Date of Award

8-2013

Document Type

Thesis

Degree Name

Master of Science (MS)

Legacy Department

Plant and Environmental Science

Advisor

Agudelo, Paula

Committee Member

Wells , Christina E.

Committee Member

Scott , Simon W.

Abstract

Nematode glutathione S-transferases (GSTs) have been implicated in plant-nematode interactions as effector proteins with an important role in the establishment of feeding sites. Studies with root-knot nematode in Arabidopsis thaliana suggest that GSTs may protect the nematode against oxidative plant defenses and modulate plant responses to parasitism. Our objective was to study the function of a GST from reniform nematode (Rotylenchulus reniformis) in soybean. We used a virus-induced gene silencing (VIGS) system, utilizing a Bean pod mottle virus (BPMV)-based vector and a partial sequence of gsts-1 from Meloidogyne incognita to silence the putative reniform nematode homolog. The effect of silencing this gene on reniform nematode infection was evaluated by inoculating treated soybean plants with 3,000 nematodes per plant. The reproduction factor was calculated thirty-five days after inoculation, and the experiment was conducted three times. Gene silencing was assessed by qRT-PCR at 0, 2, and 4 days after nematode inoculation (dai), using pecific primers for the reniform nematode gsts gene and for reniform nematode 18S ribosome. Hydrogen peroxide concentration in the roots was measured at 0 and 2 dai, using a fluorometric assay. Roots from the treated and untreated plants were fixed and sectioned for observations on the histopathology of infection. Reproduction on the plants inoculated with the silencing virus construct was significantly lower than in controls inoculated plants, suggesting this gene of reniform nematode plays an important role in the infection of soybean. Hydrogen peroxide concentration 2 dai in nematode-infected roots with the silenced gene was two times higher than that in roots without the silenced gene. We suggest that plant cells response to reniform nematode infection by producing superoxide and its dismutation product, hydrogen peroxide, both of which are toxic to plant-parasitic nematodes. The observed behavior of reactive oxygen species (ROS), cell wall thickening, and callose deposition support the possibility of this nematode-secreted protein potentially acting as a microbe-associated molecular pattern.

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