Detection of phytoplasma in tomato and pepper plants by electron microscopy and molecular biology based methods

 Om-Hashem M. El-banna 1, M. S. Mikhail 1 , Azza G. Farag 2and   A. M. S. Mohammed *

1 Plant  Pathol.  Dept., Fac. Agric., Cairo Univ., Egypt. 2 Virus and phytoplasma Res., Plant  Pathol. Res. Inst. Agric. Res. Centre, Giza, Egypt.

Abstract:

     Phytoplasmas causing big bud  on tomato and witches’-broom symptoms on pepper were isolated from naturally infected plants. The detected phytoplasmas were transmitted by; grafting and dodder into healthy tomato and pepper plants and by dodder into periwinkle plants. Electron microscopy of ultrathin section prepared from petioles of infected plants revealed phytoplasma units ranging in diameters from 0.4 to 0.8 nm inside phloem tissues. DNA extracted from symptomatic samples was used as template for amplification  of  products of 1.8 kb using primer pair P1/P7 and 1.2 kb using primer pair R16F2/R2 by direct and nested-PCR, respectively. Based on the nucleotide sequence of the 16S/23S rDNA intergenic spacer region (SR), 241bp and 325bp for the Egyption phytoplasma isolates of TBB and PeWB, respectively, the association with these diseases were confirmed as well as, its phylogenetic relatedness to other phytoplasmas was achieved. Three concentrations of tetracycline hydrochloride i.e. 150,200,250 ppm were used to control the two diseases by two different treatments. Immersing transplants at tetracycline at 250ppm for either 15 or 30min gave best result.  Furthermore, the partial nucleotide sequence deposited at the Genbank (accession No. EU232714 and EU232715) respectively. Three concentrations of tetracycline hydrochloride i.e. 150,200,250 ppm were used to control the two diseases by two different treatments. Immersing transplants at tetracycline at 250ppm for either 15 or 30min gave best result.   This study of tomato big bud and pepper witches’-broom phytoplasmas is carried out for the first time in Egypt.