Detection of Pear Decline Phytoplasma by Polymerase Chain Reaction in Bulgaria

M. TOPCHIISKA1 and D. SAKALIEVA2
1 Fruit Growing Research Institute, BG-4000 Plovdiv, Bulgaria
2 Agricultural University, BG-4000 Plovdiv, Bulgaria

Abstract

TOPCHIISKA, M. and D. SAKALIEVA, 2001. Detection of pear decline phytoplasma by polymerase chain reaction in Bulgaria. Bulg. J. Agric. Sci., 7: 611-614

Polymerase chain reaction (PCR) and primers for phytoplasma found in trees of fruit species in Europe were used for detection and identification of the sequences of ribosomal DNA of phytoplasma – the causative agent of pear decline (PD) in pear trees. The pair of specific fO1/rO1 primers induced amplification of template DNA in six PD samples of phloem tissues of naturally infected pear trees, while the universal fU5/rU3 primers reacted positively in only two of the samples tested. The detection of the PD causative agent by PCR and group specific (fO1/rO1) and universal (fU5/rU3) primers is an appropriate method for finding phytoplasma contained in low concentration in trees of the fruit species, including pear. Restriction enzyme analyses of PCR products, produced with the fO1/rO1 and fU5/rU3 primers, identified the PD phytoplasma in trees of 6 pear varietries (i.e. Zolotistaya, Junska, Nouveau Poiteau, Forel, Progress, and Jubeleen Dar) – isolates No. 14-25A, No. 1-15A, No. 6-20A, No. 7-39A, No. 1-27A, and No. 5-5A, respectively. For the remaining 8 pear samples (including the two rose samples) that did not contain phytoplasma DNA, the results obtained were negative.
The data on phytoplasma DNA extraction, its detection and identification by PCR and RFLP analyses, and the proof of pear decline (PD) phytoplasma in pear trees have been reported for the first time in Bulgaria.

Key words: phytoplasma, DNA, PCR, RFLP, pear decline, pear