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Spying on an elusive virus.

Spying on an Elusive Virus

It's a very confusing virus: It needs a very specific insect to transmit it. Or does it? It causes a plethora of symptoms in different hosts - even no symptoms in some. And it's an exotic organism. Or is it?

To further muddle matters, the virus has had two names. Researchers in Japan refer to the virus as soybean dwarf virus (SDV) because it attacks soybeans. Investigators in Australia, New Zealand, and Tasmania have called it subterranean clover red leaf virus because it attacks clover.

But are they really strains of the same virus? And if so, could they start attacking soybeans or other crops in this country?

These questions are important to the safety of crops in this country. Agricultural Research Service plant pathologists Vernon D. Damsteegt and Oney P. Smith and biological lab technician Andrew L. Stone are tracking down some of the answers at the Foreign Disease-Weed Science Research Unit in Frederick, Maryland.

It's known that the virus is transmitted from plant to plant only by aphids - somewhat as yellow fever virus is transmitted person to person only by mosquitoes. Similarly, only the female aphid carries the virus as with virus-transmitting mosquitoes.

The aphid, a little soft-bodied sapsucker, sticks its mouthparts into leaves or stems, puncturing them and probing to find just the right spot to feed. Sucking away steadily, the aphid must feed on an infected plant for at least half an hour for the virus to move from the plant into the aphid's gut, into its blood system, and then to its salivary glands.

When the aphid feeds again, the virus picked up from the first plant is neatly deposited into the phloem of the second plant and subsequently spreads throughout the plant, infecting it. The virus must rely on the aphids to find just the right host where it can thrive.

In Japan, the virus relies on the foxglove aphid, Aulacorthum solani, to find soybean host plants. In the United States, however, the virus has not been found in soybeans. Foxglove aphids here shun soybeans. The reason for this apparent aversion remains a mystery to entomologists.

Besides having a specific aphid carrying it to specific plant species, the soybean dwarf virus in Japan consists of two strains, each producing dramatically different disease symptoms.

The SDV-Y (yellowing) strain causes the older leaves of an infected plant to turn a brilliant yellow color and the leaves to appear sparse. But when the SDV-D (dwarfing) strain attacks, the plant grows to only about a third of its normal size with leaves that become dark green and curl downward.

In the 1980's, Adrianna D. Hewings and others studying SDV-D, SDV-Y, and subterranean clover red leaf virus confirmed that all three viral strains reacted about the same way in diagnostic tests that employed antibodies.

Antibody tests such as the ELISA (enzyme linked immunosorbent assay) are often used to distinguish the presence of a certain protein in a sample. The test can be tailored to react to and thus detect virtually any protein. In the case of the SDV strains and the clover leaf virus, similar reactions indicated that they were the same virus.

Hewings, an ARS plant pathologist now at the University of Illinois, was the first in this country to purify the SDV-D and SDV-Y strains.

"Based on the test results and other information and the fact that SDV was described in the research literature before subterranean clover red leaf virus, virologists decided to classify the latter as a strain of SDV," says Damsteegt.

Virus Now What Is Expected

In the United States, the virus hasn't been found in soybeans, but it may be wrong to assume that the virus isn't here at all.

In 1983, an SDV-like pathogen was isolated from legumes by James E. Duffus, who works for the ARS Sugarbeet Production Research Laboratory in Salinas, California. This isolate was specifically transmitted by the pea aphid. Acyrthosiphon pisum. Then, SDV-like pathogens from white clover were found in the Carolinas, Florida, Kentucky, Maryland, Mississippi, and Virginia.

Very interestingly, all these isolates were transmitted by the pea aphid and produced leaf-reddening when injected into subterranean clover.

Part of Damsteegt's and Smith's job is to determine if SDV is a potential threat to U.S. soybean production. At the Frederick quarantine facility, researchers work with plant pathogens from all over the world.

Now the plant pathologists have found a way to differentiate between the two Japanese strains of SDV. "The method examines the virus' RNA chromosome using double-stranded RNA analysis," says Smith.

How the Virus Works

The virus has a six-sided protein shell that surrounds and protects a single-stranded RNA chromosome, its genetic material. When the aphid feeds, the virus infects the plant and releases this RNA. This forces the plant cells to reproduce the virus' RNA.

"Viruses aren't technically alive because they can't reproduce their own RNA," explains Damsteegt. "They need plants to do it for them."

Once the aphid injects the virus into the plant, the virus releases its RNA chromosome. The single-stranded RNA is converted to double-stranded RNA (dsRNA), as a part of the life cycle of the virus. The scientists can isolate this dsRNA and genetically "fingerprint" it with a standard laboratory technique that uses a small electrical current to separate various nucleic acid molecules into bands within a gel material.

"Using the dsRNA analysis is a very accurate way to differentiate between the D and Y strains of SDV," says Smith. Each strain produces two size-specific dsRNA's with different molecular weights that stop at particular levels on the gel.

Smith and Damsteegt now want to apply the technique to see what other soybean dwarf virus strains exist in the world.

After Duffus' identification of an SDV-like virus transmitted only by the pea aphid, interest was generated through the Southern Regional Research Project S-228, "Forage Legume Viruses," to search for SDV-like viruses and other luteoviruses in legumes in the United States. The Committee, made up of researchers from the southeastern states, is working to understand and control viruses in forage legumes.

In the past 5 years, scientists at the Frederick lab and Duffus' lab, and Michael R. McLaughlin at the Forage Research Unit, Mississippi State, Mississippi, have identified several SDV-like isolates. What damage they do to legume crops or pastures is being investigated.

So, is SDV exotic? Maybe some strains are, says Damsteegt, but for soybeans, the Japanese biotype (or subspecies) of the foxglove aphid may be more exotic than the virus it transmits.

"Where has the research on SDV taken us?" he adds. "To a better understanding of the occurrence and distribution of a previously poorly understood virus and to the realization that viruses may already occur in areas we believe they do not."

A Unique Evolutionary Relationship

Before Vernon Damsteegt could begin to figure out where the soybean dwarf virus was coming from, he first had to determine which aphid was carrying the virus.

All aphids are parthenogenic, meaning they are produced asexually as clones of their mothers. Would they then feed on the same plants? And is there a danger they could spread the virus to U.S. soybeans?

Damsteegt tested green and yellow biotypes (or subspecies) of soybean-feeding foxglove aphids from Japan, and nonsoybean feeders from California, New Zealand, and New Brunswick, Canada. The aphids were identified by Manya Stoetzel, who is with the ARS Systematic Entomology Laboratory in Beltsville, Maryland.

This foxglove aphid feeds on soybeans in Japan, carrying the virus to that crop with devastating results. If only half of the plants in a soybean field become infected with the virus, yield drops by about 40 percent.

The same aphid exists in the United States, but it seems to ignore the soybeans and favors clover and potatoes. "It appears that U.S. aphids just don't like soybeans," says Damsteegt.

The pea aphid, Acyrthosiphon pisum, also carries strains of the virus. It feeds on clovers, peas, french and broad beans, lentils, lupines, and sugar beets but doesn't like soybeans.

Damsteegt thinks that each of the aphis biotypes is virus-specific; that is, each carries only one form of the virus.

"Though the soybean dwarf virus is a group of closely related strains, those transmitted by the foxglove aphid are exotic to the United States," he says, "whereas allendemic SDV-like viruses are carried specifically by the plea aphid. So there may not be much danger to U.S. soybeans," he says.

"There is a unique evolutionary relationship, and a very specific one, between the aphids, the viruses, and the plants," says Damsteegt.

PHOTO : Soybeans being checked by plant pathologist Vernon Damsteegt are infected with soybean dwarf virus. (K-4094-7)

PHOTO : Using ethidium bromide stain and ultraviolet light, plant pathologist Oney Smith detects strain-specific dsRNA of soybean dwarf viruses. (K-4095-11)

PHOTO : Adult pea aphids, about an 1/8th inch long, transmit an SDV-like virus to clover in the United States.
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Title Annotation:soybean dwarf virus/subterranean clover red leaf virus research; includes related article
Author:Konstant, Dvora Aksler
Publication:Agricultural Research
Date:Jul 1, 1991
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