MOLECULAR FINGERPRINTING OF MICROBIAL POPULATIONS IN PHYTOPLASMA-DISEASED PLANTS
Molecular Plant Pathology
2012 Annual Report
1a.Objectives (from AD-416):
1. Characterize microbial communities associated with tissue necrosis during the terminal stage of lethal yellowing disease of palms.
2. Determine genetic variability/diversity of phytoplasmas affecting potato and other crops.
The ultimate goal of the project is to gain new knowledge of ecology, niche adaptation and pathogenicity evolution of phytoplasmas, causal agents of economically important plant diseases worldwide.
1b.Approach (from AD-416):
For Objective 1, necrotic tissues will be harvested from lethal yellows (LY) disease-affected palm trees and total DNA will be extracted from selected tissues. Microbial populations will be characterized using 454 pyrosequencing technology with primers designed to target V3 and V6 hypervariable regions of bacterial 16S rRNA genes. For Objective 2, total DNA will be extracted from symptomatic tissues of phytoplasma-infected plants, and phytoplasma DNA will be enriched through cesium chloride gradient centrifugation. Phytoplasma genome survey sequencing will be conducted using 454 pyrosequencing technology using coded primers. For both Objectives, the sequence data obtained will be analyzed and annotated using a suite of bioinformatic tools.
This cooperative research to acquire new knowledge of ecology, niche adaptation and pathogenicity evolution of phytoplasmas, cell wall-less bacteria that cause diseases in ecologically and economically important plants worldwide was initiated through the use of genome survey sequencing and metagenomic sequencing approaches. In this reporting period, preliminary results were obtained in the characterization of microbial communities associated with tissue necrosis during the terminal stage of lethal yellowing disease of palms. Studies were continued on diseased palm trees in the Caribbean region and of diseased potato plants, suspected insect vectors, and potential reservoir plants nearby the crop plants in Russia and the state of Texas, U.S.A. Additional undescribed phytoplasmas were discovered, some of which represent new genetic lineages. Genome sequencing of phytoplasma strains representing several distinct species and strains was carried out using next generation sequencing methodology. The new information gained is providing bases for enhanced understanding of the phytoplasma pathogens.