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Metabolite changes in the tree can help us detect Huanglongbing

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Research by Dr. Carolyn Slupsky, University of California, Davis
Article written by Carolyn Slupsky, Elizabeth Chin, Elizabeth Grafton-Cardwell, Peggy G. Lemaux, & Lukasz Stelinski.
Revised November 13, 2019

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What is the technique?

Metabolism is all of the processes an organism carries out in order to stay alive. There are two types of molecules involved in an organism’s metabolism: proteins and small molecular weight chemicals called metabolites. These molecules change in abundance when the metabolism of a living organism, such as a human, insect, or tree, is altered. Measuring these molecules provides a ‘snapshot’ of an organism’s metabolism. During infection, an organism’s metabolism is busy responding to the pathogen, as it works to protect itself. Thus, the ‘metabolite profile’ is different for healthy versus infected organisms. Carolyn Slupsky is developing methods to measure metabolite profiles of citrus trees, and to identify one that indicates early CLas (the bacteria that causes HLB) infection.

How can metabolites be used to identify infected trees?

Early detection of CLas infection is critical to removing infected citrus trees and reducing HLB spread. The plant response to CLas occurs throughout the plant soon after infection, even before symptoms of HLB appear in leaves, twigs, fruit or roots. The changes in metabolite composition can be measured and used to identify infected trees. The Slupsky Lab uses a technique called 1H NMR (“proton NMR”) spectroscopy to measure these changes in metabolites. Leaves are dried and ground, and metabolites are extracted using specific chemicals. The resulting extract is run on the NMR, and the output- called a ‘spectrum’- is analyzed on a computer. Each spectrum has many peaks, and each peak or group of peaks corresponds to specific metabolites. This includes sugars (e.g., glucose and sucrose), amino acids (the building blocks of proteins), and other small molecules. The area under each peak provides information about the metabolite concentration, so the pattern of metabolite peaks during CLas infection defines the metabolite profile (Fig. 1). Metabolite profiles of unknown leaf samples can be compared to known profiles of CLas-infected and non-infected leaves. Then, unknown leaf samples can be categorized as infected or non-infected. This method, requiring only one leaf per tree, allows for rapid and noninvasive sampling.

Who is working on this project?

Carolyn Slupsky, a Professor in the Department of Food Science and Technology at the University of California, Davis, and her research team are using these methods to establish and validate metabolite-based markers for early detection of HLB.

What are the challenges and opportunities?

The main challenge is field validation, because truly healthy, non-CLas-infected trees are not guaranteed in the field if the psyllid vector is present. However, this method has been successful in correctly identifying CLas-infected field trees in Texas, and more field trials using screen-protected healthy control trees are underway. Preliminary data shows that the metabolite profiles of leaves infected with citrus tristeza virus (CTV), canker, or citrus stubborn infections are different from the metabolite profile of leaves infected with CLas. Although sampling is as easy as clipping one leaf from a tree, analysis of the NMR spectrum is currently a bottleneck because this method is only semi-automated, and requires lab researcher input. Automation of NMR spectral analysis would help turn this into a high-throughput method. 

Funding source: This project is funded by the Citrus Research Board and a USDA-NIFA Hatch grant.

Watch video about HLB, created by Dr. Slupsky’s lab.