He Jiwei, Li Yuxing, Zong Shuaizhou, Qian Min, Zhang Yaru, Zhang Zhangsheng, Yu Haijie, Qu Daofeng
Food Safety Key Laboratory of Zhejiang Province, School of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou 310018, Zhejiang, China.
Jiaxing Vocational & Technical College, Jiaxing 314036, Zhejiang, China.
Sheng Wu Gong Cheng Xue Bao. 2024 Sep 25;40(9):3216-3232. doi: 10.13345/j.cjb.240094.
We analyzed the biological and genome characteristics of a phage infecting enteroinvasive (EIEC), aiming to provide resources and a reference for the prevention and treatment of EIEC. With the EIEC preserved in our laboratory as the host bacterium, one strain of phage was isolated from the effluent sample from a chicken farm in Huzhou, Zhejiang and named ΦEP1. The titer, optimal multiplicity of infection, one-step growth curve, temperature, pH value, chloroform and bile salt sensitivity of ΦEP1 were determined by the double-layer agar plate method. The morphology of the phage was observed by transmission electron microscopy. The biocontrol effects of ΦEP1 in different food matrixes and the protective effect of this phage on Caco-2 cells were tested. The phage ΦEP1 showed the optimal multiplicity of infection of 0.1, the titer of 1.3×10 PFU/mL, strong tolerance to temperature, pH, chloroform, and bile salt, and a broad host spectrum. Furthermore, it expressed lysis activity against multiple strains of multiple antibiotic-resistant pathogenic . and with different serotypes. Phage ΦEP1 had an incubation period of 10 min, an outbreak period of 80 min, and an outbreak volume of 48 PFU/cell. According to the morphology observed by transmission electron microscopy, phage ΦEP1 belonged to the order of , and it had a good protective effect on Caco-2 cells. Phage ΦEP1 had a genome of 87 182 bp with the GC content of 39.80%, 128 putative open reading frames, and no antibiotic resistance genes or virulence genes. ΦEP1 inhibited the growth of EIEC in artificially contaminated milk and beef and eliminated EIEC in cell protection experiments. It significantly increased the survival rate of Caco-2 cells and down-regulated the expression of interleukin (IL)-6 and IL-1β to reduce inflammation. We obtained an EIEC-targeting phage ΦEP1 with a high titer and strong tolerance to the environment, which provided a basis for the application of phages in food preservation and other fields.
我们分析了一种感染肠侵袭性大肠杆菌(EIEC)的噬菌体的生物学和基因组特征,旨在为EIEC的防治提供资源和参考。以本实验室保存的EIEC作为宿主菌,从浙江省湖州市某养鸡场的污水样本中分离出一株噬菌体,命名为ΦEP1。采用双层琼脂平板法测定了ΦEP1的效价、最佳感染复数、一步生长曲线、温度、pH值、氯仿和胆盐敏感性。通过透射电子显微镜观察噬菌体的形态。测试了ΦEP1在不同食品基质中的生物防治效果以及该噬菌体对Caco-2细胞的保护作用。噬菌体ΦEP1的最佳感染复数为0.1,效价为1.3×10 PFU/mL,对温度、pH、氯仿和胆盐具有较强的耐受性,宿主谱较广。此外,它对多种不同血清型的多重耐药病原菌具有裂解活性。噬菌体ΦEP1的潜伏期为10分钟,爆发期为80分钟,爆发量为48 PFU/细胞。根据透射电子显微镜观察到的形态,噬菌体ΦEP1属于 目,对Caco-2细胞具有良好的保护作用。噬菌体ΦEP1的基因组为87 182 bp,GC含量为39.80%,有128个推定的开放阅读框,无抗生素抗性基因或毒力基因。ΦEP1在人工污染的牛奶和牛肉中抑制了EIEC的生长,并在细胞保护实验中消除了EIEC。它显著提高了Caco-2细胞的存活率,并下调了白细胞介素(IL)-6和IL-1β的表达以减轻炎症。我们获得了一种效价高、环境耐受性强的靶向EIEC的噬菌体ΦEP1,为噬菌体在食品保鲜等领域的应用提供了依据。