ARS | Publication request: Using Real-Time PCR to Quantify Aster Yellows Phytoplasma in its Insect Vector; Relationship of Infectivity to Transmissibility in the Aster Leafhopper

Submitted to: American Phytopathological Society Abstracts
Publication Type: Abstract Only
Publication Acceptance Date: March 20, 2009
Publication Date: July 31, 2009
Citation: Frost, K.E., Willis, D.K., Groves, C.L., Groves, R.L. 2009. Using Real-Time PCR to Quantify Aster Yellows Phytoplasma in its Insect Vector; Relationship of Infectivity to Transmissibility in the Aster Leafhopper [abstract]. American Phytopathological Society Abstracts. 99:S38.

Technical Abstract: The aster yellows (AY) index is used to prescribe insecticide sprays that target Macrosteles quadrilineatus, or aster leafhopper (ALH), the vector of the aster yellows phytoplasma (AYp). The AY index metric is the product of the proportion of infective ALHs and the relative ALH population size at a location. PCR is now used to determine the percentage of AYp-infected ALHs but there is a discrepancy between insects that are infected and those that are infective which can lead to undue chemical applications. We hypothesize that ALH infectivity is related to the amount of AYp in insect (titer) and developed a quantitative real-time PCR (qPCR; SYBR Green) method to quantify the amount of AYp DNA its insect host. Elongation factor TU (tuf) gene and valine-tRNA ligase (valS) were used as targets for AYp and leafhopper CP6 wingless gene was used as the target for insect DNA amplification. Targets were chosen because they are (likely) present as single copies in their respective genomes. The amount of AYp target DNA was measured relative to the amount of insect target DNA to avoid influences of the DNA extraction procedure. To date, single insect extracts that have been quantified have ranged in titer from 0.006 to 0.832 AYp genomes to insect genomes. These values correspond to x copies per ng of insect of leafhopper DNA. The distribution of ratios was right-skewed meaning ALHs with low AYp titer occur more frequently than those with high titer. Using this method we plan to elucidate the dose-response relationship between ALH AYp-titer and leafhopper infectivity to improve the accuracy of the AY index.