Population Diversity of Isolated Halophilic and Halotolerant Bacteria from Hypersaline Salt Lakes and Evaluation of Ectoine Production

doi:10.13560/j.cnki.biotech.bull.1985.2021-0201

Abstract

Abstract:

To fully understand the population diversity of isolated halophilic and halotolerant bacteria in the Chaka Salt Lake,Keke Salt Lake and Xiaochaidan Salt Lake of 3 major magnesium sulfate subtype hypersaline salt lakes,cultivable halophilic and halotolerant bacteria were screened in RM medium with medium and high salinity. The 16S rRNA gene sequences of the isolated strains were amplified for species identification and canonical correspondence analysis（CCA）of environmental factors. The dominant genera were selected to construct individual phylogenetic trees,and the accumulation of secondary metabolite ectoine was measured by high performance liquid chromatography（HPLC）,and finally the strains with high-yield of ectoine were screened. Results showed that a total of 113 halophilic and halotolerant bacteria were isolated from the three major salt lakes. The dominant genera were Bacillus（42 strains）,Staphylococcus（30 strains）and Halomonas（26 strains）. Their abundance was related to the type of the salt lakes. The dominant genera Bacillus and Halomonas were mostly moderately halophilic bacteria,followed by halotolerant bacteria and weakly halophilic bacteria. Phylogenetic analysis demonstrated that Bacillus bacteria had 5 evolutionary branches,composed of 6 species,and Halomonas bacteria had 6 evolutionary branches,composed of 7 species. The accumulation of ectoine was detected by HPLC,and 7 strains with potential of high ectoine production were obtained. These results indicate that the dominant halophilic and halotolerant bacteria from the magnesium sulfate subtype hypersaline salt lakes in the Qaidam Basin are mainly Bacillus,Staphylococcus and Halomonas,and most of them are moderately halophilic bacteria. The strains with high ectoine production could be used for subsequent application research of ectoine fermentation.

Key words: Qaidam Basin, hypersaline salt lake, cultivable bacteria, population diversity, ectoine

ZHANG Tian-tian, LI Yong-zhen, SHEN Guo-ping, WANG Rong, ZHU De-rui, XING Jiang-wa. Population Diversity of Isolated Halophilic and Halotolerant Bacteria from Hypersaline Salt Lakes and Evaluation of Ectoine Production[J]. Biotechnology Bulletin, 2022, 38(1): 168-178.

Figures/Tables 5

References 55

[1]	沈硕. 青藏高原察尔汗盐湖地区可培养中度嗜盐菌的群落结构与多样性[J]. 微生物学报, 2017, 57(4):490-499.
	Shen S. Community structure and diversity of culturable moderate halophilic bacteria isolated from Qrhan Salt Lake on Qinghai-Tibet Plateau[J]. Acta Microbiol Sin, 2017, 57(4):490-499.
[2]	Han R, Zhang X, Liu J, et al. Microbial community structure and diversity within hypersaline Keke Salt Lake environments[J]. Can J Microbiol, 2017, 63(11):895-908. doi: 10.1139/cjm-2016-0773 URL
[3]	Liu W, Zhang GJ, Xian WD, et al. Halomonas xiaochaidanensis sp. nov., isolated from a salt lake sediment[J]. Arch Microbiol, 2016, 198(8):761-766. doi: 10.1007/s00203-016-1235-3 URL
[4]	郑绵平, 刘喜方. 青藏高原盐湖水化学及其矿物组合特征[J]. 地质学报, 2010, 84(11):1585-1600.
	Zheng MP, Liu XF. Hydrochemistry and minerals assemblages of salt lakes in the Qinghai-Tibet plateau, China[J]. Acta Geol Sin, 2010, 84(11):1585-1600.
[5]	Sun YJ, Lai ZP, Long H, et al. Quartz OSL dating of archaeological sites in Xiao Qaidam Lake of the NE Qinghai-Tibetan Plateau and its implications for palaeoenvironmental changes[J]. Quat Geochronol, 2010, 5(2/3):360-364. doi: 10.1016/j.quageo.2009.02.013 URL
[6]	朱德锐, 韩睿, 石晴, 等. 青藏高原盐湖细菌群落与超盐环境因素的相关性[J]. 中国环境科学, 2017, 37(12):4657-4666.
	Zhu DR, Han R, Shi Q, et al. The correlation between bacterial communities in salt lakes on the Qinghai-Tibet Plateau and super-salt environmental factors[J]. China Environ Sci, 2017, 37(12):4657-4666.
[7]	郑绵平, 刘喜方, 赵文. 西藏高原盐湖的构造地球化学和生物学研究[J]. 地质学报, 2007, 81(12):1698-1708.
	Zheng MP, Liu XF, Zhao W. Tectonogeochemical and biological aspects of salt lakes on the Tibetan Plateau[J]. Acta Geol Sin, 2007, 81(12):1698-1708.
[8]	李璐, 郝春博, 王丽华, 等. 巴丹吉林沙漠盐湖微生物多样性[J]. 微生物学报, 2015, 55(4):412-424.
	Li L, Hao CB, Wang LH, et al. Microbial diversity in salt lakes of Badain Jaran Desert[J]. Acta Microbiol Sin, 2015, 55(4):412-424.
[9]	曹军卫, 沈萍, 李朝阳. 嗜极微生物[M]. 武汉: 武汉大学出版社, 2004.
	Cao JW, Shen P, Li (C/Z)Y. Extremophiles[M]. Wuhan: Wuhan University Press, 2004.
[10]	Mukherjee P, Mitra A, Roy M. Halomonas rhizobacteria of Avicennia marina of Indian sundarbans promote rice growth under saline and heavy metal stresses through exopolysaccharide production[J]. Front Microbiol, 2019(10):1207.
[11]	周延, 王芳, 王琳. 盐湖开发对柴达木盆地盐湖湖水细菌的多样性影响[J]. 化工进展, 2013, 32(S1):234-239.
	Zhou Y, Wang F, Wang L. Influence of salt lake exploitation on the bacterial diversity of the salt lakes in the Qaidam Basin[J]. Chem Ind Eng Prog, 2013, 32(S1):234-239.
[12]	Gan LZ, Li XG, Zhang HM, et al. Preparation, characterization and functional properties of a novel exopolysaccharide produced by the halophilic strain Halomonas saliphila LCB169T[J]. Int J Biol Macromol, 2020, 156:372-380. doi: 10.1016/j.ijbiomac.2020.04.062 URL
[13]	Wang J, Zhou J, Wang Y, et al. Efficient nitrogen removal in a modified sequencing batch biofilm reactor treating hypersaline mustard Tuber wastewater:The potential multiple pathways and key microorganisms[J]. Water Res, 2020, 177:115734. doi: 10.1016/j.watres.2020.115734 URL
[14]	Begmatov SA, Selitskaya OV, Vasileva LV, et al. Morphophysiological features of some cultivable bacteria from saline soils of the Aral sea region[J]. Eurasian Soil Sci, 2020, 53(1):90-96. doi: 10.1134/S1064229320010044 URL
[15]	Wang Q, Cao Z, Liu Q, et al. Enhancement of COD removal in constructed wetlands treating saline wastewater:Intertidal wetland sediment as a novel inoculation[J]. J Environ Manage, 2019, 249:109398. doi: 10.1016/j.jenvman.2019.109398 URL
[16]	王慧敏, 姚倩倩, 李月月, 等. 渗透压冲击下中度嗜盐菌Halomonas sp. 四氢嘧啶类相容性溶质的合成与释放[J]. 微生物学通报, 2018, 45(4):744-752.
	Wang HM, Yao QQ, Li YY, et al. Synjournal and release of ectoines in a moderate halophile Halomonas sp. Y subjected to osmotic shocks[J]. Microbiol China, 2018, 45(4):744-752.
[17]	Nayak PK, Goode M, Chang DP, et al. Ectoine and hydroxyectoine stabilize antibodies in spray-dried formulations at elevated temperature and during a freeze/thaw process[J]. Mol Pharmaceutics, 2020, 17(9):3291-3297. doi: 10.1021/acs.molpharmaceut.0c00395 URL
[18]	Chookietwattana K, Yuwa-Amornpitak T. Data on soil properties and halophilic bacterial densities in the Na Si Nuan secondary forest at kantharawichai district, maha sarakham, Thailand[J]. Data Brief, 2019, 27:104582. doi: 10.1016/j.dib.2019.104582 pmid: 31673585
[19]	Ning Y, Wu X, Zhang C, et al. Pathway construction and metabolic engineering for fermentative production of ectoine in Escherichia coli[J]. Metab Eng, 2016, 36:10-18. doi: 10.1016/j.ymben.2016.02.013 URL
[20]	赵婉雨, 杨渐, 董海良, 等. 柴达木盆地达布逊盐湖微生物多样性研究[J]. 地球与环境, 2013, 41(4):398-405.
	Zhao WY, Yang J, Dong HL, et al. Microbial diversity in the hypersaline dabuxun lake in Qaidam basin, China[J]. Earth Environ, 2013, 41(4):398-405.
[21]	刘文, 杨渐, 吴耿, 等. 青藏高原北部湖泊沉积物中基于不同碳源可培养细菌多样性[J]. 盐湖研究, 2016, 24(2):92-101.
	Liu W, Yang J, Wu G, et al. Diversity of cultivable bacteria based on different carbon sources in the sediments of lakes on northern Qinghai-Tibet plateau[J]. J Salt Lake Res, 2016, 24(2):92-101.
[22]	张欣, 刘静, 沈国平, 等. 基于高通量测序研究青藏高原茶卡盐湖微生物多样性[J]. 微生物学通报, 2017, 44(8):1834-1846.
	Zhang X, Liu J, Shen GP, et al. Illumina-based sequencing analysis of microbial community composition in Chaka Salt Lake in Qinghai-Tibet Plateau[J]. Microbiol China, 2017, 44(8):1834-1846.
[23]	柴丽红, 崔晓龙, 彭谦, 等. 青海两盐湖细菌多样性研究[J]. 微生物学报, 2004, 44(3):271-275.
	Chai LH, Cui XL, Peng Q, et al. Bacterial diversity of two salt lakes in Qinghai[J]. Acta Microbiol Sin, 2004, 44(3):271-275.
[24]	Kumar S, Stecher G, Tamura K. MEGA7:molecular evolutionary genetics analysis version 7. 0 for bigger datasets[J]. Mol Biol Evol, 2016, 33(7):1870-1874. doi: 10.1093/molbev/msw054 URL
[25]	Chen YH, Lu CW, Shyu YT, et al. Revealing the saline adaptation strategies of the halophilic bacterium Halomonas beimenensis through high-throughput omics and transposon mutagenesis approaches[J]. Sci Rep, 2017, 7(1):13037. doi: 10.1038/s41598-017-13450-9 URL
[26]	田磊, 张芳, 沈国平, 等. Ectoine高产菌株Halomonas sp. 的鉴定及紫外诱变选育[J]. 生物学杂志, 2020, 37(4):31-35.
	Tian L, Zhang F, Shen GP, et al. Identification of high-yielding strain Halomonas sp. XH26 for producing ectoine and UV mutagenesis breeding[J]. J Biol, 2020, 37(4):31-35.
[27]	Chen J, Liu P, Chu X, et al. Metabolic pathway construction and optimization of Escherichia coli for high-level ectoine production[J]. Curr Microbiol, 2020, 77(8):1412-1418. doi: 10.1007/s00284-020-01888-6 pmid: 32189048
[28]	刘静, 张欣, 沈国平, 等. 青藏高原小柴旦盐湖微生物群落结构及多样性[J]. 水生态学杂志, 2017, 38(5):55-64.
	Liu J, Zhang X, Shen GP, et al. Microbial community structure and diversity of Xiaochaidan salt lake on the Tibetan Plateau[J]. J Hydroecology, 2017, 38(5):55-64.
[29]	Naghoni A, Emtiazi G, Amoozegar MA, et al. Microbial diversity in the hypersaline Lake Meyghan, Iran[J]. Sci Rep, 2017, 7(1):11522. doi: 10.1038/s41598-017-11585-3 pmid: 28912589
[30]	Escudero L, Oetiker N, Gallardo K, et al. A thiotrophic microbial community in an acidic brine lake in Northern Chile[J]. Antonie Van Leeuwenhoek, 2018, 111(8):1403-1419. doi: 10.1007/s10482-018-1087-8 pmid: 29748902
[31]	Jacob JH, Hussein EI, Shakhatreh MAK, et al. Microbial community analysis of the hypersaline water of the Dead Sea using high-throughput amplicon sequencing[J]. MicrobiologyOpen, 2017, 6(5):e00500. doi: 10.1002/mbo3.2017.6.issue-5 URL
[32]	Tazi L, Breakwell DP, Harker AR, et al. Life in extreme environments:microbial diversity in Great Salt Lake, Utah[J]. Extremophiles, 2014, 18(3):525-535. doi: 10.1007/s00792-014-0637-x URL
[33]	Almeida-Dalmet S, Sikaroodi M, Gillevet P, et al. Temporal study of the microbial diversity of the north arm of great salt lake, Utah, US[J]. Microorganisms, 2015, 3(3):310-326. doi: 10.3390/microorganisms3030310 pmid: 27682091
[34]	Boutaiba S, Hacene H, Bidle KA, et al. Microbial diversity of the hypersaline Sidi ameur and himalatt salt lakes of the Algerian Sahara[J]. J Arid Environ, 2011, 75(10):909-916. pmid: 21909172
[35]	Jaiani E, Kusradze I, Kokashvili T, et al. Microbial diversity and phage-host interactions in the Georgian coastal area of the black sea revealed by whole genome metagenomic sequencing[J]. Mar Drugs, 2020, 18(11):558. doi: 10.3390/md18110558 URL
[36]	刘国红, 刘波, 林乃铨, 等. 芽孢杆菌的系统进化及其属分类学特征[J]. 福建农业学报, 2008, 23(4):436-449.
	Liu GH, Liu B, Lin NQ, et al. Phyletic evolution and taxonomic characteristics of Bacillus[J]. Fujian J Agric Sci, 2008, 23(4):436-449.
[37]	刘国红, 刘波, 王阶平, 等. 基于可培养方法分析云南腾冲小空山火山谷芽胞杆菌分布特征[J]. 微生物学报, 2019, 59(6):1063-1075.
	Liu GH, Liu B, Wang JP, et al. Analysis of the distribution characteristics of Bacillus in the volcanic valley of Xiaokong Mountain in Tengchong, Yunnan based on cultivable methods[J]. Acta Microbiol Sin, 2019, 59(6):1063-1075.
[38]	朱德锐, 刘建, 韩睿, 等. 青海湖嗜盐微生物系统发育与种群多样性[J]. 生物多样性, 2012, 20(4):495-504. doi: 10.3724/SP.J.1003.2012.10224
	Zhu DR, Liu J, Han R, et al. Population diversity and phylogeny of halophiles in the Qinghai Lake[J]. Biodivers Sci, 2012, 20(4):495-504.
[39]	Neifar M, Chouchane H, Najjari A, et al. Genome analysis provides insights into crude oil degradation and biosurfactant production by extremely halotolerant Halomonas desertis G11 isolated from Chott El-Djerid salt-lake in Tunisian desert[J]. Genomics, 2019, 111(6):1802-1814. doi: 10.1016/j.ygeno.2018.12.003 URL
[40]	李坤珺, 龙健. 山西运城盐湖嗜盐细菌的系统发育与种群多样性[J]. 贵州农业科学, 2015, 43(11):95-101.
	Li KJ, Long J. Biodiversity of halophilic BacteriaIn Yuncheng salt lake of Shanxi Province[J]. Guizhou Agric Sci, 2015, 43(11):95-101.
[41]	León MJ, Hoffmann T, Sánchez-Porro C, et al. Compatible solute synjournal and import by the moderate halophile Spiribacter salinus:physiology and genomics[J]. Front Microbiol, 2018, 9:108. doi: 10.3389/fmicb.2018.00108 URL
[42]	Chen PW, Cui ZY, Ng HS, et al. Exploring the additive bio-agent impacts upon ectoine production by Halomonas salina DSM5928T using corn steep liquor and soybean hydrolysate as nutrient supplement[J]. J Biosci Bioeng, 2020, 130(2):195-199. doi: 10.1016/j.jbiosc.2020.03.011 URL
[43]	Krutmann J. Ultraviolet A radiation-induced biological effects in human skin:relevance for photoaging and photodermatosis[J]. J Dermatol Sci, 2000, 23(Suppl 1):S22-S26. doi: 10.1016/S0923-1811(99)00077-8 URL
[44]	Graf R, Anzali S, Buenger J, et al. The multifunctional role of ectoine as a natural cell protectant[J]. Clin Dermatol, 2008, 26(4):326-333. doi: 10.1016/j.clindermatol.2008.01.002 URL
[45]	Sydlik U, Gallitz I, Albrecht C, et al. The compatible solute ectoine protects against nanoparticle-induced neutrophilic lung inflammation[J]. Am J Respir Crit Care Med, 2009, 180(1):29-35. doi: 10.1164/rccm.200812-1911OC URL
[46]	Abdel-Aziz H, Wadie W, Abdallah DM, et al. Novel effects of ectoine, a bacteria-derived natural tetrahydropyrimidine, in experimental colitis[J]. Phytomedicine, 2013, 20(7):585-591. doi: 10.1016/j.phymed.2013.01.009 pmid: 23453305
[47]	Thuoc D, Hien TT, Sudesh K. Identification and characterization of ectoine-producing bacteria isolated from Can Gio mangrove soil in Vietnam[J]. Ann Microbiol, 2019, 69(8):819-828. doi: 10.1007/s13213-019-01474-7
[48]	Chen WC, Yuan FW, Wang LF, et al. Ectoine production with indigenous Marinococcus sp. MAR2 isolated from the marine environment[J]. Prep Biochem Biotechnol, 2020, 50(1):74-81. doi: 10.1080/10826068.2019.1663534 URL
[49]	Sajjad W, Qadir S, Ahmad M, et al. Ectoine:a compatible solute in radio-halophilicStenotrophomonassp. WMA-LM19 strain to prevent ultraviolet-induced protein damage[J]. J Appl Microbiol, 2018, 125(2):457-467. doi: 10.1111/jam.13903 pmid: 29729069
[50]	Anburajan L, Meena B, Sreelatha T, et al. Ectoine biosynjournal genes from the deep sea halophilic eubacteria, Bacillus clausii NIOT-DSB04:Its molecular and biochemical characterization[J]. Microb Pathog, 2019, 136:103693. doi: 10.1016/j.micpath.2019.103693 URL
[51]	Chen Q, Zhang LH, Li XL, et al. Poly-β-hydroxybutyrate/ectoine co-production by ectoine-excreting strain Halomonas Salina[J]. Process Biochem, 2014, 49(1):33-37. doi: 10.1016/j.procbio.2013.09.026 URL
[52]	Zhao Q, Li S, Lv P, et al. High ectoine production by an engineered Halomonas hydrothermalis Y2 in a reduced salinity medium[J]. Microb Cell Fact, 2019, 18(1):184. doi: 10.1186/s12934-019-1230-x pmid: 31655591
[53]	Sauer T, Galinski EA. Bacterial milking:a novel bioprocess for production of compatible solutes[J]. Biotechnol Bioeng, 1998, 57(3):306-313. pmid: 10099207
[54]	艾丽, 王川, 江科, 等. 古盐井中嗜盐菌的分离及四氢嘧啶的制备[J]. 生物技术, 2018, 28(6):597-602.
	Ai L, Wang C, Jiang K, et al. Isolation of halophilic bacteria from ancient salt well and production of ectoine[J]. Biotechnology, 2018, 28(6):597-602.
[55]	姚倩倩, 顾向阳. 中度嗜盐菌Halomonas sp. 的分离和鉴定及其高产四氢嘧啶特性研究[J]. 南京农业大学学报, 2017, 40(1):109-115.
	Yao QQ, Gu XY. Isolation, identification and the high ectoine-producing characteristics of a moderately halophilic bacterium Halomonas sp. W2[J]. J Nanjing Agric Univ, 2017, 40(1):109-115.

培养基 Culture medium	分离菌株数量Isolated strain number			总计 Total
培养基 Culture medium	茶卡盐湖 Chaka Salt Lake	柯柯盐湖 Keke Salt Lake	小柴旦盐湖Xiaochaidan Salt Lake	总计 Total
5% NaCl	38	18	14	70
10% NaCl	9	22	12	43

培养基 Culture medium	分离菌株数量Isolated strain number			总计 Total
培养基 Culture medium	茶卡盐湖 Chaka Salt Lake	柯柯盐湖 Keke Salt Lake	小柴旦盐湖Xiaochaidan Salt Lake	总计 Total
5% NaCl	38	18	14	70
10% NaCl	9	22	12	43

门Phylum	属Genus	5%氯化钠 5% NaCl	10%氯化钠 10% NaCl	菌株数量Strain number			总计Total
门Phylum	属Genus	5%氯化钠 5% NaCl	10%氯化钠 10% NaCl	茶卡盐湖 Chaka Salt Lake	柯柯盐湖 Keke Salt Lake	小柴旦盐湖 Xiaochaidan Salt Lake	总计Total
Firmicutes	Bacillus	32	10	20（42.6%）	16（40%）	6（23.1%）	42
	Staphylococcus	15	15	8（17%）	17（42.5%）	5（19.3%）	30
	Enterococcus	3	0	0（0%）	3（7.5%）	0（0%）	3
	Planococcus	1	0	0（0%）	0（0%）	1（3.8%）	1
	Oceanobacillus	1	0	1（2.1%）	0（0%）	0（0%）	1
Proteobacteria	Halomonas	11	15	10（21.3%）	3（7.5%）	13（50%）	26
	Pantoea	6	0	6（12.8%）	0（0%）	0（0%）	6
	Idiomarina	1	3	2（4.2%）	1（2.5%）	1（3.8%）	4
总计Total		70	43	47	40	26	113

门Phylum	属Genus	5%氯化钠 5% NaCl	10%氯化钠 10% NaCl	菌株数量Strain number			总计Total
门Phylum	属Genus	5%氯化钠 5% NaCl	10%氯化钠 10% NaCl	茶卡盐湖 Chaka Salt Lake	柯柯盐湖 Keke Salt Lake	小柴旦盐湖 Xiaochaidan Salt Lake	总计Total
Firmicutes	Bacillus	32	10	20（42.6%）	16（40%）	6（23.1%）	42
	Staphylococcus	15	15	8（17%）	17（42.5%）	5（19.3%）	30
	Enterococcus	3	0	0（0%）	3（7.5%）	0（0%）	3
	Planococcus	1	0	0（0%）	0（0%）	1（3.8%）	1
	Oceanobacillus	1	0	1（2.1%）	0（0%）	0（0%）	1
Proteobacteria	Halomonas	11	15	10（21.3%）	3（7.5%）	13（50%）	26
	Pantoea	6	0	6（12.8%）	0（0%）	0（0%）	6
	Idiomarina	1	3	2（4.2%）	1（2.5%）	1（3.8%）	4
总计Total		70	43	47	40	26	113

Classification of bacteria	Strain code	Salinity range /（mol·L^-1）	Optimal salinity /（mol·L^-1）	Classification of halophiles	Ectoine accumulation /（mg·g ^-1CDW）	Strain code	Salinity range /（mol·L^-1）	Optimal salinity /（mol·L^-1）	Classification of halophiles	Ectoine accumulation /（mg·g ^-1 CDW）
Bacillus	C10	0.5-2.5	0.5	Moderate halophile	80.08	K15	0-2.5	1.5	Moderate halophile	147.75
	C11	0-2.0	2.0	Moderate halophile	220.56	K17	0.5-2.5	1.0	Moderate halophile	10
	C12	0-3.0	0.5	Moderate halophile	64.47	K26	0-2.5	1.5	Moderate halophile	ND
	C15	0.5-3.0	0.5	Moderate halophile	324.69	K27	0-2.5	0.5	Moderate halophile	18.20
	C18	0-1.5	0.5	Slight halophile	ND	K28	0-2.0	0.5	Slight halophile	32.26
	C20	0-1.5	0	Halotolerant	ND	K29	0-1.5	0.5	Slight halophile	30.81
	C22	0-1.5	0	Halotolerant	ND	K30	0-2.5	1.0	Moderate halophile	18.50
	C23	0-2.0	0	Halotolerant	ND	K31	0-2.0	0.5	Slight halophile	77.60
	C24	0-2.5	0	halotolerant	31.93	K32	0-2.0	1.0	Moderate halophile	100.35
	C25	0-3.0	1.0	Moderate halophile	80	K37	0-2.5	1.5	Moderate halophile	135.84
	C26	0 -2.0	0	halotolerant	ND	K39	0-2.5	0.5	Moderate halophile	23.88
	C27	0-3.0	1.0	Moderate halophile	ND	K40	0-3.0	0	Halotolerant	ND
	C29	0-3.0	0	Halotolerant	ND	K41	0-3.0	0.5	Moderate halophile	10.91
	C30	0-2.0	0	Halotolerant	ND	K42	0-3.0	0	Halotolerant	ND
	C33	0-3.0	0.5	Moderate halophile	50.95	K48	0-3.0	0	Halotolerant	ND
	C37	0-2.5	0	Halotolerant	ND	X5	0.5-3.0	1.0	Moderate halophile	70.09
	C42	0.5-3.0	1.0	Moderate halophile	65.46	X10	0-2.5	0.5	Moderate halophile	19.82
	C43	0.5-3.0	1.5	Moderate halophile	133.33	X11	0-2.0	0.5	Slight halophile	23.99
	C50	0-1.5	0	Halotolerant	ND	X20	0-2.0	0	Halotolerant	ND
	C52	0-1.0	0	Halotolerant	ND	X22	0-2.5	0	Halotolerant	ND
	K11	0-2.5	1.5	Moderate halophile	275.20	X33	0-3.0	0	Halotolerant	ND
Halomonas	C8	0-2.5	0.5	Moderate halophile	30.85	X4	0-3.0	1.0	Moderate halophile	125.18
	C9	0-3.0	1.5	Moderate halophile	ND	X6	0-3.0	1.0	Moderate halophile	53.98
	C13	0-3.0	0.5	Moderate halophile	122.38	X9	0-3.0	0.5	Moderate halophile	41.80
	C17	0.5-3.0	0.5	Moderate halophile	18.66	X21	0-3.0	1.0	Moderate halophile	100.39
	C34	0-3.0	0.5	Moderate halophile	15.71	X26	0-3.0	1.0	Moderate halophile	369.10
	C39	0-3.0	0	Halotolerant	ND	X27	0-3.0	1.5	Moderate halophile	239.51
	C40	0.5-3.0	2.0	Moderate halophile	11.25	X28	0-3.0	1.0	Moderate halophile	296.56
	C41	0-3.0	0.5	Moderate halophile	32.35	X31	0-3.0	1.5	Moderate halophile	41.37
	C44	0-3.0	2.5	Moderate halophile	21.82	X32	0-3.0	0.5	Moderate halophile	65.23
	C51	0-2.5	0	halotolerant	ND	X34	0-3.0	0.5	Moderate halophile	57.61
	K5	0-3.0	0.5	Moderate halophile	9.98	X35	0-3.0	1.0	Moderate halophile	99.65
	K14	0-2.0	1.5	Moderate halophile	96.81	X36	0-3.0	1.0	Moderate halophile	208.56
	K38	0-3.0	0.5	Moderate halophile	46.30	X39	0-3.0	0.5	Moderate halophile	41.26