|Yokomi, Raymond - Ray|
|SELVARAJ, VIJAYANADRAJ - Foreign Agricultural Service (FAS, USDA)|
|MAHESHWARI, YOGITA - Foreign Agricultural Service (FAS, USDA)|
|RATTNER, RACHEL - Cooperative Agricultural Support Services|
|GODFREY, KRISTINE - University Of California, Davis|
|HAJERI, SUBHAS - Central California Tristeza Eradication Agency|
Submitted to: Citrograph
Publication Type: Trade Journal
Publication Acceptance Date: 1/7/2021
Publication Date: 3/16/2021
Citation: Yokomi, R.K., Selvaraj, V., Maheshwari, Y., Rattner, R., Godfrey, K.E., Chen, J., Hajeri, S. 2021. Improvements to HLB and stubborn testing. Citrograph. 12(2):64-68.
Technical Abstract: Huanglongbing (HLB) control in California is based, in part, on testing citrus trees and eradicating ‘Candidatus Liberibacter asiaticus’ (CLas)-positive trees. This is complicated by the presence of Spiroplasma citri, which is the causal agent of citrus stubborn disease (CSD), since CSD symptoms can resemble those of HLB. These trees will test negative for CLas, but this may raise concern that this is a false negative result. The objective of this study was to develop a protocol to detect and differentiate both pathogens in a single test. Using specific primers and different fluorescent probes for each pathogen, a duplex qPCR assay was developed that detects and differentiates both pathogens simultaneously. Because conventional qPCR assays provide only a relative quantity of the pathogen target, a duplex droplet digital (ddPCR) assay was developed that measures the exact number of DNA targets in the sample and can serve as a confirmation test. These tests can be used by regulators and diagnosticians to improve reliable detection of CLas. In addition, whole genome sequencing of seven strains of S. citri were determined and examined for new gene targets to improve detection of S. citri.