CHARACTERIZATION OF PLUM POX STRAINS FROM REGHIN AREA -RFLP METHOD

Abstract

SUMMARY Plum Pox virus is the causal agent of the sharka d isease, considered the most pathogen viral of stone fruit crops in Europe. The impact of the disease makes a strong limiting factor for propagation material movement. Infected Prunus trees are the major source of inoculum. The virus is transmitted from them either by grafti ng or non-persistently by the aphid vectors Aphis spiraecola and Myzus persicae. The number of trees becoming infected in an orchar d is directly related, in a given season, to numbers of winged aphids. These aphids probe or feed on infected leaves, then fly to other trees where they again probe or feed. Symptoms may appear on leaves, petals, fruits and stones. They are particularly clear on leaves i n spring: mild light green discoloration, chlorotic spots, bands or rings, vein clearing or y ellowing, or even leaf deformation. In this study I collected twenty PPV isolates from two orchards situated in Reghin County, based on typical symptoms on leaves. Total RNA was extracted from fresh material plant using the RNeasy Plant Minikit. Than the viru s infection was confirmed by molecular methods. For the general detection of PPV, primers located near the 3` end of the coat protein (CP) gene were used to amplify a 243 bp fragment: P1: 5`-3` ACC GAG ACC ACT ACA CTC CC and P2: 5`-3` CAG ACT ACA GCC TCG CCA GA (Wetzel et al., 1991). For RFLP analysis the PCR products was treated with RsaI endonuclease. The primers P1/P2 confirmed the presence of the vir us in all the samples with RT-PCR analysis. Using RFLP method we could distinguish the two strains, D and M. Following the digestion with the restriction enzyme Rsa I we detected fifteen samples infected with PPV-D, two samples with PPV-M and three samples which presented PPV-Rec(PPV-D+PPV-M). Conclusions: All isolates collected present infecti on with PPV; PPV-D stain is most spred that PPV-M