|RATTNER, RACHEL - Cooperative Agricultural Support Services|
|HAJERI, SUBHAS - Central California Tristeza Eradication Agency|
|GODFREY, KRIS - University Of California, Davis|
|Yokomi, Raymond - Ray|
Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 9/13/2021
Publication Date: 10/6/2021
Citation: Rattner, R., Hajeri, S., Godfrey, K., Yokomi, R.K. 2021. Recombinase polymerase amplification coupled with lateral flow assay for rapid field detection of “Candidatus Liberibacter asiaticus” in citrus plants. California Citrus Conference. Citrus Research Board.
Technical Abstract: Huanglongbing is a destructive citrus disease that affects production worldwide. “Candidatus Liberibacter asiaticus” (CLas), a phloem-limited bacterium, is associated with Huanglongbing in the United States. The current standard for CLas detection is realtime quantitative polymerase chain reaction (qPCR) using either CLas 16S rRNA gene- or ribonucleotide reductase (RNR)-gene specific primers/probes. qPCR requires well-equipped laboratories and trained personnel which is not convenient for rapid field detection of CLas. Recombinase Polymerase Amplification (RPA) assay is a fast, portable alternative to PCR. In this study, an RPA assay was developed to detect CLas in crude citrus extracts without the need for DNA purification. Primers were designed to amplify a region of the RNR gene of CLas by RPA analysis. The use of a fluorescent labeled probe allowed for detection of the amplicon in real-time. This assay detected a positive sample as early as in seven minutes. Additionally, the RPA assay was combined with a lateral flow device (LFD) for a point-of-use assay that is field deployable. This assay was specific to CLas and was sensitive to Ct 34 when compared with qPCR. This assay will be important for distinguishing CLas-infected trees in California from other pathogens that cause similar disease symptoms.