1.11.35
CHARACTERIZATION OF THREE PLUM POX VIRUS ISOLATES IN THE CZECH REPUBLIC

P K0MiNEK, R HAK and J POLAK

Research Institute of Crop Production, Division of Phytomedicine, DrnovskS 507, 161 06, Prague 6 - Ruzyne, Czech Republic

Background and objectives
Plum pox virus (PPV) is currently the most harmful plant virus in the Czech Republic. It is widely distributed in plum, apricot and peach trees. Two prevalent strains with different pathogenicity in plant species, PPV-M and PPV-D, are present in Europe. Herbaceous and woody indicator plants, polyclonal antibodies, Western blotting and restriction fragment length polymorphism (RFLP) were used for the differentiation of PPV strains in the Czech Republic.

Materials and methods
Three PPV isolates originating from plum, apricot and peach were maintained in Nicotiana benthamiana and Prunus persica var. GF 305 plants. Chenopodium quinoa, Chenopodium foetidum, Prunus persica var. Siberian C and Prunus persica var. GF 305 were used for biological differentiation of the isolates. PPV isolates were purified by differential and sucrose density gradient centrifugation. Polyclonal antisera were prepared and antibodies suitable for ELISA isolated. Affinity of the isolates to homologous and heterologous antibodies was evaluated by DAS-ELISA. Western blotting was used to differentiate virus strains on the basis of mobility of viral coat protein in SDS-PAGE [2]. The presence of specific restriction sites [1] was determined by RFLP of PCR-amplified coat protein gene of the three PPV isolates.

Results and conclusions
The apricot isolate caused no symptoms in C. quinoa leaves when inoculated with phosphate-veronal buffer. On the other hand, plum and peach isolates induced local necrotic lesions. C. foetidum reacted on infection with all three isolates with chlorotic-necrotic local lesions. Similarly, no differences among the isolates were found using P. persica var. Siberian C. On the contrary, P. persica var. GF 305 revealed severe reaction when inoculated with the apricot isolate (similar to PPV-M strain ) while plum and peach isolates induced only mild reactions (characteristic for PPV-D strain ). On the basis of the above results it seems that plum and peach isolates are similar in their biological properties and differ from the apricot isolate. The three PPV isolates showed similar affinity to all polyclonal antibodies tested. This signals their relevance to the same serotype. Coat protein subunits of apricot and peach isolates had similar mobility in SDS-PAGE and appear to be 38-kDa proteins as detected by Western blotting. They obviously belong to the PPV-M strain. The plum isolate had two bands of 37 and 39 kDa, and seems to be an intermediate type between PPV-M and PPV-D strains. It can be concluded that the Czech isolates of PPV from apricot and peach belong to the PPV-M strain. The isolate from plum is an intermediate type of PPV-D and PPV-M strains.

References
1. Deborrd G, Maiss E, Jelkmann W, 1995. Acta Horticulturae 386, 253-262.
2. Pasquini G, Mazzei M, Pilotti M, Barba M, 1995. Acta Horticulturae 386, 346-353.