甘蔗黄叶病毒病早期快速RT-PCR诊断和鉴定

DIAGNOSIS AND IDENTIFICATION OF PRE-SYMPTOMS OF SUGARCANE YLS WITH RT-PCR AND DAS-ELISA

甘蔗黄叶病毒病 (YLS)是一种由黄症病毒 (Luteovirus)引起 ,随种蔗传带的新病害。利用由黄症病毒组特征片段克隆测序设计的特异性引物对YLSR 4 62和YLSF 111开发出一种可以早期快速检测无症甘蔗繁殖材料带毒情况的反转录多聚酶链式反应技术 (RT -PCR)。对来自世界各地和美国甘蔗主产区的 14 0个甘蔗品种及育种材料进行了检测 ,黄症叶片样品RT -PCR阳性检出率为 10 0 % ,而无症样品带毒率为 5 5 .17%。与DAS -ELIS相比 ,显症和无症样品的检测符合度分别为 76.4 7%和 5 6.2 4 % ,表明分子诊断较血清学方法灵敏。而单纯采用叶片糖分锤度单一指标诊断YLS是不可靠的。美国的主栽甘蔗品种和大部分育种材料 (95 .0 % )对于YLS均表现感病或带毒。

A novel RT-PCR approach based on the specific upstream/downstream primers of YLSR 462 & YLSF111 was developed for detecting asymptomatic yellow leaf syndrome (YLS) of sugarcane breeding lines and commercial varieties. Of the 140 parental and commercial clones from Louisiana and Texas of USA and other parts of the world, only six were YLSLV (YLS Luteovirus) free as detected by RT-PCR and DAS-ELISA. Higher Brix readings were associated with symptom expression (8.4) and a high level of YLS (100%), and lower average Brix reading (4.2) often involved asymptomatic expression and a lower level of YLS detection by both RT-PCR and DAS-ELISA. Some (55.17%) of asymptomatic crossing progenies/cultivars with lower Brix readings as assayed appeared to be YLS positive, indicating that Brix readings alone are not reliable enough for YLS detection. RT-PCR proved more sensitive than DAS-ELISA for detecting pre-symptom sugarcane leaf of YLS. Ninety-five percent of the crossing lines and varieties of USA used in this experiment were susceptible to YLSLV.