一株产L-天冬氨酸酶大肠埃希氏菌的噬菌体分离和生物学特性

Isolation and characterization of a phage against an Escherichia coli strain producing L-aspartase

【目的】从大肠埃希氏菌CICC 11021S发酵液中分离一株噬菌体,对其生物学特性进行研究。【方法】采用双层平板法分离噬菌体CICC 80003;利用透射电镜观察噬菌体形态;提取噬菌体基因组,核酸内切酶处理并进行凝胶电泳;分析噬菌体最佳感染复数、一步生长曲线、p H和温度稳定性、宿主谱。考察CICC 80003对CICC 11021S生长和L-天冬氨酸酶活力的影响。【结果】CICC 80003噬菌斑圆形透明,有明显晕环;头部规则,直径约50-60 nm,尾部长约120-130 nm;基因组能被核酸内切酶Bam H I和Mlu I切开;最佳感染复数0.1,潜伏期5 min,裂解期25 min,平均裂解量约86个;最适p H值8.0;90°C温育15 min,噬菌体全部失活;能裂解大肠埃希氏菌和沙门氏菌的部分菌株。发生噬菌体污染时,CICC 11021S无法正常生长,基本检测不到L-天冬氨酸酶活力。【结论】CICC 80003属于长尾噬菌体科ds DNA噬菌体,液体环境中能够彻底裂解大肠埃希氏菌CICC 11021S。

[Objective] A phage was isolated from Escherichia coli CICC 11021 S fermentation media, and its biological characterization was studied. [Methods] A phage designated CICC 80003 was isolated using the double-layer agar culture method. Phage morphology was observed by transmission electron microscopy. Gel electrophoresis of phage genome was carried out after genome extraction and treatment by endonucleases. We investigated the optimal multiplicity of infection, one-step growth curve, p H and temperature stability, and host range. Additionally, we analyzed the effects of CICC 80003 on cell growth and L-aspartase activity of CICC 11021 S. [Results] The phage plaque was round and transparent, and showed clear aureole. The phage had a regular head of 50-60 nm in diameter and a long tail with 120-130 nm in length. The phage genome could be cut by endonucleases Bam H I and Mlu I. The optimal multiplicity of infection was 0.1. The one-step growth curve showed a latent period of 5 min and a rise period of 25 min. The average burst size was about 86 phage particles per infected cell. The optimal p H was 8.0. The phage was completely inactivated when incubated at 90 ℃ for 15 min. Some of Escherichia coli and Salmonella spp. strains could be split by CICC 80003. No cell growth and L-aspartase activity was detected when phage contamination occurred. [Conclusion] CICC 80003 was a ds DNA phage and belonged to the family Siphoviridae, which could completely split E. coli CICC 11021 S in broth.