冻存人血中分离的5株布鲁杆菌的生物学特征分析  

Biological characteristics of five strains of Brucella isolated in frozen human blood samples

作　　者：王锐泽 皮鑫 张翠[1] 刘新彬[1] 关超玲 赵倩[1] 赵硕[1] 李萌[1] 甄清[1] 

机构地区：[1]吉林大学公共卫生学院流行病与卫生统计学教研室,长春130021

出　　处：《中华地方病学杂志》2015年第11期805-807,共3页Chinese Journal of Endemiology

基　　金：国家自然科学基金青年基金（81102163）;吉林省卫生厅科研课题（20122057）

摘　　要：目的了解布鲁杆菌在冻存血液中的存活情况。方法在松原市疾病预防控制中心布鲁杆菌病（简称布病）门诊就诊的人群中，收集有发热症状及有流行病学接触史的患者血液28份（加入到柠檬酸钠抗凝管），-20℃保存。对冻存不同时间的血液样本进行布鲁杆菌分离培养，疑似菌落进行革兰染色及镜检，并与布鲁杆菌阳性血清、A因子、M因子进行血清凝集试验；采用多重PCR方法，使用布鲁杆菌属、种的引物对菌株进行鉴定。结果在冻存时间不同的28份血液中，有5份样本在分离培养后的第4～5天出现小菌落。肉眼下，菌落透明湿润或为乳白菌苔，革兰染色均为阴性，可与布鲁杆菌阳性血清、A因子、M因子血清发生凝集，初步判定这5株菌为羊种布鲁杆菌生物3型。5份分离出布鲁杆菌的血液样本冻存时间最长为17d，最短的6d。牛、羊种布鲁杆菌多重PCR结果均出现223bp目的条带，牛种还含有1条488bp的目的条带，羊种为310bp的目的条带，5株待测菌株的扩增结果与羊种布鲁杆菌的扩增结果一致。结论布鲁杆菌可以在冻存血液中存活一定时间．采用多重PCR方法，利用属、种鉴别，可以为布病的诊断与治疗提供依据。Objective To understand the survival situation of BruceUa in frozen human blood samples. Methods Blood samples of outpatients with brucellosis were from the Songyuan Center for Disease Control and Prevention. Twenty-eight blood samples of patients who had a symptom of fever and epidemiological contact history were collected, which were kept in tubes containing sodium citrate anticoagulation, and stored in the fridge at - 20 ℃. Isolation and culture of pathogen in frozen human blood samples were carried out. Suspected colonies were detected by Grams stain and microscope examination, then further tested by serum agglutination test with Brucella, A and M factors positive serum. The strains were detected with Brucella tautonym primers by multiplex-PCR amplification. Results In the twenty-eight blood samples frozen at different times, colonies were found in five samples after isolation and culture of pathogen for 4 - 5 d. The colonies were arranged in transparence, moist or milky lawn; the Grams staining was negative; serum agglutination test with Brucella, A and M factors positive serum were positive. Five strains of Brucella were preliminary considered as Brucella melitensis type 3. The freezing time of Brucella being isolated in five human blood samples was from 6 to 17 d. The results of multiplex-PCR showed that a band of 223 bp could be amplified from strains of Brucella abortus and Brucella melitensis, and that a band of 488 bp could be amplified from strain of BruceUa abortus, and 310 bp from Brucella melitensis. The results of five tested strains were identical with those of Brucella melitensis. Conclusion Brucella could be survived in frozen human blood samples for a certain time, and multiplex-PCR amplification with Brucella tautonym primers could provide a basis for diagnose and treatment of Brucella.

关 键 词：冻存血 布鲁杆菌 生物学 多重PCR 

分 类 号：R516.7[医药卫生—内科学]