 010-0195 Akita, Japan; dInstitute for Integrative Biology of the Cell, UMR 9198, CNRS, Commissariat à l’Energie Atomique et aux Énergies Alternatives (CEA), Université Paris-Sud, 91198 Gif-sur-Yvette, France; eResearch Center for Zoonosis Control, Hokkaido University, 001-0020 Sapporo, Japan; fBioproduction Research Institute, National Institute of Advanced Industrial Science and Technology, Tsukuba Center, 305-8566 Tsukuba, Japan; and gComputational Bio Big Data Open Innovation Laboratory, National Institute of Advanced Industrial Science and Technology, 062-8517 Sapporo, Japan Edited by Joan E. Strassmann, Washington University in St. Louis, St. Louis, MO, and approved September 30, 2019 (received for review July 18, 2019) Despite the omnipresence of specific host–symbiont associations microbe competition on the evolution and stabilization of host– with acquisition of the microbial symbiont from the environment, symbiont specificity is very scarce. little is known about how the specificity of the interaction evolved The bean bug Riptortus pedestris (Heteroptera: Alydidae) is and is maintained. The bean bug Riptortus pedestris acquires a associated with a Burkholderia symbiont that is confined in specific bacterial symbiont of the genus Burkholderia from environ- symbiosis-specific crypts located in the posterior midgut region Burkholderia mental soil and harbors it in midgut crypts.
  • Reclassification of Four Polynucleobacter Necessarius Strains As Representatives of Polynucleobacter Asymbioticus Comb. Nov., Polynucleobacter Duraquae Sp

    Reclassification of Four Polynucleobacter Necessarius Strains As Representatives of Polynucleobacter Asymbioticus Comb. Nov., Polynucleobacter Duraquae Sp

    International Journal of Systematic and Evolutionary Microbiology (2016), 66, 2883–2892 DOI 10.1099/ijsem.0.001073 Reclassification of four Polynucleobacter necessarius strains as representatives of Polynucleobacter asymbioticus comb. nov., Polynucleobacter duraquae sp. nov., Polynucleobacter yangtzensis sp. nov. and Polynucleobacter sinensis sp. nov., and emended description of Polynucleobacter necessarius Martin W. Hahn,1 Johanna Schmidt,1 Alexandra Pitt,1 Sami J. Taipale2 and Elke Lang3 Correspondence 1Research Institute for Limnology, University of Innsbruck, Mondseestrasse 9, A-5310 Mondsee, Martin W. Hahn Austria [email protected] 2Lammi Biological Station, University of Helsinki, Pa€aj€ arventie€ 320, 16900 Lammi, Finland 3Leibniz Institut DSMZ-Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Inhoffenstraße 7B, D-38124 Braunschweig, Germany Genome comparisons based on average nucleotide identity (ANI) values of four strains currently classified as Polynucleobacter necessarius subsp. asymbioticus resulted in ANI values of 75.7– 78.4 %, suggesting that each of those strains represents a separate species. The species P. necessarius was proposed by Heckmann and Schmidt in 1987 to accommodate obligate endosymbionts of ciliates affiliated with the genus Euplotes. The required revision of this species is, however, hampered by the fact, that this species is based only on a description and lacks a type strain available as pure culture. Furthermore, the ciliate culture Euplotes aediculatus ATCC 30859, on which the description of the species was based, is no longer available. We found another Euplotes aediculatus culture (Ammermann) sharing the same origin with ATCC 30859 and proved the identity of the endosymbionts contained in the two cultures. A multilocus sequence comparison approach was used to estimate if the four strains currently classified as Polynucleobacter necessarius subsp.