Microneedle assisted delivery of the cloned components of cucurbit leaf crumple virus in yellow squash (Cucurbita pepo)

Cucurbit leaf crumple virus (CuLCrV) is among the prominent viruses infecting cucurbits in the USA. Attainable procedures of virus inoculation to crops are prerequisite for screening of resistance against the virus. Because mechanical (non-vector-mediated) infection by cucurbit leaf crumple virus (CuLCrV) is inefficient in economically important crops, screening for CuLCrV resistance is currently laborious and time-consuming using transmission by viruliferous whiteflies. We constructed an infectious partial tandem repeat construct of an isolate of CuLCrV from Georgia, USA, in the plant expression binary vector pCambia2300 and transformed it into Agrobacterium tumifaciens strain EHA105. Agroinfiltration of this construct into the abaxial surface of the leaves of common bean (Phaseolus vulgaris L.) produced a systemic infection characteristic of CuLCrV, although this approach was not successful for yellow squash. However, we report a very efficient and reproducible inoculation procedure established in squash when the leaves were injured with a microneedle and rubbed it with cell suspension harbouring the infectious viral construct.