摘要
目的从我国东北地区全沟硬蜱中分离新的伯氏疏螺旋体菌株,了解其基因型及分布。方法应用BSKH培养基从硬蜱中分离菌株,PCR扩增5S^23SrRNA基因间隔区,进行限制性片段长度多态性分析及序列测定。结果新分离出伯氏疏螺旋体25株,其中11株来自内蒙古自治区,13株来自黑龙江省,1株来自吉林省。RFLP结果显示其中22株属于Borreliagarinii基因型,3株属B.afazenii基因型,1株产生了新的酶切带型,序列测定表明其属于B.garinii。结论东北地区的伯氏疏螺旋体菌株为B.garinii和B.afzelii两种基因型,B.garinii为主要基因型。采用5S^23SrRNA间隔区RFLP分型方法对菌株进行分型研究时应结合序列分析方法,以保证分型结果的可靠性。
Objective To isolate Borrelia burgdorferi sensu lato（ B. b. s. 1. ） from ticks collected from Northeastern Forest Areas of China and to determine their genospecies and distribution. Methods BSKH medium was used to isolate B. b. s. 1. from ticks. 5S-23S rRNA intergenic spacer from all the isolates was amplified by polymerase chain reaction（PCR）. Restriction fragment length polymorphism（RFLP） method and sequencing analysis based on 5S-23S rRNA intergenic spacer were used for genotyping, identifying genotypes. Results A total of 25 strains were isolated, of which 11,13 and 1 were obtained from Great Xing-an Mountains forest in Inner Mongoha Autonomous Region, Small Xing-an Mountains forest in Heilongjiang province and Chang Bai Mountains in Jilin province, respectively. A target amplicon with 254 bp in size was generated from all 25 isolates. The 25 PCR products were digested using Mse Ⅰ and yielded 4 different RFLP patterns： B, C, D and N. Of the 4 patterns, N seemed to be a new one（a new RFLP pattern）. Among them, 21 isolates belonged to B. garinii and were divided into two different patterns： pattern B and C; 3 isolates belonged to B. arielii and fell into one pattern D; another isolate, named as VH10, show a unique pattern. The phylogenitic tree deduced from the sequences of all the 25 strains showed the isolates belonging to 4 branching groups. One branching group comprised 3 isolates：of B. afzelii , the others comprised 22 isolates belonged to B. garinii . All 3 B. afzelii isolates were obtained from Weihe area of Heilongjiang province. Twenty two B. garinii isolates were obtained from Inner Mongolia Autonomous Region, Heilongjiang province and Jilin province respectively. Conclusion In Northeastern Forest Areas of China, there are two genuspecies, and B. garinii is dominant one. In order to genotype the isolates accurately, RFLP analysis basing on 5S-23S rRNA intergenic spacer should be combined with sequence analysis after PCR assay.
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2006年第5期404-408,共5页
Chinese Journal of Microbiology and Immunology
基金
国家科技攻关计划课题(2003BA712A05-01)
