Detection of Apple Chlorotic Leaf Spot and Apple Stem Grooving Viruses Using RT-PCR

Abstract

Reverse transcriptase polymerase chain reaction (RT-PCR) procedures were used to amplify fragments specific for apple chlorotic leaf spot virus (ACLSV) and apple stem grooving virus (ASGV). Products visible in agarose gels were produced using templates consisting of as little as 5 pg of total nucleic acid extracted from infected apple leaves. Both viruses were also detected in fruit, bark, and root tissues of infected apples. These results suggest the viruses are distributed throughout actively growing trees. The systems were further validated by sequencing fragments amplified from leaves of a apple, cherry, and Asian pear trees growing in South Carolina. Comparisons with published sequences indicate that the fragments from the three ACLSV isolates have very high (≥97.6%) percent identity values to the corresponding region of a plum isolate and moderately high (≥81.4%) identities to an apple isolate from Japan. The fragments amplified from the ASGV isolates were found to be similar (≥88.5% identities) to the corresponding region of an apple ASGV isolate from Japan.