耐甲氧西林金黄色葡萄球菌裂解性噬菌体PhMASA-WJ的生物学特性及其对小鼠败血症的疗效

目的探讨耐甲氧西林金黄色葡萄球菌(methicillin-resistant Staphylococcus aureus,MRSA)噬菌体PhMASA-WJ的生物学特性及其对小鼠败血症的疗效,为开发高效低毒的抗菌药物奠定实验基础。方法以MRSA(USA300)为宿主菌,从地下污水中分离筛选出1株噬菌体PhMASA-WJ,双层琼脂平板培养法测定噬菌体效价,点滴法测定噬菌体裂解谱,确定最佳感染复数(multiplicity of infection,MOI),提取噬菌体全基因组并进行酶切鉴定,电镜观察噬菌体形态,并检测噬菌体的体外裂解效果、安全性及其对小鼠败血症的疗效。结果 PhMASA-WJ在USA300菌苔上形成直径1～2 mm的圆形、透明且边界清晰的噬菌斑,PhMASA-WJ效价为7. 1×10⁸ PFU/m L,其为专一裂解金黄色葡萄球菌的噬菌体,最佳MOI为0. 001,属于RNA病毒,基因组大小约23 000 bp。电镜观察显示,PhMASA-WJ属于有尾噬菌体目肌尾噬菌体科。动物实验证实,PhMASA-WJ对小鼠无毒副作用,败血症小鼠经PhMASA-WJ...

Biological characteristics of methicillin-resistant Staphylococcus aureus lytic phage PhMASA-WJ and its curative effect on septicemia in mice

Objective To investigate the biological characteristics of methicillin-resistant Staphylococcus aureus(MRSA)phage PhMASA-WJ and its curative effect on septicemia in mice so as to provide experimental basis for the development of high-efficiency and low-toxicity antibiotics.Methods Bacteriophages PhMASA-WJ was isolated and screened from sewage using MRSA(USA300)as host,and determined for titer by double layer agar plate culture and for splitting spectrum by dropping method,based on which the multiplicity of infection(MOI)was optimized.The whole genome of phage was extracted and identified by restriction analysis,then observed by electron microscopy and evaluated for lytic effect in vitro,safety and curative effect on septicemia in mice.Results PhMASA-WJ formed round,transparent and well-defined plaques at diameters of 1~2 mm on USA300 moss and a titer of 7.1×10^8 PFU/m L,which lyzed S.aureus specifically.At an optimal MOI of 0.001,PhMASA-WJ belonged to RNA virus,of which the whole genome was about 23000 bp in length.Electron microscopy showed that PhMASA-WJ belonged to Caudovirales myoviridae family.Animal experiments showed no toxic effect of PhMASA-WJ on mice.The survival rate of mice with septicemia after treatment with PhMASA-WJ was 60%,while all the mice in control group died,indicating that PhMASA-WJ improved the survival rate of mice significantly(P<0.001).Conclusion PhMASA-WJ showed strict host specificity,narrow splitting spectrum and strong splitting ability,which significantly improved the survival rate of mice with septicemia.It provided an experimental basis for clinical treatment of MRSA.