Empleo de plantas de pepino como amplificador biológico para la detección del viroide del enanismo del lúpulo (HSVd) en cítricos

Hop stunt viroid (HSVd) is the etiological agent of citrus cachexia and it can cause considerable losses in this crop. In Cuba, HSVd has been reported occurring in a high percentage of plants in different citrus areas, and works concerning its detection have been mainly based on biological assays and their combination with nucleic acid analysis by polyacrilamida gel electrophoresis and nucleic acid hybridization. These methods involve a previous step of inoculating a citrus biological amplifier before nucleic acids can be analyzed. It imposes a restrictive requirement of incubation for at least three months. The objective of the present work was to develop a new methodology for detecting HSVd in citrus plants using cucumber plants (Cucumis sativus L. var. Japanese) as an intermediate biological amplifier that requires a shortest incubation time than the amplifier presently used without affecting its effectiveness. Positive citrus plants controls from field were evaluated and compared with reference methods. The results defined a methodology for HSVd diagnosis in citrus where the intermediate biological amplification in cucumber plants is combined with detection by nucleic acid hybridization. This methodology showed an effectiveness of 98.9% and required a replication time of the viroid in the plant of 14 days, shorter than that presently required. Its use could diminish the time of decision taken of control measures and the risk period, both aspects that are of great importance for the specialized services that guarantee the propagation material phytosanitary quality.