Molecular Detection Of Phytoplasma Research Articles

Molecular detection of phytoplasmas in witches’ broom diseased Ziziphus oenoplia from Andhra Pradesh, India

Ziziphus oenoplia is a wild jujube evergreen small tropical fruit tree that grows along the roadsides. During a survey in 2021, this jujube trees exhibited severe witches’ broom symptoms, therefore, an investigation was carried out, to verify the presence of phytoplasmas in the symptomatic plants. Total DNA was extracted from symptomatic leaves using a CTAB method and a PCR assay was performed using 16S rRNA gene specific primer pairs P1/P7 and R16F2n/R16R2. Amplicons of about 1.2 kb were consistently amplified from all the symptomatic plant samples in nested PCR assays, while no amplification was observed in non-symptomatic samples. Virtual RFLP analysis of 16S rDNA sequences allowed the classification of this strain into 16SrV group. The phylogenetic analysis of 16Sr DNA sequence of the detected phytoplasma strain indicates its close clustering with ‘Candidatus Phytoplasma balanitae’ and ‘Ca. P. ziziphi’ and further analyses on other housekeeping gene are necessary to define its affiliation.

Molecular detection of phytoplasmas in potato psyllids in ecuador

Since the potato purple top appeared and the potato psyllid was reported in Ecuador, crop losses have been significant. So far, phytoplasmas enclosed in two ribosomal groups (16SrI-F, 16SrII) and Candidatus Liberibacter solanacearum’ were molecularly identified in symptomatic potato plants. Later, Bactericera cockerelli, a potato psyllid, was also first reported and correlated to the potato purple top disease. Molecular identification of the pathogens involved in this disease has been quite elusive. Samples of potato psyllids were analyzed in this study resulting in the identification of phytoplasmas enclosed in group 16SrXI, and others related to Candidatus Phytoplasma prunorum’ and Ca. P. solani’. Potato purple top in Ecuador is a complex disease that constitutes a severe threat for the potato production in the region.

Molecular detection of phytoplasma from phyllody infected sesame and its vector orosius albicinctus distant

Asymptomatic and symptomatic plants were collected from the phyllody infected sesame field during kharif 2012. From the infected plants, leaves, flowers, stems, seed capsules and roots were selected both from symptoms and symptomless branch. Leafhoppers were also collected from the same field with insect collection net. Total genomic DNA from asymptomatic and symptomatic plants was extracted following CTAB (Cetyl Trimethyl Ammonium Bromide) method with some modifications and from leafhoppers with phenol-chloroform extraction protocol. Amplification of band size 1.8 kb and 1.2 kb was obtained from symptomatic plant and its vector Orosius albicinctus using phytoplasma specific primer pair P1/P7 and R16F2n/R16R2, respectively in first and second round PCR. The phytoplasma could be detected only from stems of symptomless branch of infected plant while in symptomatic plant's branch it was amplified from stem and flower. Thus the status of leafhopper, O. albicinctus as a vector of phytoplasma that cause sesame phyllody was confirmed. Further, it was observed that the symptomless parts of phyllody infected plants also harbour phytoplasma.

Indonesia

Witches’ broom disease caused by phytoplasma is a very serious disease on peanut (Arachis hypogaea) which may potentially lead to high yield loss. Insects are the most important agents of phytoplasma transmission in the field. The objective of this research was to examine the potential role of leafhoppers species as insect vector of phytoplasma and to determine their transmission characteristic. Two species of leafhopper i.e. O. argentatus and Empoasca sp. (both belong to Hemiptera: Cicadellidae) were chosen for this study. The methodology involved were transmission study of phytoplasma by O. argentatus and Empoasca sp., and molecular detection of phytoplasma by PCR technique to confirm the association of pathogen, insect vector and symptomatic plants. The result showed that specific symptom was observed when using O. argentatus in the transmission study with number of insect as low as 1 insect per plant, whereas Empoasca sp. was not able to transmit the disease. Incubation period of phytoplasma in the host plant was affected by the number of insect, i.e. the more insect vector the shortest incubation period. The phytoplasma was successfully detected using P1/P7 primer in symptomatic plants as well as in the insect vector.Key words: Empoasca sp., leafhoppers, polymerase chain reaction

First Report on Molecular Detection of Phytoplasmas in Papaya in Cuba.

Phytoplasmas and rickettsias have been associated with recent nonviral papaya diseases (1,2). Forty leaf samples with symptoms similar to papaya bunchy top (PBT) (1) and Australian papaya diseases (2) were collected from surveys done in Cuban papaya areas of Havana and Villa Clara provinces. Thirty nine plants showed typical PBT symptoms, while 35 plants also exhibited symptoms previously described for phytoplasma diseases (2) such as yellowing, crinkling, necrosis and deformation of older leaves, phyllody, virescence, short internodes, reduced latex flow, stunted crown leaves, and abscission of the fruit. Samples were evaluated by transmission electron microscopy, nested polymerase chain reaction (nPCR) with 16S ribosomal RNA phytoplasma primers SN910601/DH6-R16F2n/R16R2, PCR with rickettsial succinate dehydrogenase gene (sdhA) primers PBT1/PBT2, HaeIII, RsaI, and AluI enzyme restriction, and phytoplasma intergenic sequence analyses. Rickettsia PCR amplifications of 750 bp were obtained for samples with PBT symptoms, while the 35 phytoplasma-infected samples yielded amplifications of 1,250 bp. Restriction profiles and a 98% homology in the intergenic sequence (GenBank Accession No. AY257547) confirmed the presence of apple proliferation phytoplasma group. Electron microscopy analysis evidenced the presence of particles similar to rickettsia and phytoplasma pleomorfic bodies in more than 50% of samples analysed. Also, those similar to potyvirus and rhabdovirus were observed in 22.5%. To our knowledge, these results are the first report on the molecular detection of phytoplasmas in papaya in Cuba and suggest a possible concomitance among phytopathogens detected.