粪肠球菌噬菌体vB_EfaP_IME195的生物学特性及其全基因组分析

Characterization and genome analysis of a novel bacteriophage vB_EfaP_IME195 infecting Enterococcus faecalis

摘要:

【目的】以粪肠球菌为宿主菌，从医院的污水中筛选出相应的粪肠球菌噬菌体vB_EfaP_IME195，简称IME195，研究其生物学特性；并通过高通量测序得到其全基因组，深入研究其基因组学特征。【方法】以临床的耐药粪肠球菌为宿主菌，利用医院污水筛选噬菌体并纯化；对噬菌体IME195生物学特性进行了深入研究，包括电镜观察噬菌体形态、最佳感染复数、一步生长曲线、噬菌体IME195对紫外线的敏感度、对温度的耐受程度、对pH的耐受程度、对氯仿是否敏感；通过蛋白酶K/SDS法提取噬菌体IME195全基因组；Ion Torrent高通量测序；测序后进行噬菌体全基因组序列组装、注释、进化分析和比较分析。【结果】通过噬菌体梯度稀释，双层培养基平板法得到噬菌斑边缘分明、斑体透明的裂解性噬菌体IME195，最佳感染复数为0.01，一步生长曲线显示IME195的潜伏期为30 min，暴发量为11。该噬菌体对紫外线比较敏感，对5%浓度的氯仿不敏感，噬菌体对高温比较敏感，该噬菌体在pH 6.0?8.0范围内具有良好的裂解活性；电镜观察结果显示该噬菌体属于尾病毒目短尾噬菌体科；全基因组分析表明：噬菌体IME195基因组大小只有18 607 bp (GenBank登录号为KT932700)，G+C含量仅为33%。Blastn比对结果表明，该噬菌体和GenBank中的噬菌体vB_Efae230P-4只有82%的相似性。对噬菌体IME195进行了全基因组功能注释和进化分析。【结论】分离鉴定了一株粪肠球菌噬菌体，进行了生物学特性、全基因组测序和生物信息学深入分析，为噬菌体治疗多重耐药细菌奠定了基础。

Abstract:

[Objective] Multidrug-resistant strains of Enterococcus faecalis are becoming increasingly common worldwide. Therefore, we aimed to find novel agents to control antibiotic-resistant E. faecalis. We isolated a lytic bacteriophage from hospital sewage. [Methods] Phage morphology was observed using transmission electron microscopy (TEM), and its genome was sequenced using the Ion Torrent sequencing platform. Genome annotation and comparative and evolutionary analyses were performed following assembly of the complete genome sequence. [Results] Lytic bacteriophage vB_EfaP_IME195 was successfully isolated from infected E. faecalis host cells. TEM analysis indicated that the isolated bacteriophage resembled members of the Podoviridae family. The complete genome of vB_EfaP_IME195 was 18 607 bp long, and was comprised of circular dsDNA (GenBank accession No. KT932700). The genome had a G+C content of 33%, and contained only 27 coding sequences. BLASTn analysis showed that vB_EfaP_IME195 shared the highest homology (82%) with E. faecalis phage vB_Efae230P-4. [Conclusion] The isolation and characterization of this novel lytic E. faecalis phage provides the basis for potential treatment alternatives for multidrug-resistant E. faecalis infection.