一株耐盐碱细菌肺炎克雷伯氏菌(Klebsiella pneumoniae) NP36的分离鉴定及全基因组分析

Isolation, identification, and whole-genome analysis of a bacterium Klebsiella pneumoniae NP36 with saline-alkali tolerance

摘要:

【背景】 近年来，大庆地区土壤盐碱化问题日渐严峻，严重威胁作物生长。盐碱土中存在的耐盐碱促生菌可缓解盐碱环境对作物的胁迫。【目的】 从水稻根际土壤中获取耐苏打盐碱的促生菌资源以提高水稻对盐碱环境的抗性，促进水稻生长。【方法】 利用改良后的培养基筛选获得耐盐碱细菌，利用功能培养基测定其促生能力；通过16S rRNA基因序列分析鉴定菌株，并结合全基因组分析菌株基因组信息。【结果】 筛选得到菌株NP36，鉴定结果为肺炎克雷伯氏菌(Klebsiella pneumoniae)，其在pH 3.0−11.0及苏打盐80−200 mmol/L的LB培养基中均可生长。菌株NP36具有产吲哚乙酸(indole-3-acetic acid, IAA)能力，可达(87.27±0.11) mg/L；具有溶无机磷能力，最大有效磷增量为(93.22±1.94) mg/L。相较于无菌水浸种处理，1.0×10⁷ CFU/mL菌悬液浸种显著促进水稻胚根和侧根的生长。通过全基因组测序分析发现，菌株NP36具有合成有机酸和运输磷酸盐的基因。此外，在菌株NP36基因组中还发现氨基酸合成和K⁺转运的相关基因，分析菌株NP36通过细胞氧化还原反应和维持渗透压来抵抗盐碱胁迫。【结论】 本研究验证了菌株NP36对水稻的促生能力，并在基因组水平分析菌株NP36解磷和耐盐碱的机制，为开发耐盐碱促生菌生物肥料提供了理论依据和菌种资源。

Abstract:

[Background] In recent years, the soil salinization in Daqing has become increasingly serious, which threatens crop growth. The presence of plant growth-promoting bacteria with saline-alkali tolerance can alleviate the saline-alkali stress on crops. [Objective] To obtain the strain resources with soda saline-alkali tolerance from rice rhizosphere soil for improving rice tolerance to saline-alkali environments and promote rice growth. [Methods] The modified medium was used for screening of saline-alkali-tolerant bacteria, the ability of which to promote plant growth was determined in a functional medium. The isolate was identified by 16S rRNA gene sequencing and the genomic information of the strain was analyzed based on the whole genome sequence. [Results] The strain NP36 was screened out and identified as Klebsiella pneumoniae, which was capable of growing in the LB medium at pH 3.0−11.0 and soda salts of 80−200 mmol/L. Strain NP36 had the ability to produce indole-3-acetic acid, the concentration of which reached up to (87.27±0.11) mg/L. In addition, it had the ability to solubilize inorganic phosphorus, with a maximum available phosphorus increment of (93.22±1.94) mg/L. Compared with the treatment of seed soaking with sterile water, the seed soaking with 1.0×10⁷ CFU/mL bacterial suspension significantly promoted the growth of radicle and lateral roots of rice. The whole genome sequence analysis showed that strain NP36 carried the genes for the synthesis of organic acids and the transport of phosphates. In addition, the genes related to amino acid synthesis and K⁺ transport were also identified in the genome of strain NP36. The results indicated that strain NP36 could respond to saline-alkali stress by redox reaction and maintenance of osmotic pressure. [Conclusion] We verified the rice growth-promoting ability of strain NP36 and analyzed the mechanism of phosphorus dissolution and saline-alkali tolerance of strain NP36 at the genome level. The findings provide a theoretical basis and strain resources for the development of biofertilizer based on the bacteria with saline-alkali tolerance and plant growth-promoting effects.