一株广域pH耐受性微小噬菌体αα的分离、纯化及生理学特性分析

Isolation,Purification and Physiological Characteristics of Microviridae Bacteriophage αα with a Broad pH Resistance

本研究采用双层平板法从污水中分离纯化出一株烈性沙门氏菌噬菌体αα。利用电镜观察其形态特征,通过琼脂糖凝胶电泳分析其核酸组成,并对其生理学特性进行分析,包括一步生长曲线、温度耐受性、pH耐受性、裂解谱、最佳离子浓度和最佳感染复数。结果表明,该烈性噬菌体αα有一个直径为20 nm的头部,无尾部结构,其核酸类型为单链DNA。由此推断,噬菌体αα属于微小噬菌体科。该噬菌体无明显潜伏期,裂解期为60 min,在pH2.10～11.45内和温度40～50℃内效价分别保持在10~6～10~8和10~8pfu/mL左右,最佳感染复数值为10^(-4),钙镁离子均能促进该噬菌体的液体增殖,其中培养基中含10 mmol/L MgCl_2时促进效果最佳,与未添加离子的对照组相比提高了1.5个log。研究结果为噬菌体的多样性及其应用等方面的研究奠定了基础。

A novel lytic Salmonella bacteriophage （hereafter refer to as phage）, named αα was isolated from sewage samples via the overlay method.The morphology of phage αα was checked via transmission electron microscope（TEM） and its nucleic acid composition was checked via agarose gel electrophoresis. In addition, the physiological characteristics of phage αα were also determined,including one - step growth curve, temperature stability, pH stability, host range, optimal ion concentration and optimal multiplicity of infection（MOI） in the liquid proliferation. As a result, the electron micrograph revealed that phage αα was 20 nm in diameter and had no tail, and its nucleic acid was a single - stranded DNA, which proved it to belong to Microviridae family.Phage αα had no significant latent period and the lysis time was 60 min. For pH and thermal stability, it showed that titers of phage aa could be 106-108 pfu/mL with pH varied from 2.10 to 11.45, and 108 pfu/mL between 40 to 50℃ in temperature.In the liquid proliferation, on the one hand, the optimal MOI was 10-4, showing its strong multiplicity.On the other hand, magnesium could facilitate the proliferation of αα, and adding 10 mmol/L MgCl2 had a best effect on the liquid proliferation.The results laid a foundation for the study of phage diversity and its application.