Comparison of diagnostic techniques for the detection and differentiation of Cherry leaf roll virus strains for quarantine purposes

Some strains of Cherry leaf roll virus (CLRV) are considered as quarantine pests in New Zealand. CLRV was detected in seven plant host species: Actinidia chinensis, Hydrangea macrophylla, Malus domestica, Plantago major, Ribes rubrum, Rubus idaeus and Rumex sp. collected from New Zealand between 2005 and 2012. Biological, serological and molecular techniques were compared for the detection and differentiation of CLRV isolates. The biological analysis revealed differences in symptomatology and disease severity among the isolates. The five isolates tested by ELISA were serologically related to each other using polyclonal antisera with only one out of four commercially-available antisera successfully detecting all of them. The phylogenetic analysis of sequences obtained from parts of the coat protein, polymerase and 3ʹ-untranslated regions revealed that the New Zealand CLRV isolates clustered into two closely related but distinct phylogenetic groups with some isolates grouping differently depending on the gene studied. The New Zealand CLRV isolates were clearly distinct to overseas isolates found in phylogenetic groups A, D and E. The conventional RT-PCR using primers targeting the CLRV coat protein coding region is recommended for determining sequence differences between strains. These findings will be useful in making regulatory decisions with regard to the testing requirements and the CLRV strains to be regulated in New Zealand.