Citrus Greening

Researchers seeking weapons in the fight against citrus greening are delving into the genetic code of the disease’s pest vector—and arming the citrus tree.

The aim is to turn the Asian citrus psyllid’s body against itself, shutting down critical systems such as digestion, through RNA interference (RNAi) technology.

Two different projects are zeroing in on specific gene sequences unique to the target pest, developing material to be applied to citrus trees. When the psyllid feeds, this material would latch onto the targeted genes—and only those sites—to disrupt their function.

Wayne Hunter, research entomologist at the U.S. Agriculture Department’s Agricultural Research Service lab in Fort Pierce, Fla., and Kerik Cox, assistant professor of tree fruit and small fruit at Cornell University in Geneva, N.Y., are leading the two RNAi projects.

Both are targeting the psyllid genes essential to producing enzymes necessary to digest the insect’s food. Other possibilities include disrupting the psyllid’s reproductive and nervous systems.

Borrowing from honeybee experience

Hunter’s research builds on earlier work to help protect honeybees against viruses linked to colony collapse disorder, resulting in a vaccine now available from Miami-based Beeologics LLC.

“This is a new breakthrough in insect pest management and disease problems,” Hunter says.

Applying it to citrus psyllid and greening means treating the tree like a patient, designing a vaccine that deals with a particular problem, he says. It’s a technique that can be adapted to other pests and crops.

What’s behind RNAi technology?

RNAi technology makes use of a virus-fighting defense mechanism found in all organisms, from humans to insects, says Robert Shatters, a research molecular biologist at the ARS lab in Fort Pierce.

Viruses produce double-stranded RNA.

In an organism infected with a virus, that defense mechanism seeks out and disables any material with double-stranded RNA.

Giving the citrus psyllids such material that matches their genetic code means they essentially destroy themselves, Shatters says.

Hunter’s approach doesn’t involve transgenics.

Lab tests show a low persistence rate of the RNAi molecule, fading after a few months from detectable rates in the plant.

“The idea would be to treat the tree in winter when the food supply for the pest is limited,” Hunter says. That period draws the greatest concentration of psyllids to citrus trees for feeding, which in turn should result in a larger impact on psyllid populations.

Psyllids die within seven to 10 days of ingesting the material, he says. A later booster treatment might be needed to extend full psyllid control throughout the season.

The low persistence rate also means that by the time growers harvest their crop or processors turn it into juice, “the molecule is gone,” he says.

Added safety stems from physiological differences between mammals and insects. The genetic sequences being targeted aren’t found in humans—or even in unrelated insects, including beneficials.

Hunter and Cox are taking different approaches in how they deliver the disruptive molecules to the psyllid.

A very targeted approach

At Cornell, Cox is developing citrus plant material that will express the desired interfering RNA as long as the tree lives.

His best prospects will be shipped later this year to Florida to grow in test plots.

The RNAi molecules show up only in the plant’s phloem, where the psyllid feeds, and never appear in the fruit.

“It’s very targeted,” he says. “It’s not even [directly] toxic to the [citrus psyllid] but shuts down its processes.”

Hunter is seeking the most cost-effective delivery method for his approach. He’s achieved absorption through a drench treatment at the roots in trees up to 2 1/2 meters tall, but he’s testing other options for greater efficiency.

A foliar spray is one possibility, but might require using extra material to compensate for drift, he says. Injections would use the lowest dose of all because every drop would be certain to reach the tree.

“We know we need to get it down to $400 per acre or less,” Hunter says. Planting densities will also affect costs.

A standard imidacloprid program is his baseline for cost comparisons.

“I think we should be able to closely match that,” Hunter says. “You only need a small amount of the [RNAi] molecule to get into the pest to get it—but it’s a big tree” to cover.

Growers currently rely on multiple sprays of several insecticides for protection against the citrus psyllid—and to ward off resistance, he says.

Resistance is an unlikely problem with the RNAi approach, the researchers say.

A long-term solution

Resistance develops when the pest survives a low dose of a compound and passes on the activated resistance gene to its offspring, Hunter says.

He and the others are targeting multiple spots in the psyllid’s genetic code, so that even insects with a mutation affecting one target area remain vulnerable.

“It’s impossible for the insect to change the sequence that much and still survive,” Hunter says.

Despite their optimism, the researchers acknowledge that RNAi products aimed at the citrus psyllid won’t be ready for commercial use in the next few years.

“The industry needs a solution [for greening] quickly,” Cox says. “A genetics approach isn’t the quickest, but would be a long-term solution.”

“We’re looking at a five- to six-year window,” Hunter says, adding that five years is the industry’s target date for new solutions. “This will come in at that critical point.”

Florida’s citrus industry, which has helped fund the RNAi research among more than a hundred greening-related projects, has a keen interest in speedy results, says Andrew Meadows, communications director for Florida Citrus Mutual in Lakeland.

At the same time, growers recognize that genomics and RNAi technology represent a longer-term solution, Meadows says.

“The [RNAi] concept has been proven,” Shatters says. “I’m encouraged that in a couple years we’ll have something for the industry.”