ELISA detection of Apple chlorotic leafspot virus (ACLSV) and Apple mosaic virus (ApMV) in pome fruit in comparison to RT-PCR detection and the determination of genetic variation of their virus species in South Africa
Abstract: Apple chlorotic leaf spot virus (ACLSV) and Apple mosaic virus (ApMV) are responsible for reduced yield in the South African deciduous fruit industry. These two diseases are regulated by the South African Deciduous Fruit Plant Certification Scheme whereby no trees infected with these viruses are permitted for plantings. Currently the double antibody sandwich enzyme-linked immune-sorbent assay (DAS-ELISA) is prescribed as the test method for routine detection of these viruses in plant material. In the first part of this study, detection limits of DAS-ELISA and reverse transcriptase polymerase chain reaction (RT-PCR) for ApMV and ACLSV were compared respectively. The RT-PCR was found to be more sensitive than DAS-ELISA in the detection of ACLSV and ApMV. In the second part of this study, the genetic variation of ApMV and ACLSV isolates from South Africa was investigated. Extracted RNA was used for RT-PCR of the coat protein genes which were then sequenced. Phylogenetic trees were constructed using these sequences as well as reference sequences from GenBank. Phylogenetic analysis revealed that South African isolates of ACLSV were similar to isolates from the rest of the world, and that the majority of isolates are not restricted to a particular fruit group. This indicates that cross-infection between pome- and stone fruit is a strong possibility. It is also concluded that ACLSV was imported from various regions of the world, since similarity with a number of different overseas isolates was found. Phylogenetic analysis of the coat protein gene sequences of ApMV isolates indicated that two major groups occur in South Africa. Phylogenetic analysis also revealed that South African isolates in individual clades are not restricted to a single fruit group, which indicate the risk of cross-infection.