Registration of yeast will add new biofungicide

An OSU patented yeast expected to be massaged through the EPA registration process before the year's out would add a new biofungicide for postharvest prevention of molds.

BY RENÉ FEATHERSTONE

"What good is it if it doesn't get out to the people? We like to see practical applications of our work."

Dr. Robert Spotts

It's fungus against fungus. Over here, a neat-looking guy with a well-defined edge, brightly orange in color. See how the single cell constricts on one end, forms a lopsided figure eight, then cleanly pinches off the new cell.

Over there, a messy mass of bluish fuss. Spores galore, arrayed in chains.

You've already met that bad guy--it's the blue mold featured in many a fruit shipper's nightmare, prime culprit of pear and apple decay postharvest in the box. Yuck.

Dr. Robert Spotts, plant pathology professor at Oregon State University's (OSU) Mid-Columbia Agriculture Research and Extension Center in Hood River, champions the orange fungus which isn't a mold but a yeast. An OSU-patented yeast, as it were, and now in commercial channels.

Spotts explains that a yeast can be used for mold control on fruit; it's an organic concept, namely, displacement of the bad guy by the competing good guy or allowing a "benign" yeast to enter and feed and thus block molds from taking hold.

He elaborates that mold typically spreads by breaking down the cells adjacent to the place where it gained entry into the fruit, through wounds most likely, or the abrasion of a bruise, through the stem, or through the lenticels of overripe apples. In contrast, the yeast feeds only on the immediate nutrition available, it neither spreads malignantly nor, in the normal course of events, becomes visible to the naked eye of the consumer.

Commission research

Such biocontrol is a timely approach, according to the Washington Tree Fruit Research Commission in whose Wenatchee laboratory the OSU yeast will be tested for efficacy as well this year. One of the commission's scientists, Peter Sanderson, who did his postdoctorate work with Spotts, said that most postharvest diseases are caused by one of three fungi. "The major one is blue mold, which consists of two species of Penicillium. Gray mold is caused by Botrytis cinerea. The third fungus we're looking at is Mucor piriformis, which causes Mucor rot."

Currently, only two conventional postharvest fungicides are allowed for use on pome fruits in the United States, Sanderson noted, adding that "most fungicides aren't really fungicidal, they're actually fungistatic; they prevent either the growth of fungus, or they prevent spore germination. The action of TBZ (thiabendazole), for instance, is that it inhibits tubulin synthesis--tubulins are the strands that pull apart the
contents of a cell when it divides."

Besides TBZ, captan is also used as postharvest fungus control in Pacific Northwest warehouses; however, some export markets, including Taiwan, won't allow captan, Sanderson pointed out.

On the distribution and retail end, the fruit business has become much tighter than it used to be not so long ago, he remarked. "The grocery industry had a tolerance of up to 3 percent, regarding fruit decay in the box; today, if there's a single apple in a box that shows signs of decay, the buyer asks for an adjustment in price."

Small market

In 1999, Sanderson researched the efficacy of four new synthetic postharvest fungicides, of which, he said, only one looks like "a good candidate." This year, he's testing that candidate on fruit rather than only in test tubes; but even if he can show good efficacy, it's not at all certain that the chemical will ever make it to the commercial level.

He said that's mainly because "Postharvest is the last place a company looks for a market, since there's so little sales potential--one application, that's it. And pome fruits are a minor crop to begin with."

Sanderson added that testing postharvest materials for federal pesticide registration is very expensive. "They come under more intense scrutiny than field chemicals."

Further, he said, "The Food Quality Protection Act requires a company to consider all uses [of a chemical] on different crops, rather than look at just an isolated factor." He noted that a fungicide called Scholar had shown promising efficacy in controlling postharvest mold on apples and pears. The manufacturer had received an emergency Section 18 registration from the U.S. Environmental Protection Agency (EPA) for use on stone fruit in California; however, they did not register Scholar for pome fruits because "their risk cup was full."

In view of all that, a biofungicide is likely to be welcomed by the industry. Also, biological controls are in demand due to the large expansion of Washington organic acreage. Currently, two organic materials, Aspire (a yeast) and BioSave (a bacterium that also works by competitive displacement), are available for organic postharvest decay control. Although, in Sanderson's tests, "They only showed some efficacy."

A bio-company

The same company that produces Aspire is in the process of getting the yeast that Spotts is working with on the market. Ecogen has focused on biocontrols since the company started up 15 years ago, said Art Dawson, vice president for biofungicide development at Ecogen's headquarters in Langhorne, Pennsylvania. Crymax, a line of Bts (Bacillus thuringiensis), is perhaps their best-known product, he noted. They also manufacture by fermentation AQ-10 (Ampelomyces quisqualis), an obligate parasite fungus that was registered for use on grapes in 1995, and for use on tree fruits, nursery stock, and hops in 1997.

Whereas Aspire is a "first-generation biofungicide," the OSU yeast, which the company now calls EXC 9001, rates as a "second- or third-generation biofungicide," Dawson said. "This product is able to compete directly against synthetic fungicides. It has a broader pathogen range than TBZ. It's been shown that up to 50 percent of Penicillium has been able to build resistance against TBZ. Conversely, with the biological control, resistance is not an issue, because our biofungicide is not a poison but an antagonist."

Dawson is confident that Ecogen will obtain EPA registration for EXC 9001 before the year is out. "We'll register the product with the WSDA [Washington State Department of Agriculture] for organic certification immediately thereafter." At that point, too, the company will have a regular tradename for marketing the bioagent.

A history of patience

Spotts credits one of his former research associates with helping him discover the yeast. "Dr. Tara Chand-Goyal came to us with a strong background in biological control. He had a keen eye for subtle differences [in microflora]."

Back in 1993, Chand-Goyal and Spotts crisscrossed eastern Washington and Oregon in a minivan. "We collected fruit from all the major fruit-growing districts in the Northwest," Spotts recalls. "We looked for poorly maintained orchards."

They brought back a sampling of apples and pears, picked at random into plastic bags at "maybe 20 different stops." In the lab, they separated the microorganisms from the fruits, which is done by washing the fruits and then growing cultures from the wash water. "There were thousands of [microflora] colonies on the plates--it was a zoo."

As it happened, the yeast that would eventually test well as a biocontrol against mold came from the old U.S. Department of Agriculture's orchard at Yakima, Washington.

"It was on the surface of a Bartlett pear," said Spotts.

The scientific name for the yeast is Cryptococcus infirmo-miniatus, CIM for short. The testing, said Spotts, was a long and tedious process. The CIM was grown on a petri dish, then it was ascertained that the growth was "a pure culture." Then, a piece of fruit was sterilized on the surface, and punctured. Into that puncture wound, they released a tiny bit of CIM. "Then, we put that fruit into a moist chamber to see what happened."

The next series of tests were for efficacy, Spotts said. On one plate they grew the mold fungi, on the other the yeast. "Of the yeast we used a concentration of 200 million cells per milliliter." Yeast cells and mold spores were then mixed in a test tube. The mixture was applied to a wounded piece of fruit. The control was a pure mold fungus strain on a wounded piece of fruit.

Further tests were undertaken in the extreme conditions that apples and pears experience in real life, Spotts said, including the cold temperatures of storage, and the about 60°F. of the drying tunnel.

Nicely enough, indications are that CIM works against all mold invasions, the blue and the gray as well as Mucor rot, plus, it's effective against molds on cherries, too.

As you can imagine, plant pathology is a science for the patient person. Spotts initially was interested in forestry, which he studied at Colorado State University in Fort Collins. After graduating, he participated in a project that looked at a forest phenomenon near Denver, at the time called "tip-burn disease," on pines. As it turned out, it wasn't a disease but a physiological disorder caused by salts in the soil. But the project's focus on plant diseases is what "moved me from forestry over to pathology," Spotts remarked. He earned his doctorate in plant pathology at Pennsylvania State University.

Getting the product out

Having isolated a biocontrol agent in collaboration with Chand-Goyal was a research coup, but "What good is it if it doesn't get out to the people?" Spotts said. "We like to see practical applications of our work."

The first step toward that was to check the database of the Centers for Disease Control, Atlanta, Georgia. The yeast is not listed as a known threat to human health, they found.

After applying for a U.S. patent, Spotts then looked for a company that would be good to work with. "Merck was our first thought, because that company was the manufacturer of TBZ. When we combined our yeast with a low rate of TBZ, we saw that a synergism takes place. That made us think that Merck would be a good candidate to license to. Away we went, but then Novartis purchased Merck. At first Novartis was still interested, but, eventually, they decided that the market for our yeast would not be big enough."

All along, they'd also been talking with Ecogen. They licensed their yeast to that company finally in the spring of 1999.

Since then, Ecogen has formulated the yeast as a WDG (wettable dispersible granular), so Spotts's ongoing work is made easier. "Generally, a gram of WDG product contains about 10 billion cells."

This year, Spotts will "scale up" his yeast research. The OSU facility at Hood River has a small packing line, packing from boxes not bins, he said. On the packing line, they'll also do more experiments with CIM/EXC 9001 as an on-line spray. He's certain that conventional warehouses are likely to use the product once it's EPA-registered. Already, he remarks, "Two of the three major [fruit] packers here in Hood River use Aspire and Biosave, added on top of their regular fungicide."

In the overall picture, Spotts predicted that "Warehouses are willing to spend some money on decay control, because it's only a small fraction of the overall expense." His hunch is that the most efficient means to control decay is by an integrated program of synthetic and biological materials. "I'm stressing that to the industry."

He's also looking at field applications of the yeast.

Meanwhile, the next generation of plant pathologists is already keen on getting in on the fungal battle. "I have a high school senior intern, Andy Miller," Spotts related. "He goes around the grocery stores and asks for fruit they'd be throwing out because of decay. He's brought all kinds of fruits in here--papaya, grapes, star fruit. We're testing our yeast on those fruits. So far, the yeast has shown good efficacy against molds on all those kinds of fruit." m

Pesticide commission revises proposal process

The Washington State Commission on Pesticide Registration has announced changes to its process of soliciting requests for proposals to support pest control in Washington.

These changes relate primarily to the expansion of the commission's mandate to include projects that support research, implementation, and demonstration of any aspect of integrated pest management and pesticide resistance management in Washington.

State lawmakers created the commission in 1995 to assist users of pesticides in obtaining and maintaining pesticide registrations for minor uses in Washington. To more fully meet the state's pest management needs, the 1999 legislature authorized the commission to fund a wider array of projects. Washington State University was named fiscal agent for funds made available to the commission.

The most significant new or emphasized requirements for requests include:

--Proposals must be received by the administrator 30 days prior to the meeting at which they are to be reviewed;

--Projects including both old and new mandate activities must clearly define the proportion or amount of the budget dedicated to each mandate;

--Commission meetings alternately hear old and new mandate proposals;

--A one-to-one match of commission funding is not mandatory but is encouraged and may influence proposal funding. User group support must be adequately demonstrated;

--A representative of the requesting group must be present at the meeting at which the request is made.

Commission Administrator Alan Schreiber is available to assist with proposal preparation and to answer questions.

The commission Web site <www. wscpr.org> provides meeting dates and specific examples of funded projects and a listing of all projects funded since 1988. The commission can be contacted at (509) 543-9757 or e-mail Schreiber at <aschreib@cbvcp.com>.

The 1997 Ag Census revealed some interesting facts about Washington State agriculture:

* The average age of Washington's farmers continues to increase-- from 53.1 years in 1992 to 54.2.

* 53% of farm operators considered farming to be their principal operation.

* 12%of farm operators are female.

* 52% of farm operators worked off the farm.

* 34% of farm operators worked 200 days or more off the farm.

* There were 15.2 million acres of Washington farmland.

* The average size farm was 523 acres.

Washington Agricultural Statistics Service