Rapid On-Site Detection of the Huanglongbing/Citrus Greening Causal Agent 'Candidatus Liberibacter as i a tic us' by Amplify RP~R, a Novel Rapid Isothermal Nucleic Acid Amplification Platform

摘要

Huanglongbing (HLB) has become the most serious threat to the citrus industry worldwide. Although the disease first emerged in the southeastern part of Asia, it is now well established in Brazil, Florida and California, which are the world's most important citrus production areas. The disease is associated with three insect-vectored, obligate intracellular bacteria of the genus Candidates Liberibacter'. These bacteria reside in the host phloem cells and they are transmitted by psyllids. Early pathogen detection is important to protect free areas from pathogen introduction. Unfortunately, the disease has a long incubation period, during which the pathogen is at low concentration and non-systemically distributed. This makes diagnostic and eradicationdifficult. Agdia has recently developed a new, rapid, molecular detection technique, AmplifyRP~R, allowing PCR-level sensitivity detection within minutes, in the field. AmplifyRP~R uses a recombinase-polymerase methodology for DNA amplification at a single temperature. In contrast to conventional or real-time PCR, AmplifyRP~R has no DNA purification requirements, requires no thermocycling, and results can be read using small, user-friendly devices. A portable fluorescence reader or a lateral flow device (similar to Agdia's ImmunoStrip~R) can be used to visualize results in as little as 30 minutes, compared to several hours for conventional PCR. AmplifyRP~R eliminates the need for expensive PCR equipment, a large number of chemicals reagents, and the need for technically trained staff. Given its characteristics, AmplifyRP~R is the ideal tool to monitor HLB progression by early detection of Ca. Liberibacter in citrus trees or in vector insects.