|MAHESHWARI, YOGITA - Foreign Agricultural Service (FAS, USDA)|
|SELVARAJ, VIJAY ANAND RAJ - Foreign Agricultural Service (FAS, USDA)|
|HAJERI, SUBHAS - Central California Tristeza Eradication Agency|
|RAMADUGU, CHANDRIKA - University Of California|
|Yokomi, Raymond - Ray|
Submitted to: Molecular Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/14/2017
Publication Date: 5/25/2017
Citation: Maheshwari, Y., Selvaraj, V., Hajeri, S., Ramadugu, C., Keremane, M.L., Yokomi, R.K. 2017. Novel development of a lateral flow immunoassay for rapid field detection of citrus tristeza virus. Molecular Biotechnology. doi: 10.1007/s12600-017-0591-0.
Interpretive Summary: Citrus tristeza virus (CTV) is a quarantined pathogen in California. Rapid detection and elimination is critical to limit spread of CTV which occurs by grafting and by aphid vectors. A user friendly, lateral flow immunoassay was developed for detection using technology similar to that used for human pregnancy testing with dipsticks. The lateral flow immunoassay was validated by comparison with two standard assays by assessing CTV infection in samples from 52 citirus trees. CTV was readily detected in infected field trees in the 10-minute test. As no specialized equipment is needed, the test may be used in the field or nursery by growers to assess CTV infection.
Technical Abstract: Maintenance of virus-free citrus in nurseries and orchards is essential to control spread of aphid-borne Citrus tristeza virus (CTV) in California. A lateral flow assay (LFA) test strip with a polyclonal antiserum made from virus particles produced in Nicotiana benthamiana plants inoculated with an infectious recombinant cDNA clone of CTV (rCTV) was developed. Affinity purified IgG to rCTV virions and Mouse IgG were conjugated with gold nanoparticles and coated onto a glass fiber conjugate pad. The test strip used CTV IgG (1mg/ml) and anti-mouse IgG (0.5 mg/ml) in the test line and control line, respectively. The LFA detected CTV within 10 minutes and was as sensitive as Enzyme-linked Immunoassay (ELISA). The LFA was tested with different strains of CTV such as the genotypes T30, T36, VT, and RB, from the greenhouse and field with 100% agreement with ELISA and Reverse Transcription quantitative Polymerase Chain Reaction assays. The LFA offered cost-effective detection of CTV in the field or nursery and was usable by non-skilled personnel without any laboratory equipment. Citrus plants identified by the LFA can be rogued to limit further spread.