Date: Fri 15-Dec-1995

Publication: Bee

Author: ANDREA

Quick Words:

chestnut-genetic-experiment

Full Text:

with cut: Genetically Engineering The American Chestnut Back Into Existence

B Y A NDREA Z IMMERMANN

The American chestnut tree. Once it accounted for 40 percent of New England's

hardwood forests; today it has been reduced to an under story specimen,

susceptible to a blight fungus that creates lethal, sunken cankers.

But, for the first time in this country, state and federal regulatory

commissions have permitted a genetically engineered biological control to be

tested outside lab with the expectation that it will spread.

Dr Sandra Anagnostakis of the Connecticut Agricultural Experiment Station in

Hamden is heading the field work, which is focused in four acres of the

Housatonic State Forest in Sharon. The mycologist hopes the spread of this

biological control, coupled with perpetuation of a more resistant strain of

chestnut through cross-breeding, will allow the American chestnut and hybrids

to be reinstated as canopy trees in the northeast.

"I'm sure there are many good economic reasons to restore the chestnut tree -

as a valuable timber crop, or to control the blight on orchard cultivars. But

the main value is to increase diversity of the forest," said Dr Anagnostakis.

"Improving diversity is intrinsically a good thing, a healthy thing." For

instance, there will be a trickle down effect whereby wild animals will

benefit from the food supply.

Arrival Of The Blight

In the 1880s a fungus was brought into the country on imported Japanese

chestnut trees. The blight fungus infects through wounds and kills the stem of

native chestnuts and chinquapins. "The trees usually sprout from the base

forming a clump of sprouts which become wounded, infected, die, and sprout

again," said Dr Anagnostakis. The Connecticut Agricultural Experiment Station,

which was established in 1875, has been working on the problem since the

beginning of the epidemic, planting resistant Asian chestnut species and

crossing them with susceptible American trees, she added.

In the 1950s and 60s, scientists were looking for stop-gap measures - such as

the application of chemicals - that would allow farmers to produce a crop

despite pests and pathogens. Studies led to better understanding of the

biology of pathogens so "we'd know when to hit to get the most effective

[temporary] control."

Any time continuous treatment is needed, it is considered a "therapy," said

the scientist. "It was clear for a long time that chemicals were not going to

be effective against chestnut blight," she said. So they sought a way to

change the biological system to bring the pathogen into balance with it's

host. Such a "biological control" could be put into the forest and not only

survive but multiply to affect widespread change.

In 1968, a scientist at the experiment station who was working on chestnut

trees, brought a diseased specimen to Dr Anagnostakis because fungi was her

expertise. Little research had been done on the fungus since the turn of the

century, and there was virtually no information on how it grew. So she and

another colleague grew specimens and performed many experiments including the

first genetic work on the fungus.

Biological Control

In 1972, a French scientist, Jean Grente, shared with Dr Anagnostakis samples

of a diseased chestnut fungus that he had imported from Italy. "We

demonstrated what was wrong with the fungus was it had a virus" - a natural

biological control, she said. Although it wasn't lethal, the virus prevented

the fungus from penetrating the chestnut stem and killing the tree, while

spreading through the fungal population.

A search ensued in this country for hypovirulent strains, and similar

"healing" cankers were found in Tennessee, Michigan, Virginia, and West

Virginia, which yielded strains of the blight fungus that were less able to

kill chestnut trees, she said.

"When scientists put bits of a hypovirulent blight fungus into holes in the

bark around killing cankers, viruses can move into the virulent strains that

caused the cankers. The cankers then stop expanding, and the tree's natural

defenses of walling off invaders succeeds in protecting the tree's living

cambium," said Dr Anagnostakis. "Once hypovirulence has been established in a

chestnut blight population, hypovirulent spores are moved around in orchards

and in the forests by every creature that moves up and down the trees. It is

likely that everything that walks across the cankers can pick up

virus-infected spores on feet, fur, feathers, and beaks, and move them to new

cankers."

Even very susceptible American chestnut trees can be kept alive through

biological control, and can continue to breed. This works well in orchards,

but is less effective in forests where many trees are competing for light,

moisture, and nutrients, the scientist said. Not all spores produced by the

natural hypovirulent contain the virus, and different strains of the blight

fungus prevent easy transmission of those that do.

Genetic Engineering

"An even better solution, would be to have hypovirulence viruses enter the

spores produced after mating, when many new strain types may be produced by

genetic recombination," said Dr Anagnostakis. "Our newest tool in fighting the

blight are new strains of the blight fungus with the genes of a hypovirulence

virus inserted among the genes of the fungus."

Dr Donald Nuss of the University of Maryland has produced these strains and

studied them in the laboratory. Dr Anagnostakis has collaborated with Dr Nuss

and is doing the field work here in Connecticut with the help of two

assistants. They first tested the strains on potted American chestnut trees

and a variety of other woody plants in an insect-proof greenhouse at the

experiment station. The strains survived, but did not cause any disease, she

said.

State and federal regulations restrict the use of anything produced through

genetic engineering. Usually, scientists would have to offer assurance that

whatever testing was done would not permit a strain to spread. "We want it to

spread and affect biological control," said Dr Anagnostakis, who got approval

to start work in June of 1994.

"We know the natural virus has four genes; we have the DNA sequence," said Dr

Anagnostakis. "But we're hoping to find out what genes in the fungus are

responsible for killing the tree; and what genes are responsible for making

the fungus unable to kill the tree."

The Housatonic State Forest in Sharon is an "ideal test site" because there is

diverse terrain, a lot of chestnuts in all directions and within close

proximity to each other, logging can be controlled, and few people traverse

the area. "As we find the strain spreading, we can see if it is spreading

faster north than south, or up hill than down hill," she said. "We can see how

site conditions are affecting it." For example, areas with drier soil can be

discerned by the types of shrubs growing there.

Every American chestnut tree in the four-acre plot that is one inch or more in

diameter will be numbered and mapped; about a third have been documented thus

far. While her assistants John Shepard and Pamela Sletten mark the location,

Dr Anagnostakis checks every tree. If she finds a canker, she will note it is

"Canker No 1 on Tree No 28" and use a non-toxic china marker to make a

permanent notation directly on the bark of the tree.

In order to do a scientific evaluation, Dr Anagnostakis needed to track

cankers on trees that appeared at the same time. So she actually inoculated

the trees with the killing strain of fungus and then treated one in each pair

of trees. Most of the test trees are three inches in diameter.

Cross Breeding

"I don't think [biological control] is the entire answer. We are breeding

chestnuts to get resistance from Asian trees into American trees," said Dr

Anagnostakis, who has begun to implement a cross-pollination breeding scheme

proposed by Dr Charles Burnham, the well known American corn geneticist.

"People forget how recent our knowledge is of how genes behave and segregate,"

said Dr Anagnostakis. "The idea of directed breeding with tree crops has come

about within the past 50 years."

The scientist said her success with the first cross tentatively proves

Burnham's theory. "Now we're going to have chestnut trees that look like

American chestnuts but have as much resistance as Asian chestnuts... We still

need hypovirulence because these trees will cross pollinate with American

Chestnut, and all offspring will be intermediates," she said. "Hypovirulents

will allow intermediates to [mature enough to] cross and eventually the tree

population will change."

The experiment station has the largest collection of species of chestnuts in

the world. Scientists from around the world ask Dr Anagnostakis for nuts and

specimens.

"Molecular biology is going to have ramifications for all kinds of plant

disease. Nuss has been able to put a virus in other fungi, which means we will

be able to control other plant disease," she said.

"The science is fascinating," admitted Dr Anagnostakis, "but I really want to

save the chestnut trees."