Detection of Citrus Psorosis Virus by dsRNA and molecular hybridization

Citrus psorosis virus is the causal agent of one of the most important viral citrus diseases in the world and has been detected from the citrus orchards in the east of Mazandaran province, IRAN. Because of the simultenious presence of various strains of this virus in infected citrus samples, molecular detection methods like PCR are not able to detect this virus with high accuaracy. In this research, we introduce a method for the precise detection of Citrus psorosis virus.

Detection and Identification of infected samples performed by triple antibody sandwich indirect-ELISA (TAS-ELISA) method and monoclonal antibody from symptomatic Thomson orange and Unshiu tangerine samples with suspicious symptoms to CPsV infection in Sari, Neka and Ghaemshahr districts of Mazandaran. Infection in samples tested by ELISA and confirmed by RT-PCR. Extraction of CPsV-dsRNA (Double-strand RNA) from infected samples was done by CF-11 column. Molecular hybridization performed by DIG labeled cDNA probe.

Exploring the electrophoretic pattern of dsRNA extracted from the CPsV infected citrus samples by using ELISA and RT-PCR methods, indicating the presence of CPsV genomes with the molecular weights of 920 bp and 2500 bp in infected samples. The presence of CPsV in the samples was also confirmed by using molecular hybridization and designed probe. 

The major advantages of the hybridization method relating to the dsRNA electrophoretic mobility is the simultenious detection of the all citrus psorosis virus strains within the infected citrus samples. This way can separate virus isolates.