Cilia Lab at International Citrus Greening Conference

Assistant Professor Michelle Cilia and four members of her lab attended the International Conference for Huanglongbing last week. They joined hundreds of other researchers in Orlando, Fla. Feb. 9-13, to present their research on this disease, which causes citrus greening and is threatening Florida’s citrus crops.

Huanglongbing prevents oranges and other citrus fruits from ripening, resulting in bitter green fruit. Scientists believe that a bacterial species called C. Liberibacter asiaticus causes the disease, which is spread by a sap-sucking insect called the Asian citrus psyllid. Once the bacteria infect a tree, it can take up to three years to show symptoms, giving the organism more time to spread through the orchard before anyone is aware of the problem. The disease is fatal to citrus trees and has no known treatment.

At the conference, Cilia, who is also a Research Molecular Biologist at the USDA Agricultural Research Service, gave a talk on how C. Liberibacter asiaticus, or CLas, manipulates the Asian citrus psyllid to promote its own spread. She made this discovery using proteomics, which is the study of all the proteins within a cell, tissue or organism. She found that the bacteria cause metabolic syndrome in the insect, which she hypothesizes will trigger it to behave in ways that spread CLas throughout the grove. Her lab has developed a new collaboration with BTI Associate Professor Frank Schroeder, to further explore this idea.

Jared Mohr, a junior chemistry major at Cornell University, presented a poster on the use of proteomics for early detection of HLB. Researchers typically try to detect CLas DNA in tree leaves, but the bacteria are not evenly distributed throughout the tree. For these tests to be successful, the researchers must be lucky enough to pick the infected leaves, or the test could give a false negative result.

Instead of looking for CLas directly, Mohr used a mass spectrometer to find proteins that the whole tree makes to respond to an infection. By analyzing leaves from infected and uninfected trees from the Contained Research Facility at the University of California, Davis, he found that the trees change their production level of dozens of proteins—out of more than 3,000 detected—but the work is still preliminary.

Mohr presented his work on optimizing this technique and his use of a method called machine learning, which can take proteomic data from infected and uninfected plants and then develop an algorithm to decide if a new sample has the bacteria or not.

“You can just feed it proteomic data,” said Mohr. “It develops a scoring algorithm that assigns a tree some probability of being infected.”

In recognition of Mohr’s excellent work, the American Phytopathological Society awarded him the 2015 Frank L. Howard Undergraduate Research Fellowship, given each year to a single undergraduate. He will present his work again this summer at the APS meeting in Pasadena, California.

John Ramsey, a postdoctoral researcher in the lab, gave a talk on his work using Protein Interaction Reporter technology to find critical proteins that CLas uses to infect the Asian citrus psyllid.

“The disease is acquired by the insect from feeding on phloem sap of infected trees,” said Ramsey. “For the pathogen to be transmitted to a healthy plant, the pathogen first has to travel through the psyllid, crossing multiple cell membranes and evading the insect immune response.”

Protein Interaction Reporter technology allows him to fish out key proteins that CLas uses to start a successful infection and get detailed information on how these proteins interact. Collaborators in the lab of James Bruce, at the University of Washington, developed the technology. Other members of Cilia’s lab are using the technique to study viruses transmitted by aphids.

“It’s a cutting edge approach that has no parallel among other protein interaction discovery approaches,” said Cilia.

“The downstream application will be to design inhibitors to interfere with that interaction,” said Ramsey. “This is a control strategy to specifically perturb protein interactions that are critical to successful transmission of the pathogen.  Ultimately we are working to deliver a treatment to growers which would render the insect vector incapable of transmitting the pathogen to their trees.”

Jaclyn Mahoney, a research assistant, is taking a different approach with her investigations into HLB. She presented her work on psyllids raised on orange jasmine. These plants don’t usually carry the bacteria and when psyllids feed on it they tend to have low levels of the bacteria, and are less likely to pass it on. No one knows why this is the case, but Mahoney is looking for differences in the proteins generated by psyllids when fed on orange trees, compared to when they eat orange jasmine.

“This is more of a discovery proteomics experiment to figure out what are the differences between psyllids raised on these different plants,” said Mahoney. “Hopefully we’ll be able to see some interesting proteins that might have a very strong influence on the ability to vector Liberibacter.”

Annie Kruse, a new graduate student in the lab enrolled in the Plant Pathology and Plant-Microbe Interaction Section at Cornell, also attended the meeting. Kruse solidified a new collaboration with Robert Shatters, a USDA Agricultural Research Service Molecular Biologist and psyllid expert. She also interacted extensively with citrus growers, gaining valuable experience communicating science to the grower community. Kruse concluded the meeting with a visit to the USDA Horticultural Research Lab in Fort Pierce, Florida to learn techniques from the Shatters’ lab that will enable her to complete her thesis research.

The Cilia lab receives funding from the California Citrus Research Board, the Boyce Thompson Institute, the USDA Agricultural Research Service and the USDA SCRI program for molecular research on HLB.