|VIDAL, E - VALENCIAN INSTITUTE FOR AGRICULTURAL RESEARCH|
|Yokomi, Raymond - Ray|
|MORENO, A - INSTITUTO DE AGRICULTURA|
|BERTOLINI, E - VALENCIAN INSTITUTE FOR AGRICULTURAL RESEARCH|
|CAMBRA, M - VALENCIAN INSTITUTE FOR AGRICULTURAL RESEARCH|
Submitted to: International Organization of Citrus Virologists Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: 10/22/2010
Publication Date: 11/7/2010
Citation: Vidal, E., Yokomi, R.K., Moreno, A., Bertolini, E., Cambra, M. 2010. Tissue-print real-time RT-PCR for accurate detection of Citrus tristeza virus. Validation and comparison with Tissue print-ELISA [abstract]. International Organization of Citrus Virologists Proceedings. 31:s97.
Technical Abstract: Citrus tristeza virus (CTV) causes one of the most important virus diseases of Citrus species. The control of CTV in Spain and Central California is based on planting virus-free citrus on CTV-tolerant or -resistant rootstocks. However, quarantine and certification programs are still essential to avoid importation and propagation of severe strains of CTV. There is a zero tolerance of CTV in nursery blocks in Spain and Central California. These programs need sensitive, specific and reliable CTV-detection methods. The use of PCR-based procedures for diagnosis of plant pathogens have been developed, nevertheless, all new methods must be validated by comparison with the accepted standard technique. Thus, the tissue-print real-time RT-PCR was compared with the validated method of tissue print-ELISA with 1,395 samples from mature citrus trees and nursery plants. The total agreement between techniques was significant (P < 0.01) with a Cohen’s kappa index of 0.77 ± 0.03. The reliability of each technique was estimated using a nursery plot of 658 C. macrophylla seedlings in December 2008. Plants with discordant results were retested in July 2009 by the two methods and by indexing in Mexican lime. A plant was considered infected by CTV when a positive test was observed by at least by two of the three techniques used. The estimate for tissue print-ELISA for sensitivity was 0.8015 with a specificity of 0.9962 and a positive and negative likelihood ratio of 216.42 and 0.199, respectively. The estimate for tissue-print real-time RT-PCR for sensitivity was 0.982 with a specificity of 0.8519 and a positive and negative likelihood ratio of 6.63 and 0.021, respectively. These results show tissue-print real-time RT-PCR was more sensitive than tissue-print ELISA and validates its use to detect CTV for diagnostic purposes.