Abstract
An extremely halophilic archaeon, strain ZY21T, was isolated from a subterranean rock salt sample in Yunnan, China. Colonies of strain ZY21T on nutrient-rich agar plates are orange, wet and transparent. Cells are pleomorphic, motile, Gram-stain negative and lyse in distilled water. Cells can grow at 20–55 °C (optimum 42 °C), in the presence of 15–30% (w/v) NaCl (optimum 18–20%) and at pH 6.0–9.5 (optimum 7.5). Mg2+ is required for growth (optimum 0.3 M). The major polar lipids of strain ZY21T are phosphatidylglycerol, phosphatidylglycerol sulfate and phosphatidylglycerol phosphate methyl ester, sulfated mannosyl-glucosyl-glycerol diether-1 and seven unidentified glycolipids. Sequence similarity searches with the 16S rRNA gene and rpoB′ gene showed that strain ZY21T is closely related to Halobellus rufus CBA1103T (sequence similarities: 97.5% for 16S rRNA gene and 93.3% for rpoB′ gene). The DNA G+C content of strain ZY21T was determined to be 63.0 mol% based on the draft genome sequence. Genome-based sequence similarity analysis showed that the values of the ANI, AAI, and DDH were far below the boundary for delineation of new species. Phenotypic, chemotaxonomic characteristics and phylogenetic properties suggest that strain ZY21T represents a novel species in the genus Halobellus, for which the name Halobellus captivus sp. nov. is proposed. The type strain is ZY21T (= CGMCC 1.16343T = NBRC 113439T).
![](http://media.springernature.com/m312/springer-static/image/art%3A10.1007%2Fs10482-019-01332-1/MediaObjects/10482_2019_1332_Fig1_HTML.png)
![](http://media.springernature.com/m312/springer-static/image/art%3A10.1007%2Fs10482-019-01332-1/MediaObjects/10482_2019_1332_Fig2_HTML.png)
Similar content being viewed by others
Abbreviations
- ANI:
-
Average nucleotide identity
- AAI:
-
Average amino-acid identity
- DDH:
-
DNA–DNA hybridization
- CGMCC:
-
China General Microbiological Culture Collection Center
- NBRC:
-
NTE Biological Resource Center (Japanese)
- MES:
-
2-Morpholinoethanesulfonic acid
- PIPES:
-
1,4-Piperazine bis (ethanesulfonic acid)
- CHES:
-
2-(Cyclohexylamino) ethanesulfonic acid
- CAPS:
-
N-Cyclohexyl-3-aminopropanesulfonic acid
- DMSO:
-
Dimethylsulfoxide
- OD:
-
Optical density
- DDH:
-
DNA–DNA hybridization
- TLC:
-
Thin-layer chromatography
References
Auch AF, von Jan M, Klenk HP, Göker M (2010) Digital DNA–DNA hybridization for microbial species delineation by means of genome-to-genome sequence comparison. Stand Genom Sci 2:117–134
Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, Lesin VM, Nikolenko SI, Pham S, Prjibelski AD, Pyshkin AV, Sirotkin AV, Vyahhi N, Tesler G, Alekseyev MA, Pevzner PA (2012) SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing. J Comput Biol 19:455–477
Cha IT, Yim KJ, Song HS, Lee HW, Hyun DW, Kim KN, Seo MJ, Kim D, Choi JS, Lee SJ, Bae JW, Rhee SK, Choi HJ, Rhee JK, Nam YD, Roh SW (2014) Halobellus rufus sp. nov., an extremely halophilic archaeon isolated from non-purified solar salt. Antonie Van Leeuwenhoek 105:925–932
Chun J, Oren A, Ventosa A, Christensen H, Arahal DR, da Costa MS, Rooney AP, Yi H, Xu XW, De Meyer S, Trujillo ME (2018) Proposed minimal standards for the use of genome data for the taxonomy of prokaryotes. Int J Syst Evol Microbiol 68:461–466
Corcelli A, Lobasso S (2006) Characterization of lipids of halophilic archaea. In: Rainey FA, Oren A (eds) Methods in microbiology, extremophiles. Elsevier, Amsterdam, pp 585–613
Cui HL, Zhou PJ, Oren A, Liu SJ (2009) Intraspecific polymorphism of 16S rRNA genes in two halophilic archaeal genera, Haloarcula and Halomicrobium. Extremophiles 13:31–37
Cui HL, Yang X, Gao X, Xu XW (2011) Halobellus clavatus gen. nov., sp. nov. and Halorientalis regularis gen. nov., sp. nov., two new members of the family Halobacteriaceae. Int J Syst Evol Microbiol 61:2682–2689
Cui HL, Yang X, Zhou YG, Liu HC, Zhou PJ, Dyall-Smith ML (2012) Halobellus limi sp. nov. and Halobellus salinus sp. nov., isolated from two marine solar salterns. Int J Syst Evol Microbiol 62:1307–1313
Deja-Sikora E, Gołębiewski M, Kalwasińska A, Krawiec A, Kosobucki P, Walczak M (2019) Comamonadaceae OTU as a remnant of an ancient microbial community in sulfidic waters. Microb Ecol 78:85–101
Dussault HP (1955) An improved technique for staining red halophilic bacteria. J Bacteriol 70:484–485
Felsenstein J (1985) Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39:783–791
Gupta RS, Naushad S, Baker S (2015) Phylogenomic analyses and molecular signatures for the class Halobacteria and its two major clades: a proposal for division of the class Halobacteria into an emended order Halobacteriales and two new orders, Haloferacales ord. nov. and Natrialbales ord. nov., containing the novel families Haloferacaceae fam. nov. and Natrialbaceae fam. nov. Int J Syst Evol Microbiol 65:1050–1069
Gupta RS, Naushad S, Fabros R, Adeolu M (2016) A phylogenomic reappraisal of family-level divisions within the class Halobacteria: proposal to divide the order Halobacteriales into the families Halobacteriaceae, Haloarculaceae fam. nov., and Halococcaceae fam. nov., and the order Haloferacales into the families, Haloferacaceae and Halorubraceae fam. nov. Antonie van Leeuwenhoek 109:565–587, and Erratum in: Antonie van Leeuwenhoek 2016, 109:1521–1523
Gutiérrez C, González C (1972) Method for simultaneous detection of proteinase and esterase activities in extremely halophilic bacteria. Appl Microbiol 24:516–517
Hall TA (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucl Acids Symp Ser 41:95–98
Hartmann R, Sickinger HD, Oesterhelt D (1980) Anaerobic growth of halobacteria. Proc Natl Acad Sci USA 77:3821–3825
Kates M (2010) Techniques of lipidology: isolation, analysis, and identification of lipids, 3rd edn. NewportSomerville, Ottawa
Kumar S, Stecher G, Tamura K (2016) MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol 33:1870–1874
Marmur J (1961) A procedure for the isolation of deoxyribonucleic acid from micro-organisms. J Mol Biol 3:208–218
Medlar AJ, Toronen P, Holm L (2018) AAI-profiler: fast proteome-wide exploratory analysis reveals taxonomic identity, misclassification and contamination. Nucleic Acids Res 46:W479–W485
Meier-Kolthoff JP, Göker M, Spröer C, Klenk HP (2013) When should a DDH experiment be mandatory in microbial taxonomy? Arch Microbiol 195:413–418
Oren A, Ventosa A, Grant WD (1997) Proposed minimal standards for description of new taxa in the order Halobacteriales. Int J Syst Bacteriol 47:233–238
Papke RT, White E, Reddy P, Weigel G, Kamekura M, Minegishi H, Usami R, Ventosa A (2011) A multilocus sequence analysis approach to the phylogeny and taxonomy of the Halobacteriales. Int J Syst Evol Microbiol 61:2984–2995
Papke RT, Corral P, Ram-Mohan N, de la Haba RR, Sánchez-Porro C, Makkay A, Ventosa A (2015) Horizontal gene transfer, dispersal and haloarchaeal speciation. Life (Basel) 5:1405–1426
Pérez-Davó A, Aguilera M, González-Paredes A, Luján Jiménez-Pranteda M, Monteoliva-Sánchez M (2015) Halobellus ramosii sp. nov., an extremely halophilic archaeon isolated from a saline-wetland wildfowl reserve. Int J Syst Evol Microbiol 65:3847–3852
Qiu XX, Mou YZ, Zhao ML, Zhang WJ, Han D, Ren M, Cui HL (2013) Halobellus inordinatus sp. nov., from a marine solar saltern and an inland salt lake of China. Int J Syst Evol Microbiol 63:3975–3980
Rosenzweig WD, Peterson J, Woish J, Vreeland RH (2000) Development of a protocol to retrieve microorganisms from ancient salt crystals. Geomicrobiol J 17:185–192
Smibert RM, Krieg NR (1994) Phenotypic characterization. In: Gerhardt P (ed) Methods for general and molecular bacteriology. American Society for Microbiology, Washington, DC, pp 607–654
Ventosa A, Quesada E, Rodriguez-Valera R, Ruiz-Berraquero F, Ramos-Cormenzana A (1982) Numerical taxonomy of moderately Gram-negative rods. J Gen Microbiol 128:1959–1968
Ventosa A, de la Haba RR, Sánchez-Porro C, Papke RT (2015) Microbial diversity of hypersaline environments: a metagenomic approach. Curr Opin Microbiol 25:80–87
**ao W, Wang ZG, Wang YX, Schneegurt MA, Li ZY, Lai YH, Zhang SY, Wen ML, Cui XL (2013) Comparative molecular analysis of the prokaryotic diversity of two salt mine soils in southwest China. J Basic Microbiol 53:942–952
Yoon SH, Ha SM, Lim JM, Kwon SJ, Chun J (2017) A large-scale evaluation of algorithms to calculate average nucleotide identity. Antonie Van Leeuwenhoek 110:1281–1286
Zhang WJ, Han D, Qiu XX, Zhao ML, Mou YZ, Cui HL, Li ZR (2013) Halobellus rarus sp. nov., a halophilic archaeon from an inland salt lake of China. Antonie Van Leeuwenhoek 104:377–384
Zhao ML, Qiu XX, Zhang WJ, Han D, Cui HL, Li ZR (2014) Halobellus litoreus sp. nov., a halophilic archaeon isolated from a Chinese marine solar saltern. Curr Microbiol 68:156–160
Acknowledgements
We thank Professor Zhu L. Yang from the Kunming Institute of Botany, Chinese Academy of Sciences for the help in sample collection, and we also thank Dr. Hao Yan from the College of Life Sciences, Anhui Normal University, for his technical assistance.
Funding
This work was supported by grants from the National Natural Science Foundation of China (31460003), the Anhui Provincial Key Lab. of the Conservation and Exploitation of Biological Resources (591601), the China Scholarship Council (201808340054) and the Department of Education Anhui Province, China.
Author information
Authors and Affiliations
Contributions
SC conceived the project, analysed the data, and drafted the manuscript. SS, YX, FC, and JL performed the experiment. SS and YX critically revised the manuscript. All authors read and approved the final manuscript.
Corresponding author
Ethics declarations
Conflict of interest
The authors declare that they have no conflict of interest.
Ethical statement
No specific ethical or institutional permits were required to conduct sampling and the experimental studies did not involve endangered or protected species.
Additional information
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Chen, S., Sun, S., Xu, Y. et al. Halobellus captivus sp. nov., an extremely halophilic archaeon isolated from a subterranean salt mine. Antonie van Leeuwenhoek 113, 221–231 (2020). https://doi.org/10.1007/s10482-019-01332-1
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10482-019-01332-1