Delivery of maize mosaic virus to planthopper vectors by microinjection increases infection efficiency and facilitates functional genomics experiments in the vector

The corn planthopper, Peregrinus maidis, not only causes direct damage to plants by feeding, but also transmits maize mosaic virus (MMV) to the plant hosts. The virus is transmitted in a propagative manner but the acquisition of MMV by the vector feeding on infected plants can result in low acquisition and inoculation efficiency. Here, we increased the acquisition efficiency by delivering the virus directly into the hemocoel through microinjection, which resulted in efficient virus infection of the insect and transmission to maize. We found that delivery of virus by injection of 10 ng MMV (50 nl, 200 μg/ml virions) into P. maidis resulted in 93% transmission efficiency. In dose-response experiments, MMV abundance in insects and transmission efficiency decreased as the amount of virus inoculum delivered into the hemocoel was reduced. Examination of virus distribution in the vector using immunolabeling and confocal microscopy revealed similar tissue distributions in the injected insects when compared to those of previous studies using feeding on plants for virus acquisition. The utility of virus inoculation by microinjection for functional analysis in virus-vector interaction was explored. Co-microinjection of MMV virions and the dsRNA of PI3Kδ (a transcript that is less abundant in MMV-infected insects), resulted in a reduction in PI3Kδ expression and higher virus titers in P. maidis. These findings demonstrated that virus microinjection is a robust method for obtaining large numbers of infected planthoppers that are competent in transmitting MMV and, in combination with RNAi, could significantly facilitate the functional analysis of P. maidis-MMV interactions.