Molecular detection and characterization of phytoplasma infecting brinjal (solanum melongena L.)
Loading...
Date
2015
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
Department of Plant Pathology, College of Agriculture, Vellayani
Abstract
The study entitled “Molecular detection and characterization of phytoplasma infecting Brinjal (Solanum melongena L.) was conducted at the Department of Plant Pathology, College of Agriculture, Vellayani, with the objectives to study the symptom development, transmission, molecular detection and characterization of phytoplasma infecting brinjal and its relationship with phytoplasma diseases of other crop plants. Brinjal little leaf (BLL), collected from the Crop museum, College of Agriculture, Vellayani and catharanthus little leaf (CLL) obtained from Coimbatore were maintained for further studies.
Symptomatology revealed the characteristic little, narrow, soft, glabrous and smooth leaves produced as clusters along with yellowing, proliferation of axillary shoots, shortened internodes, stunted bushy or rosette appearance and phyllody, the conversion of floral parts into leaf like structures.
The graft transmission was found to be 100% successful while the percentage transmission by dodder was only 10% in brinjal and 20% in catharanthus. Phytoplasma was maintained in vivo in plants by grafting and in vitro by culturing the infected explants on MS media supplemented with 0.2 mg l-1 BAP, 0.6 mg l-1 NAA and 0.4 mg l-1 IAA.
Biochemical analysis of healthy and diseased plants revealed that the contents of protein, phenol and chlorophyll were reduced in the inoculated plants as a result of phytoplasma infection. Carbohydrate content in brinjal increased immediately after inoculation and then decreased. The activity of peroxidase (PO) was enhanced in the inoculated plants while that of polyphenol oxidase (PPO) was reduced. The activity of phenyl alanineammonialyase (PAL) was reduced immediately after the inoculation, but enhanced at 30 and 60 days after inoculation (DAI).
91
92
The electrophoretic analysis of proteins using SDS-PAGE revealed the presence of two extra protein bands in the infected samples with molecular weights of 29 kDa (Kilo Dalton) and 43 kDa. The isozyme pattern analysis of peroxidase using native PAGE revealed two isoperoxidase bands in the inoculated plants with Relative mobility (Rm) values, 0.17 and 0.47, but a single band in healthy plants with Rm value of 0.17.
Molecular detection was done using nested PCR. PCR products of ~1.8 kb (Kilo base) were obtained in direct PCR with phytoplasma universal primer pair P1/P7 and the nested PCR with P1/P7 followed by R16F2n/R16R2 amplified the fragment of size 1.2 kb. The presence of phytoplasma in tissue culture plants was also confirmed using nested PCR.
Comparative nucleotide sequence analysis of brinjal and catharanthus isolates with the existing data base from NCBI revealed a 100% homology with brinjal little leaf phytoplasma isolates from Haryana and IARI and 99% homology with potato witches’ broom, potato purple top, tomato big bud phytoplasma etc.
The 16S rDNA sequences of BLL and CLL phytoplasma shared 99.7% similarity with that of ‘Candidatus Phytoplasma trifolii (Ca. Phytoplasma trifolii)’. Thus the two phytoplasma isolates were identified as the related strains of ‘Ca. Phytoplasma trifolii’.
Description
MSc
Keywords
Citation
173777