2010 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 One, 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 Two, 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.
The cooperative research was initiated to acquire new knowledge of ecology, niche adaptation and pathogenicity evolution of phytoplasmas, causal agents of economically important plant diseases worldwide. Metagenomic sequencing and genome survey sequencing approaches and bioinformatics tools have been used to characterize the population structures of the microbial communities associated with affected tissues of phytoplasma-diseased palm, potato, and other plants, gaining insights into interactions among phytoplasmas, their hosts, and other microbes sharing the same niche, and gaining an understanding of what roles a specific host (plant or insect vector) and the associated microorganisms have in the development of a specific phytoplasma populations and the disease induced. In this reporting period, we have conducted extensive field studies to identify diseased palm and potato plants, suspected insect vectors, and potential reservoir plants (mainly herbaceous weeds nearby the crop plants). We have collected samples from diseased palm trees in Caribbean region and diseased potato plants in Russia and the in Texas. Samples of five suspected insect vector species and more than 20 weed species were also collected. Total DNA was extracted from the collected samples. Primers and adaptors for metagenomic sequencing were designed and synthesized. This project was monitored by conference calls, regularly scheduled meetings, and exchanges of ideas, materials, and data.