耐碳青霉烯类鲍曼不动杆菌裂解性噬菌体PA103的生物学特性及全基因组分析  

Biological characteristics and whole genome analysis of carbapenem-resistant Acinetobacter baumannii lyticphagePA103

作　　者：张改 靳静 陈松建 王书伟 李振江 刘肖 李亚辉 张欢欢 王山梅[2] ZHANG Gai;JIN Jing;CHEN Songjian;WANG Shuwei;LI Zhenjiang;LIU Xiao;LI Yahui;ZHANG Huanhuan;WANG Shanmei(Pathogenic Biology and Immunology Teaching and Research Office of Henan Medical College,Zhengzhou 451191,China;Laboratory of Henan Provincial People's Hospital)

机构地区：[1]河南医学高等专科学校病原生物学与免疫学教研室,河南郑州451191 [2]河南省人民医院检验科

出　　处：《中国病原生物学杂志》2024年第10期1182-1188,1193,共8页Journal of Pathogen Biology

基　　金：河南省高等学校重点科研项目(No.23B310012);河南省科技厅科技发展计划项目(No.222102310449)。

摘　　要：目的 探讨耐碳青霉烯类鲍曼不动杆菌(carbapenem-resistant Acinetobacter baumannii,CRAB)噬菌体PA103的生物学特性及全基因组信息。方法 以CRAB临床菌株A103为宿主菌,采用双层平板法在市政污水中分离得到1株裂解性噬菌体PA103;通过双层平板法观察噬菌斑形态;通过透射电镜观察噬菌体形态特征;调查PA103的噬菌谱、一步生长曲线及对温度和酸碱的稳定性;应用蛋白酶K/SDS法提取噬菌体基因组DNA;对PA103进行基因组测序、拼装;用RAST在线预测软件对基因组中可能存在的开放读码框进行预测;用MEGA 11.0软件以末端酶大亚基构建进化树,用Mauve软件进行比较基因组学分析。结果 试验分离得到1株可以裂解CRAB的裂解性噬菌体,命名为PA103;它可以在宿主菌A103的菌苔上形成直径约2 mm完全透明的噬菌斑,周围环绕宽为3~4 mm的半透明晕环;透射电镜显示PA103属于有尾噬菌体,头部和尾部直径长度约64 nm和120 nm;一步生长曲线显示PA103的潜伏期约为40 min,爆发期约为15 min,爆发量约为17 PFU/cell;噬菌体PA103在50、60℃,pH 5.0~9.0的环境下,均能保持相对稳定的活性;PA103的基因组全长45 225 bp, G+C含量为37.82%,全基因组预测含86个开放阅读框,其中29个为功能蛋白,其余57个被注释为假定蛋白;共线性分析结果显示噬菌体PA103与噬菌体LZ35的局部共线性区域的数目、方向、排列顺序和长度非常相似;进化树构建结果显示噬菌体PA103应属于Obolenskviru属成员。结论 新型噬菌体PA103具有较强的鲍曼不动杆菌裂解能力,可为噬菌体防治耐碳青霉烯类鲍曼不动杆菌的感染提供参考。Objective To investigate the biological characteristics and whole genome information of carbapenem-resistant Acinetobacter baumannii(CRAB)phage PA103.N Methods Using CRAB clinical strain A103 as the host bacterium,a lytic phage PAl03 was isolated from municipal wastewater using a double-layer plate assay technique;Observe the morphology of plaques using the double-layer plate assay technique;Observe the morphological characteristics of phages through transmission electron microscopy;Investigate the phage spectrum,one-step growth curve,and stability to temperature and acidity/alkalinity of PA103;Extract phage genomic DNA using proteinase K/SDS method;Perform genome sequencing and assembly on PA103;Use RAST online prediction software to predict potential open reading frames in the genome;Construct an evolutionary tree using MEGA 1l.O software with terminal enzyme subunits,and perform comparative genomics analysis using Mauve software.ResultsA lytic phage capable of cleaving CRAB was isolated from the experiment and named PA103;It can form a wholely transparent plaque with a diameter of about 2 mm on the bacterial coat of host bacterium A1o3,surrounded by a translucent halo with a width of 3-4 mm;Transmission electron microscopy showed that PA103 belongs to a tail bacteriophage,with a head and tail diameter length of approximately 64 nm and 120 nm,respectively;The one-step growth curve shows that the latent period of PAi03 is about 40 minutes,the burst period is about 15 minutes,and the burst size is about 17 PFU/cell;Phage PA103 can maintain relatively stable activity in environments of 50 C,60 C,and pH 5.0-9.0;The genome of PA103 has a total length of 45225 bp and a G+C content of 37.82%.The whole genome prediction contains 86 open reading frames,of which 29 are functional proteins and the remaining 57 are annotated as hypothetical proteins;The results of collinearity analysis showed that the number,direction,arrangement order,and length of local collinear regions between phage PA103 and phage LZ35 were very simil

关 键 词：耐碳青霉烯类鲍曼不动杆菌 噬菌体 生物学特性 基因组分析 

分 类 号：R378[医药卫生—病原生物学]