福建省鼠感染钩端螺旋体调查和分离株基因种的鉴定

Infectious status of Leptospira in the rat population and genospecies identification of Leptospira strains isolated in Fujian Province

目的了解福建省鼠感染钩端螺旋体(钩体)现状及分离株基因种的特征,为钩体病防控提供科学依据。方法2015-2019年根据疫情选取14个调查点,采用笼夜法捕鼠并采集心血或肾脏标本进行钩体培养,测定血清抗体;对鼠中分离保存的23株钩体菌株提取DNA,PCR扩增16S rRNA基因、PCR产物纯化测序,与GenBank数据库收录的钩体17种基因型进行比对确定基因种,并用Mega 6.0软件构建系统进化树并进行序列分析。结果福建省鼠种以黄毛鼠为主要优势鼠种,占40.87%(499/1221)、其次依次是黄胸鼠19.57%(239/1221)、针毛鼠16.79%(205/1221)和褐家鼠8.85%(108/1221),钩体血清抗体阳性率为29.32%(207/706),以黄毛鼠最高,为42.16%(121/287),不同地区鼠种构成及鼠钩体感染率不同。23株菌株均扩增出16S rRNA基因的目标条带1484 bp,测序及序列分析属于致病性的基因种有13株,其中L.borgpetersenii(伯氏)占8株,L.interrogans(问号)占5株,属于非致病的基因种10株。钩体菌株分离自福建省的优势鼠种(黄毛鼠、黄胸鼠、针毛鼠)及不同地区(闽东、闽南、闽西和闽北)。结论福建省鼠类感染钩体普遍,以致病性基因种(L.borgpetersenii和L.interrogans)为主,也携带非致病性基因种,需加强钩体病传染源啮齿动物的监测工作。

This study aimed to explore the nature of infection and the genospecies of Leptospira in Fujian Province,to provide a scientific basis for the prevention and control of Leptospirosis.From 2015 to 2019,14 areas were selected according to the leptospirosis epidemic in Fujian.Rats were captured with the squirrel-cage method,and their sera and organs were collected for pathogen culture and antibody detection.DNA from 23 Leptospira strains isolated from the rats was extracted by alkaline lysis.The 16S rRNA genes were amplified with PCR and sequenced.These sequences were compared with those from 17 representative Leptospira strains in GenBank,and phylogenetic analyses were performed in Mega 6.0 software.The Rattus losea was the most frequent rodents,accounting for 40.87%(499/1221),followed by R.flavipectus 19.57%(239/1221),Niviventer fulvescens 16.79%(205/1221)and R.norvegicus 8.85%(108/1221),respectively.The rat serum antibody positivity rate was 29.32%(207/706),and the highest infection rates of Leptospira were detected in Rattus losea(42.16%,121/287).The distribution of rat species and infection rate varied among areas.The target bands(1484 bp)of the 16S rRNA gene for the 23 isolates were amplified and successfully sequenced.The BLAST results revealed the genospecies of 13 isolates included in the pathogenic clade.Eight isolates were most related to the genospecies L.borgpetersenii,and five isolates were related to the genospecies L.interrogans.The other ten isolates belonged to non-pathogenic species.The Leptospira strains were isolated from dominant mouse species(Rattus losea,R.flavipectus and Niviventer fulvescens)in different regions in Fujian Province and infected the dominant rodents.The genospecies of Leptospira strains isolated from rats had a ubiquitous distribution in various geographic areas in Fujian Province.They belonged mostly to pathogenic species(L.borgpetersenii and L.interrogans)and also were related to non-pathogenic species.Surveillance of rodents infected with Leptospirosis must be strengthened.