Abstract：The resources of saline-alkaline tolerant microorganisms can promote soil nutrient circulation, enhance the water reservation and salinity tolerance ability of the plant, they have important application in remediation of saline-alkaline soil. Previously, a saline-alkaline resistant bacterial strain BFL-3 was isolated, the purpose of this study was to clarify the genetic evolution relationship, saline-alkaline tolerance and mechanism of the strain. The strain BFL-3 was identified by morphology and 16S rRNA sequence analysis. The growth characteristics of strain BFL-3 under different NaCl content and pH stresses were determined by optical density measurement. Besides, the whole genome of strain BFL-3 was sequenced, followed by gene function annotation and bioinformatics analysis. qPCR was applied to verify the expression of genes related to saline-alkaline tolerance of BFL-3 in medium containing 5% NaCl and pH 9. The results showed that the strain BFL-3 was identified as Arthrobacter rhombi, it could tolerant 0.5%~10% of NaCl, the tolerance range of pH value was 5~10. The strain contained 35 scaffolds, the total sequence length was 3 476 199 bp, G+C mol% was 69.15%, coding 3 170 genes, including 50 tRNA genes and 7 rRNA genes. Gene annotation showed that strain BFL-3 was enriched in a large number of genes in amino acid and carbohydrate metabolism pathways, and a certain number of genes in ion transporter related pathways, suggesting that the strain adopts 2 strategies of solute accumulation and ion balance in response to saline-alkaline stress. When the NaCl content was 5% and the pH value was 9, the expression level of trehalose synthase gene (treS), diaminobutyrate-2-oxoglutarate transaminase gene (ectB), glutamate dehydrogenase gene (gdhA), glycine betaine/L-proline ABC transporter permease gene (proW), Trk system potassium transporter gene (trkA) and choline ABC transporter permease gene (opuBD) were up-regulated, 11.0, 6.2, 6.0, 13.1, 7.4 and 6.3 times of the control group, respectively (P<0.01). These results indicated that strain BFL-3 could alleviate the saline-alkaline stress through the synthesis and accumulation of cytocompatible substances (such as trehalose, ectoine and glutamate) and regulating ion balance in the cell. This study clarified the saline-alkaline tolerance characteristics of BFL-3, explored its saline-alkaline tolerance mechanism, provided scientific basis for the further application of this strain in remediation of saline-alkaline soil.
[1] 李娟, Uwaremwe C, 冷艳, 等 . 2017. 节杆菌属细菌处理有机物和重金属污染物的研究进展[J]. 环境科学与技术,40(10): 89-97. (Li J, Uwaremwe C, Leng Y, et al.2017. Progress on the study of biodegradation of organic pol‐ lutants and adsorption of heavy metals with Arthrobacter strains[J]. Environmental Science & Technology, 40(10): 89-97.) [2] 刘国锋, 徐增洪, 么宗利, 等 . 2019. 冲水灌溉对西北硫酸盐型土壤中盐分离子变化的影响研究[J]. 干旱区资源与环境, 33(03): 118-123. (Liu G F, Xu Z H, Yao Z L, et al.2019. Effects of irrigation on the salt ions in sulfate - type saline-alkali soil[J]. Journal of Arid Land Resourc‐ es and Environment, 33(03): 118-123.) [3] 逄焕成, 李玉义, 严慧峻, 等 . 2009. 微生物菌剂对盐碱土理化和生物性状影响的研究[J]. 农业环境科学学报, 28(5): 951-955. (Pang H C, Li Y Y, Yan H J, et al.2009. Effects of inoculating different microorganism agents on the improvement of salinized soil[J]. Journal of Agro- Environment Science, 28(5): 951-955.) [4] 王伟伟, 唐鸿志, 许平 . 2015. 嗜盐菌耐盐机制相关基因的研究进展[J]. 微生物学通报 , 42(3): 550-558. (Wang W W, Tang H Z, Xu P.2015. Salt-tolerance related genes in halophilic bacteria and archaea[J]. Microbiology China,42(3): 550-558.) [5] 张新建, 张广志, 杨合同 . 2016. 节杆菌环境适应性的基因组学研究进展[J]. 微生物学报 , 56(04): 570-577. (Zhang X J, Zhang G Z, Yang H T.2016. Genomics basis of Ar- throbacter spp. environmental adaptability-A review[J]. Acta Microbiologica Sinica, 56(04): 570-577.) [6] Chang P, Gerhardt K E, Huang X D, et al.2014. Plant growth- promoting bacteria facilitate the growth of barley and oats in salt-impacted soil: Implications for phytoremedi‐ ation of saline soils[J]. International Journal of Phytore‐ mediation. 16: 1133-1147. [7] Chen Y G, Tang S K, Zhang Y Q, et al.2009. Arthrobacter halodurans sp. nov., a new halotolerant bacterium isolat‐ed from sea water[J]. Antonie van Leeuwenhoek, 96(1): 63-70. [8] Deen S G, Bondici V F, Essilfie-Dughan J, et al.2018. Biotic and abiotic sequestration of selenium in anoxic coal waste rock[J]. Mine Water and the Environment, 37(4): 825-838. [9] Ding L X, Taketo H, Akira Y.2009. Four novel Arthrobacter species isolated from filtration substrate[J]. International Journal of Systematic and Evolutionary Microbiology, 59(4): 856-862. [10] Ding X W, Liu K H, Lu Y X, et al.2019. Morphological, tran‐ scriptional, and metabolic analyses of osmotic-adapted mechanisms of the halophilic Aspergillus montevidensis ZYD4 under hypersaline conditions[J]. Applied Micro‐ biology and Biotechnology, 103: 3829-3846. [11] Dsouza M, Taylor M W, Turner S J, et al.2015. Genomic and phenotypic insights into the ecology of Arthrobacter from antarctic soils[J]. BMC Genomics, 16(1): 1-18. [12] Fang H, Hu B, Nie Y, et al.2017. The complete genome of Di‐ etzia timorensis ID05-A0528T revealed the genetic ba‐ sis for its saline-alkali tolerance[J]. Journal of Biotech‐ nology, 241: 11-13. [13] Grigoryan A A, Bondici V F, Kryachko Y, et al.2021. Draft genome sequence of Arthrobacter sp. strain 260, isolated from a uranium tailings management facility in North‐ ern Saskatchewan, Canada[J]. Microbiology Resource Announcements, 10(26): e00360-21. [14] Gupta P, Reddy G S N, Delille D, et al.2004. Arthrobacter gan-gotriensis sp. nov. and Arthrobacter kerguelensis sp. nov. from Antarctica[J]. International Journal of Systematic And Evolutionary Microbiology, 54(6): 2375-2378. [15] Kageyama A, Takahashi Y, Morisaki K, et al.2008. Arthrobac- ter oryzae sp. nov. and Arthrobacter humicola sp. nov[J]. International Journal of Systematic and Evolutionary Microbiology, 58(1): 53-56. [16] Keddie R M, Collins M D, Jones D.1986. Genus Arthrobacter[C]//, Sneath P H A, Mair N S, Sharpe M E, Holt J G (eds.). Bergey's manual of systematic bacteriology. New York: Springer Verlag, pp: 1288-1301. [17] Li F, Song W J, Wei J P, et al.2016. Comparative proteomic analysis of phenol degradation process by Arthrobacter[J]. International Biodeterioration & Biodegradation, 110: 189-198. [18] Lin J T, Liang H B, Yan J W, et al.2017. The molecular mech‐ anism and post-transcriptional regulation characteristic of Tetragenococcus halophilus acclimation to osmotic stress revealed by quantitative proteomics[J]. Journal of Proteomics, 168: 1-14. [19] Morino M, Suzuki T, Ito M, et al.2014. Purification and func‐ tional reconstitution of a seven-subunit mrp-type Na+/H+ antiporter[J]. Journal of Bacteriology, 196(1): 28-35. [20] Niewerth H, Schuldes J, Parschat K, et al.2012. Complete ge‐nome sequence and metabolic potential of the quinal‐ dine-degrading bacterium Arthrobacter sp. Rue61a[J]. BMC Genomics, 13(1): 534-534. [21] Osorio C R, Barja J L, Hutson R A, et al.1999. Arthrobacter rhombi sp. nov. isolated from greenland halibut (Rein- hardtius hippoglossoides)[J]. International Journal of Sys‐ tematic Bacteriology, 49(3): 1217-1220. [22] Padan E, Schuldiner S.1994. Molecular physiology of the Na+/H+ antiporter in Escherichia coli[J]. Journal of Exper‐ imental Biology, 196(7): 443-456. [23] Ren L, Jia Y, Ruth N, et al.2015. Complete genome sequence of an aromatic compound degrader Arthrobacter sp. YC -RL1[J]. Journal of Biotechnology, 219: 34-35. [24] Singh K.2016. Microbial and enzyme activities of saline and sodic soils[J]. Land Degradation & Development. 27: 706-718. [25] Vandera E, Samiotaki M, Parapouli M, et al.2015. Compara‐ tive proteomic analysis of Arthrobacter phenanthreniv- orans Sphe3 on phenanthrene, phthalate and glucose[J]. Journal of Proteomics, (113): 73-89. [26] Xu L H, Shi W X, Zeng X C, et al.2017. Draft genome se‐ quence of Arthrobacter sp. strain B6 isolated from the high-arsenic sediments in Datong Basin, China[J]. Stan‐ dards in Genomic Sciences, 12(1): 1-7. [27] Yu X Y, Zhang L, Ren B, et al.2015. Arthrobacter liuii sp. nov., resuscitated from Xinjiang desert soil[J]. Interna‐ tional Journal of Systematic and Evolutionary Microbi‐ ology, 65(3): 896-901. [28] Zhang Y, Ge S J, Jiang M Y, et al.2014. Combined bioremedi‐ ation of atrazine-contaminated soil by Pennisetum and Arthrobacter sp. strain DNS10[J]. Environmental Sci‐ ence and Pollution Research, 21(9): 6234-6238. [29] Zhi X Y, Li W J, Stackebrandt E.2009. An update of the struc‐ ture and 16S rRNA gene sequence-based definition of higher ranks of the class Actinobacteria, with the propos‐ al of two new suborders and four new families and emended descriptions of the existing higher taxa[J]. In‐ ternational Journal of Systematic and Evolutionary Mi‐ crobiology, 59(3): 589-608.
