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POSTERS: Pathogen detection, quantification and diagnosis

Targeted early detection of citrus Huanglongbing causal agent ‘Candidatus Liberibacter asiaticus’ before the symptoms expression
Sheo Shankar Pandey - Citrus Research and Education Center, University of Florida. Nian Wang- Citrus Research and Education Center, University of Florida

Citrus Huanglongbing (HLB) is the most severe disease of citrus plants caused by Candidatus Liberibacter asiaticus (Las) and transmitted by the insect vector Asian citrus psyllid (ACP). To date, no effective curative measure is available against HLB. For citrus production areas without HLB or with low HLB disease incidence, removal of Las inoculum is critical to prevent HLB spread. Such strategy requires robust early diagnosis of HLB for inoculum removal to prevent ACP acquisition and transmission of Las. However, early diagnosis of HLB is challenging because the citrus trees remain asymptomatic for several months to years after Las transmission by ACP. In this study, we report a new method for a targeted early detection of Las in Valencia sweet orange before HLB symptom expression. We take advantage of the fact that Las remains around the ACP feeding site immediately following transmission into the young flush and before flush maturation. The ACP secretes salivary sheaths at their feeding sites, which can be visualized using Coomassie brilliant blue staining due to the presence of salivary sheaths secreted by ACP. Epifluorescence and confocal microscopy indicate the presence of salivary sheaths beneath the blue spots on ACP-fed-leaves. Quantitative real-time PCR and conventional PCR assays are able to detect Las in the ACP feeding surrounding areas as early as 2-20 days after ACP feeding. This finding lays a foundation to develop much-needed tools for early diagnosis of HLB before symptom expression, thus assisting Las inoculum removal and preventing HLB from spreading.