Abstract

Bacterial ghosts (BGs) are gram-negative bacteria cell membranes without cytoplasmic content obtained by lysis as a result of cell-wall perforation mediated by the φX174 bacteriophage E protein. Introducing into a lytic plasmid lysis gene from other bacteriophages, which target alternative molecular targets, in addition to the E protein gene of bacteriophage φX174, can lead to an increase in the efficiency of lysis and, in some cases, to an increase in the immunogenicity of preparations. Escherichia coli strains carrying plasmids with various combinations of genes encoding protein E of phage φ174 with cassettes of lytic genes of the “choline–endolysin” systems of phages λ or L-413C, providing different degrees of destruction of the cell wall, were obtained for the subsequent selection of the most promising lytic structures. The formation of E. coli ghosts and the release of cell contents was confirmed by transmission electron microscopy. When the cultivation temperature was increased from 28 to 42°C, the lysis of E. coli cultures DH5α/pEYR'-E, DH5α/pEYR'-Y-K, DH5α/pEYR'-E-Y-K, and DH5α/pEYR'-E-Sam7-R-Rz was observed. Lysis was practically not detected during the growth of the DH5α/pEYR'-S-R-Rz culture, as in the control strain DH5α/pEYR'. The results of the analysis of ultrathin sections of BGs preparations by transmission electron microscopy (TEM) were comparable with the data on optical density and the results of inoculation of induced cultures of engineered strains, and also made it possible to assess the fine structure of bacterial cells carrying various combinations of lytic phage genes. BGs of gram-negative bacteria are promising for the creation of highly effective inactivated candidate vaccines that could replace currently available bacterial heat-inactivated and formol vaccines.

中文翻译：

---

噬菌体裂解酶在细菌幽灵生成过程中的效率

摘要

细菌影 (BGs) 是革兰氏阴性细菌的细胞膜，由于 φX174 噬菌体 E 蛋白介导的细胞壁穿孔，通过裂解获得细胞质内容物。将其他噬菌体的裂解基因引入裂解质粒裂解基因，除了噬菌体 φX174 的 E 蛋白基因外，这些噬菌体还靶向替代分子靶标，可导致裂解效率增加，在某些情况下，免疫原性增加准备工作。大肠杆菌携带质粒的菌株具有编码噬菌体 φ174 蛋白 E 的基因的各种组合，以及噬菌体 λ 或 L-413C 的“胆碱-内溶素”系统的裂解基因盒，提供不同程度的细胞壁破坏，用于随后的选择最有前途的裂解结构。大肠杆菌幽灵的形成和细胞内容物的释放通过透射电子显微镜证实。当培养温度从 28°C 升高到 42°C 时，大肠杆菌的裂解观察到培养物 DH5α/pEYR'-E、DH5α/pEYR'-YK、DH5α/pEYR'-EYK 和 DH5α/pEYR'-E-Sam7-R-Rz。在 DH5α/pEYR'-SR-Rz 培养物的生长过程中几乎没有检测到裂解，就像在对照菌株 DH5α/pEYR' 中一样。通过透射电子显微镜 (TEM) 对 BGs 制备物的超薄切片进行分析的结果与光密度数据和工程菌株诱导培养物的接种结果具有可比性，也使得评估细菌的精细结构成为可能携带多种裂解噬菌体基因组合的细胞。革兰氏阴性菌 BGs 有望用于创建高效灭活候选疫苗，可以替代目前可用的细菌热灭活疫苗和甲醛疫苗。