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Immunofluorescent Localization of Tobacco Ringspot Nepovirus in the Vector Nematode Xiphinema americanum

An indirect immunofluorescent technique was developed to localize tobacco ringspot nepovirus (TRSV) in the vector nematode Xiphinema americanum sensu stricto. A population of this nematode that efficiently transmitted TRSV was given an acquisition access period of 10 days on TRSV-infected cucumber. Treatment of fragments of viruliferous nematodes with a polyclonal antiserum against TRSV followed by fluorescein isothiocyanate-conjugated goat anti-rabbit immunoglobulin G resulted in virus-specific bright fluorescence only in the lumen of the stylet extension and esophagus. Virus-specific fluorescent signals were observed in the virus-retention region of 44% of the nematode fragments examined. The percentage of nematodes labeled with virus-specific fluorescence increased as the acquisition access period increased from 0 to 22 days; the increase paralleled the increase in the transmission efficiency of the nematode population. Visualization of the entire virus-retention region of individual nematodes within a population of vector or nonvector nematodes provides a rapid and simple means of monitoring specific attachment of plant viruses.