WSU scientists pursue single test for plant viruses

By Terri Reddout, WSU Extension Prosser
PROSSER, Wash. — To determine if a fruit tree has a virus currently takes a minimum of nine separate laboratory tests in combination with several biological assays, and these can identify only known viruses.
Scientists dream of running a single test to detect all viruses in a plant. That idea is closer to reality thanks to the Washington Tree Fruit Research Commission, which has funded a two-year project to evaluate a universal plant virus microarray test.
Potentially, such a test might even be able to detect previously unknown viruses.

Quicker, cheaper, more detailed

“This research could lead to quicker and cheaper testing that gives us more detailed answers than current laboratory methods,” said WSU’s Ken Eastwell, who will serve as the principle investigator on the project. Eastwell, based at WSU’s Irrigated Agriculture Research and Extension Center in Prosser, is a plant pathologist and director of the Clean Plant Center of the Northwest. “This will benefit fruit tree growers and nurseries by getting clean plant material to them sooner and at lower cost. Additionally, it will help growers because we will be able to unravel emerging disease situations in orchards much faster.”
Microray technology
Originally designed to detect viruses in humans and animals, microarray technology was modified to detect plant viruses by the USDA’s Agricultural Research Service. The new project enables a team of WSU and USDA scientists to further modify testing protocols and find ways to optimize testing conditions for fruit trees diseases.

“We know the technology works,” said James Susaimuthu of the National Clean Plant Network–Fruit Trees program, also based at WSU’s research center in Prosser. He will serve as a co-principle investigator on the project. “We’ll be determining what changes need to be made to existing testing procedures to allow the microarray to give us clear results for fruit tree viruses.”

Power of attraction
The microarray process takes advantage of the power of attraction and starts with a slide outfitted with 10,000 micro probes. Each probe is designed to attract different types of viruses. RNA is extracted from the suspect plant and also from a healthy control plant. The RNA is then copied into DNA, a more stable molecule.
The DNA from the suspect plant is colorized with a red florescent dye, while the control DNA is dyed green. The two are mixed together and a drop of the DNA-rich solution is pipetted onto the microarray chip.
In a manner similar to the way negative and positive magnets are attracted to each other, the DNA molecules seek out those probes in the microarray that are complementary to its sequence. When the molecule finds one it is attracted to, it sticks or binds to it.
Once this binding or “hybridization” is completed, excess dyed DNA is washed off and the slide is scanned in a microarray reader. The reader scans and records the red and green florescent color of each of the spots on the slide. Software analysis then compares this color pattern to that produced by known virus sequences to indicate the family, genus and species of virus.
Universal virus slides
The first phase of the project has started with the printing of the universal plant virus microarray slides. This fall the research team will start training and optimizing methods and data analysis techniques. The second year of the project will be spent evaluating the effectiveness of the test.