新疆双峰驼源枯草芽孢杆菌的分离及全基因组分析

Isolation and genome-wide analysis of Bacillus subtilis from Bactrian camels in Xinjiang

摘要:

【背景】以常规可培养方法从驼乳和驼粪中分离出3株枯草芽孢杆菌，对体外益生效果较突出的菌株F10431进行全基因组测序。【目的】探究分离菌株的益生潜力，了解其功能基因和遗传基础，为该菌的应用和研究打下理论基础。【方法】采用体外法对枯草芽孢杆菌F10431、2N2N和2N13N的产酶能力、耐受能力、抑菌效果和饲用安全性进行益生评价，利用二代+三代测序技术对其进行全基因组预测和功能注释。【结果】三株枯草芽孢杆菌有一定的耐酸、耐胆盐能力，其中，F10431耐肠液、胃液能力最强；都有较好的产蛋白酶、淀粉酶、纤维素酶和抗氧化酶的能力，菌株F10431蛋白酶活性为(3.17±0.26) nmol/(min·mL)、淀粉酶活性为(0.48±0.04) nmol/(min·mL)、纤维素酶活性为(151.02±8.05) μg/(min·mL)、超氧化物歧化酶活性为(1.81±0.31) μmol/(min·mL)；菌株F10431对金黄色葡萄球菌和大肠杆菌抑制效果明显，但对沙门氏菌抑制较弱；溶血性测试均不产生溶血环，饲用安全。【结论】本研究分离出3株益生性能优良的驼源枯草芽孢杆菌，对体外益生效果较突出的菌株F10431进行全基因组测序，将测序数据提交至NCBI获得GenBank登录号为MW721260。从基因层面阐明了其益生菌特性的功能机制和基因组特征，为相关益生菌制剂的应用提供了理论支持。

Abstract:

[Background] Three strains of Bacillus subtilis were isolated from camel milk and feces by the conventional culture method, and genome sequencing was performed for strain F10431 with strong probiotic effects in vitro. [Objective] To explore the probiotic potential, functional genes, and genetic basis of the isolates, so as to lay a theoretical foundation for the application and research of B. subtilis. [Methods] The enzyme production, stress tolerance, antimicrobial effect, and feeding safety of B. subtilis F10431, 2N2N, and 2N13N were evaluated in vitro. The second- and third-generation sequencing was performed for the whole genome and functional gene annotation of the isolates. [Results] The three strains of B. subtilis had tolerance to acids and bile salt, and strain F10431 had the strongest tolerance to intestinal and gastric fluids. All of the strains were capable of producing protease, amylase, cellulase, and antioxidant enzymes. Particularly, strain F10431 showed the protease activity of (3.17±0.26) nmol/(min·mL), the amylase activity of (0.48±0.04) nmol/(min·mL), the cellulase activity of (151.02±8.05) μg/(min·mL), and the superoxide dismutase activity of (1.81±0.31) μmol/(min·mL). The strains exerted strong inhibitory effects on Staphylococcus aureus and Escherichia coli and weak inhibitory effects on Salmonella. None of the strains produced hemolysis ring, thus being safe for feeding. [Conclusion] In this study, three camel-derived B. subtilis strains with excellent probiotic properties were isolated. F10431 with outstanding probiotic effects in vitro was sequenced, and the sequencing data were submitted to NCBI to obtain the GenBank accession number of MW721260. This study elucidated the functional mechanism and genomic characterization of F10431, providing theoretical support for the application of relevant probiotic preparations.