重庆地区蜜蜂微孢子虫的鉴定及分子遗传多样性分析

Identification and Genetic Diversity of Microsporidia Nosema ceranae Isolated from Chongqing Based on Multilocus Molecular Markers

采用PCR扩增、克隆与测序的方法,对重庆东南地区引起意大利蜜蜂(意蜂)蜂群出现明显下降趋势的病原微孢子虫的遗传标记基因SSU r DNA(小亚基核糖体脱氧核糖核酸,small subunit ribosomal DNA)和ITS(内转录间隔区,internal transcribed spacer)以及3个单拷贝座位分子标记肌动蛋白基因(Actin),热激蛋白基因(Hsp70)和RNA聚合酶基因(RPB1)进行分子水平扩增及测定,并与国内外不同地理分布的蜜蜂微孢子虫种群进行了分子遗传多样性比较及单倍型分析。结果显示,该蜜蜂微孢子虫分离种群为东方蜜蜂微孢子虫,命名为CQYX。序列分析表明东方蜜蜂微孢子虫重庆分离群与其它地理种群之间的遗传多样性水平上无差异性。高频单倍型为重庆CQYX地理群与其他不同地理种群所共有,并分化成不同的低频种群特异性单倍型,表明重庆CQYX种群和其他不同地理来源的东方蜜蜂微孢子虫种群具有共同的遗传起源,并经历了近期的快速扩增。本研究有助于进一步在分子水平上探讨意蜂中东方蜜蜂微孢子虫的地理起源。

Using PCR amplification,cloning and sequencing,the multilocus molecular markers,including small subunit ribosomal DNA（ SSU r DNA）,internal transcribed spacer（ ITS）,Actin,Hsp70 and RPB1,were obtained from microsporidia-like pathogen isolated from sick Apis mellifera with population decline in a apiary of Chongqing Southeast area. The genetic diversity and haplotypes of population based on these markers were analyzed by comparing the Chongqing isolates with other geographical isolates. The results showed that this microsporidia-like isolate from Apis mellifera of Chongqing apiary was Nosema ceranae,which then named as CQYX isolate. Further sequence analysis suggested that the level of genetic diversity couldnot be distinguished distinctly between CQYX and other different geographical isolates. Most haplotypes were derived from a frequent haplotypes and always not shared across isolates,suggesting that CQYX and other geographical isolates should have a common origin and undergo population expansion. This study would make contribution to further study the geographical origins of N. ceranae that parasitize Apis mellifera.