一株耐盐碱芽孢杆菌的分离鉴定及其对水稻的促生分析

Isolation, identification, and rice growth-promoting effect measurement of a salt-alkali-tolerant Bacillus halotolerans strain

摘要:

【背景】 盐碱地是我国重要的土地资源，利用微生物改良土壤环境并促进植物生长近年来已经成为研究的热点。【目的】 从黑龙江省大庆市盐碱地水稻根部土壤中筛选获得1株耐盐碱促生细菌，旨在改善盐碱土环境并促进水稻生长，为开发适用于东北盐碱地地区水稻生长的微生物菌剂提供良好的菌种资源与基础。【方法】 通过微生物分离培养技术筛选得到耐盐碱微生物菌株，并对其进行菌落形态观察、生理生化检测、生长曲线测定、菌株16S rRNA基因鉴定。利用多功能鉴定培养基对菌株进行解磷、产铁、产淀粉酶、耐盐碱性检测。利用平板对峙法对菌株进行抗病性检测。检测菌株产胞外聚合物(extracellular polymeric substances, EPS)、吲哚乙酸(indole-3-acetic acid, IAA)及降碱能力，并通过水稻种子平板促生实验、水培实验对菌株的促生能力进行鉴定。【结果】 从黑龙江省大庆市盐碱地水稻根部土壤中筛选得到一株耐盐碱菌株J-009，杆状，革兰氏阳性，菌落呈现白色褶皱状，菌株在接种12 h后达到生长峰值，生长pH范围8.0-10.0，最高耐盐浓度(NaCl)为5%。菌株J-009对无机磷及有机磷有一定的溶解特性，产铁载体、淀粉酶及H₂S，可利用葡萄糖、阿拉伯糖、山梨醇，不利用蔗糖与麦芽糖，过氧化氢酶、脲酶与纤维素水解酶均为阳性。16S rRNA基因序列分析判定菌株J-009为耐受盐芽孢杆菌(Bacillus halotolerans)，GenBank登录号为PV635949。平板对峙试验检测，菌株J-009对水稻稻瘟病有一定的抑制作用。菌株J-009产EPS的量为(0.581±0.083) g/g。分泌IAA的含量为(12.39±0.66) mg/L，并且菌株J-009的降碱能力可以达到13.9%。在水稻种子平板促生实验、水培实验中，相较于对照组，经菌株J-009处理后的水稻种子的根长和苗长均极显著增加。【结论】 菌株J-009具有良好的耐盐碱性及促生功能，能够有效促进盐碱环境下的水稻生长，为今后盐碱土微生物改良及菌剂研发奠定了基础。

Abstract:

[Background] Saline-alkali land is an important land resource in China. Using microorganisms to improve soil environment and promote plant growth has become a research focus in recent years. [Objective] A strain of saline-tolerant growth-promoting bacteria was selected from rice root soil in the saline-alkali land in Daqing, Heilongjiang. This study is expected to provide elite strain resources for the development of microbial agents suitable for rice growth in the saline-alkali land of Northeast China. [Methods] Saline-tolerant microbial strains were screened by microbial isolation and culture. These strains were subjected to colony morphology observation, physiological and biochemical tests, growth curve determination, and 16S rRNA gene identification. The multifunctional identification media were used to evaluate the phosphorus solubilization, siderophore production, amylase production, and salt-alkali tolerance of the target strain. The disease resistance of the strain was tested by the plate confrontation method. The abilities of the strain to produce extracellular polymetric substance (EPS) and indole-3-acetic acid (IAA) and reduce alkali were examined. Furthermore, the rice growth-promoting effect of the strain was evaluated by rice seed germination and water culture experiments. [Results] A saline-alkali tolerant strain J-009 was screened from the rice root soil in the saline-alkali land in Daqing. The strain was rod-shaped and Gram-positive, with white and wrinkled colonies. The strain reached its growth peak 12 h after inoculation, survived within the range of pH 8.0–10.0, and showed tolerance to the maximum salt (NaCl) concentration of 5%. Strain J-009 was capable of solubilizing inorganic phosphorus and organophosphorus and producing siderophores, amylase, and H₂S. It can utilize glucose, arabinose, and sorbitol but cannot utilize sucrose or maltose. The reactions of peroxidase, urease, and cellulase were all positive. Strain J-009 was identified by 16S rRNA gene sequence analysis as Bacillus halotolerans, with the GenBank accession number PV635949. Strain J-009 had a certain inhibitory effect on rice blast. This strain showed the EPS production of (0.581±0.083) g/g, the IAA secretion of (12.39±0.66) mg/L, and the alkali-reducing capacity of 13.9%. Compared with the control group, the strain J-009 treatment significantly increased the root length and plant height of rice seedlings. [Conclusion] Strain J-009 demonstrated good saline-alkali tolerance and plant growth-promoting effect, and it could effectively promote the growth of rice in saline-alkali environment. The findings lay a foundation for the microbial improvement of saline-alkali soil and the development of microbial agents in the future.