1株梅花鹿源化脓隐秘杆菌的分离鉴定及其生物学特性分析

Isolation,identification and biological characteristics analysis of a strain of Trueperella pyogenes from sika deer

为确定重庆地区1株患病梅花鹿的致病菌,并探究其主要生物学特性,本试验采集病死梅花鹿肺脏,通过细菌分离、生化鉴定和16SrDNA测序对分离菌进行鉴定,随后对其致病性、毒力基因、药物敏感性以及生长曲线等主要特性进行分析。致病菌经鉴定为化脓隐秘杆菌,对小鼠具有较强的致病性,皮下和腹腔均能感染并产生典型症状,病死小鼠肝脏、脾脏、肺脏、肾脏产生坏死、出血和炎性细胞浸润为主要特征的组织病理学变化,对小鼠的半数致死量(LD50)为6.80×10^(10) CFU/mL,将其命名为YC-1;毒力基因检测显示,YC-1菌株具有2个神经氨酸酶(NanH、NanP)、2个菌毛蛋白(FimA、FimE)和1个溶血素(PLO)共5个毒力基因,其中PLO全长1605bp(GenBank编号OK513198),氨基酸进化树与霍加狓源化脓隐秘杆菌最为接近;药敏试验显示,YC-1对多种抗生素均敏感,耐药特征不明显;细菌的生长曲线显示在37℃,含有5%胎牛血清的TSB培养基中,培养6~10h为该菌的对数生长期。本试验解析我国梅花鹿源化脓隐秘杆菌主要生物学特性有助于深入研究该菌的致病机制以及开发有效的预防制剂。

In order to determine the pathogenic bacteria of a diseased sika deer in Chongqing and explore its main biological characteristics,the lung of dead sika deer was collected,and pathogenic bacteria were isolated.Identification was conducted by biochemical experiments and 16SrDNA sequencing.Then the main characteristics such as pathogenicity,virulence gene,drug sensitivity and growth curve were analyzed.Results showed that the pathogenic bacteria belonged to Trueperella pyogenes and had strong pathogenicity to mice.Typical symptoms could be produced by subcutaneous and intraperitoneal infection and the histopathological changes characterized by necrosis,hemorrhage and inflammatory cell infiltration were mainly found in the liver,spleen,lung and kidney of the dead mice.The LD50of the isolate to mice was 6.80×10^(10) CFU/mL,and it was named YC-1.Virulence gene detection experiments showed that YC-1had five virulence factors,including two neuraminidases(NanH,NanP),two pili proteins(FimA,FimE)and one hemolysin(PLO).The total length of PLO gene was 1605bp(GenBank No.OK513198).Phylogenetic tree indicated that okapi original Trueperella pyogenes was the closest to that of YC-1.Drug sensitivity test showed that YC-1was sensitive to a variety of antibiotics,and the drug resistance characteristics were not obvious.Growth curve of the bacteria showed that the logarithmic growth period of the bacteria was 6-10hin TSB medium containing 5%fetal bovine serum at 37℃.In summary,this is the first report of Trueperella pyogenes isolated from sika deer in China.Analyzing the main biological characteristics is benefit for further study of its pathogenic mechanism and develop effective preventive agents.