柑橘黄龙病病原菌的鉴定及16S rDNA序列分析

Identification and 16S rDNA Sequence Analysis of Citrus Huanglongbing Pathogen

采用Nested-PCR技术检测顺昌果园红心柚树中柑橘黄龙病病原的携带情况,通过黄龙病病原菌16S rDNA测序和构建系统发育树比较不同类型柑橘黄龙病原菌之间的进化关系,采用特异引物扩增16S rDNA片段鉴别不同黄龙病病原菌,结合限制性内切酶酶切确定该红心柚园黄龙病病原的类型,并分析不同类型柑橘黄龙病病原16S rDNA序列差异性。结果表明:所取的12株红心柚树叶片样品中有7株树的叶片中检测出黄龙病病原,黄龙病病原的阳性检出率为58.33%,其中4个为显性性状,3个为隐性性状。柑橘黄龙病病原菌16SrDNA进化分析及限制性内切酶XbaI酶切结果表明,顺昌柑橘黄龙病病原菌属于亚洲型。不同类型柑橘黄龙病病原菌16SrDNA的序列差异性分析表明,亚洲种与非洲种的差异性为2.75%,非洲种与美洲种的差异性为3.90%,亚洲种与美洲种的差异性为3.44%。

Nested-PCR detection of Shunchang citrus Huanglongbing pathogen was carried out in this study.Phylogenetic tree based on rDNA sequencing was built to find out the evolutionary between different parts citrus HLB pathogens.Digesting the target DNA fragment of 16S rDNA with restriction enzyme XbaI was carried out to confirm the species of Shunchang Huanglongbing pathogen.The results showed that there were 7 positive detections in 12 selected samples,and the rate of positive detections was 58.33%,and four were dominant character,three were recessive character.The results showed that Shunchang citrus Huanglongbing pathogen belong to Candidatus Liberibacter asiaticus based on the 16S rDNA phylogenetic analysis and restriction enzyme XbaI digestion analysis.And 16S rDNA sequence gap analysis showed that the difference between Ca.L.asiaticus and Ca.L.africanus,Ca.L.africanus and Ca.L.americanus,Ca.L.americanus and Ca.L.asiaticus were 2.75%,3.90%,3.44%,respectively.