沙雷菌裂解性噬菌体PS2生物学特性及其全基因组测序

Characterization and genomic sequencing of PS2,a lytic phage that infects Serratia marcescens clinical isolates

目的明确一株粘质沙雷菌裂解性噬菌体PS2的生物学特性及全基因组序列。方法电镜观察PS2的形态;双层平板法分析PS2噬菌谱、生长曲线及理化稳定性;采用MiniBEST Viral RNA/DNA Extraction Kit提取噬菌体基因组,酶解作用鉴定基因组类型;构建噬菌体PS2测序文库,采用MiSeqTM系统测序,Velvet 1.2.08进行序列拼接。结果 PS2为肌尾型噬菌体。其感染S2菌株的潜伏期为21min,爆发量为70PFU。PS2在pH5到pH10的环境中具有较好的稳定性,在50℃和60℃的环境中活性也无明显变化。PS2基因组为双链DNA,全长167 266bp,平均GC含量为41.7%,共预测到276个ORF。BLASTN初步比对结果提示其与多株T4-like噬菌体基因组核酸序列具有显著相似性。结论分离鉴定了一株粘质沙雷菌属噬菌体,进行了生物学特性、全基因组测序和生物信息学初步分析,为噬菌体治疗多重耐药细菌及噬菌体生物信息学深入分析奠定了基础。

The present study focused on the characterization and genomic sequence of phage PS2 that infects Serratia marceseens clinical isolates. The morphology of phage PS2 was observed with electron microscope. The one-step growth curve, host range, and stability of PS2 were investigated. In addition, Phage DNA was extracted from the purified phage particles using a MiniBEST Viral RNA/DNA Extraction Kit. DNA sample was analyzed by digesting with restriction enzymes. The phage DNA was used for constructing the sequencing library. The library was sequenced on a MiSeqTM platform. The whole ge- nome sequence was obtained by Velvet （version： 1.2.08） assembling. Phage PS2 belongs to the Myoviridae family. The line- ar, circularly permuted, 167 266-bp double-stranded DNA genome of PS2 has high similarities to T4-1ike phages. The phage DNA contains 41.7% GC and 276 ORFs. PS2 exhibited a 21-minute latent period and 70 PFU per cell at burst size when the pathogenic S. marcescens strain S2 served as a host. Further investigation suggested that PS2 is stable in a wide pH range （pH5 to pill0） and at extreme temperatures （50 ℃ and 60 ℃） after incubation alone at different pHs and different temperatures, respectively. The paper focused on the isolation and identification of a novel lytic S. marcescens phage, the biolog- ical characteristics, the whole genome sequencing and the pre- liminary study of bioinformatics, which laid the foundation for deeply analysis to the phage therapy of multi-drug resist ant bacteria and the phage biological information.