Hairy root matrix used to culture CLas to rapidly screen antimicrobials for improved HLB management
Article written by Kranthi Mandadi, Veronica Ancona, Steve Futch, Elizabeth Grafton-Cardwell, Peggy G. Lemaux, Lukasz Stelinski, Monique Rivera & Sara Garcia-Figuera.
Revised May 20, 2019.
What is the technique?
Most plant pathogens can be studied in the laboratory. The ability to keep pathogens in petri dishes or test tubes containing nutrients in the laboratory allows the pathogen to be easily manipulated and studied. A major difficulty in studying Candidatus Liberibacter asiaticus (CLas), the bacterium thought to cause huanglongbing (HLB), has been that the bacterium can not be grown using these traditional laboratory methods. The inability to culture the HLB bacterium, CLas, in the lab makes it very difficult to develop cures for the disease. Our research team has developed a novel technique for laboratory cultivation of CLas, using so-called hairy roots, which are plant tissues that mimic a microbe’s natural environment. Hairy root cultures support CLas bacterial growth and these cultures can be used to screen antimicrobials to find compounds that control and/or suppress CLas. Once discovered, antimicrobials could be applied to the foliage or injected into the trees to prevent establishment or reduce the impact of HLB in citrus trees. The beauty of the hairy root method is that it allows researchers to rapidly screen many antimicrobials (high throughput) and urgency is necessary in combating this deadly disease.
What are the challenges and opportunities?
Currently, several antimicrobial compounds are being tested against HLB, including bactericides (antibiotics) and compounds that stimulate tree defense systems. To evaluate efficacy, the conventional, time-consuming approach is to perform foliar (leaf) sprays, trunk injections, or soil drenching, using the antibiotic, or to screen against related pathogens, such as Liberibacter cresens, that can be minimally cultured in the laboratory. However, the effectiveness of these approaches can be inconsistent and costly, especially when trying to work with mature trees, limiting the ability of researchers to screen the vast number of chemistries that exist. Our proposed CLas culturing in hairy roots and screening pipeline protocol will allow evaluation of new antimicrobial compounds (e.g., small molecules, disease resistance and susceptibility genes/proteins) against CLas much faster (higher throughput) and at a lower cost, thus saving a significant amount of resources and time. Hastening implementation of a possible successful treatment and/or therapy is critical to solving the HLB problem for the citrus industry.
Who is working on this project?
Kranthi Mandadi, Assistant Professor, Plant Pathology and Microbiology, Texas A&M AgriLife Research & Extension Center, Weslaco; San Fernando, Professor, Biological and Agricultural Engineering, Texas A&M University; Veronica Ancona, Assistant Professor, Texas A&M-Kingsville Citrus Center; Gitta Coaker, Professor, University of California (UC), Davis; Elizabeth Grafton-Cardwell, Research Entomologist, UC Riverside and Director of Lindcove Research and Extension Center; William Dawson, Professor, University of Florida (UFl) Citrus Center; Stephen Futch, Extension Agent IV, UFl Citrus Research and Education Center; Mike Irey, Director of Research and Business Development, Southern Gardens Citrus, Florida.
Funding source: This study was supported by funds from USDA-NIFA-AFRI and Texas A&M AgriLife Research Insect-Vectored Diseases Seed Grant.