大肠杆菌O157∶H7 CVa9株O抗原变异的研究

Study on O antigen variation of Escherichia coli O157:H7 CVa9 strain

目的 研究本实验室保藏的大肠杆菌O15 7:H7(E coliO15 7:H7)日本分离株CVa9O抗原的变异。方法 血清凝集试验检测CVa9菌株O和H抗原 ,区分O抗原变异株与非变异株。聚合酶链反应 (PCR)法分别检测O15 7和H7特异性基因。生化试验检测生化特性。观察菌株在麦康凯、山梨醇麦康凯、棉子糖麦康凯及ChromagarO15 7显色培养基上菌落形态 ,菌落计数法计算变异率。结果 抗 -O15 7抗血清与CVa9菌株进行玻片凝集试验出现强凝集 (CVa9- 8)和不凝集 (CVa9- 1、CVa9- 15 )两种菌落 ,试管凝集试验结果相同 ,证实不凝集菌落O抗原发生变异。抗 -HT 抗血清分别与变异株和非变异株H抗原进行试管凝集试验 ,均为强凝集。两种菌株O15 7和H7特异性基因均为阳性。变异株与非变异株生化特性基本相同 ,但变异株不发酵棉子糖。两种菌株在麦康凯及山梨醇麦康凯培养基上菌落形态特征没有区别。在ChromagarO15 7显色培养基上培养 4 8h ,变异株为浅紫红色 ,菌落周边呈毛边状 ,非变异株为紫红色 ,菌落边缘整齐。变异株在用棉子糖代替乳糖制备的棉子糖麦康凯培养基上菌落呈粉红色 ,非变异株呈红色。菌落计数显示变异率为 99 80 %。结论 CVa9菌株在保存过程中O抗原发生变异 ,菌落形态及部分生化特征亦有所改变。

Aim To detect the antigen variation of preserved E coli CVa9 strain isolated from Japan Methods Serologic agglutination assay was used to detect O and H antigens and to distinguish the O antigen variant and non variant strains Polymerase chain reaction was used to identify specific O157 and H7 genes Biochemical characteristics were tested with biochemical assay Bacterial colonies were observed on MacConkeyagar,Sorbitol MacConkey agar,raffinose MacConkey agar and Chromagar O157,counting bacterial colony and calculating variant rate Results Two kinds of colonies of strong agglutination(CVa9 8)and non agglutination(CVa9 1?CVa9 15)were observed after the reaction of O157 anti serum and CVa9 strain,both in tube agglutination assay and tube agglutination,which indicated that O antigen varianted in non agglutination colony Both H antigens of variant and non variant strain gave strong agglutination after reaction with H7 antiserum in tube agglutination assay Specific genes of O157 and H7 were positive for both strains Their biochemical characters were similar except that the variant strain didn't ferment raffinose No difference were found for the colony characters of two strains when cultured in MacConkey and sorbitol MacConkey agar After incubated in Chromagar O157 for 48 hours,variant strain became light purple with non uniform surroundings,while non variant strain,the color was purple with uniform surroundings The variant strain presented pink and the non variant strain became red when cultured in raffinose MacConkey agar in which lactose was replaced by raffinose Colony counting showed that the variant rate was 99 80% Conclusions O antigen variant occurrs during preservation,changes also in colony form and some biochemical characteristics Both strains carry specific O157 and H7 genes,the reason of loss of express of O157 antigen is under investigated