LAKE ALFRED, Fla. — University of Florida Institute of Food and Agricultural Sciences faculty have secured over $16 million in new funding to advance research projects to combat the deadly citrus greening disease that threatens the Florida citrus industry.
The funding is awarded from the USDA National Institute of Food and Agriculture Emergency Citrus Disease Research and Extension (ECDRE) program, designed to provide solutions to U.S. citrus growers.
The funding — $16,382,191 — is spread out over eight projects, five of which are funded at $1 million or more and run from two to five years. Most of the projects are collaborations with UF/IFAS faculty from the Citrus Research and Education Center in Lake Alfred, the Southwest Florida Research and Education Center in Immokalee and the University of Florida in Gainesville. A few of the projects engage scientists in California, Texas and Connecticut.
“UF/IFAS is gratified by the confidence USDA NIFA is investing in us with these grants. It’s a demonstration of the leading role UF/IFAS plays — and must continue to play — in addressing the most pressing challenges facing the citrus industry,” said J. Scott Angle, UF senior vice president for agriculture and natural resources and leader of UF/IFAS.
The largest grant, awarded to Nian Wang, professor of microbiology and cell science, for nearly $8.6 million, supports the development and delivery of HLB disease management approaches by approaching it as a pathogen-triggered immune response disease.
The ultimate goal of this integrated project is to leverage the breakthrough discovery that HLB is a chronic immune disease to develop HLB management approaches for existing groves and non-transgenic HLB-resistant and -tolerant citrus varieties for long-term, sustainable HLB control. This project was also approved as a USDA NIFA Center of Excellence and was reviewed as outstanding.
Two projects led by Assistant Professor John Chater and Professor Jude Grosser, will look at rootstocks that appear to be HLB-tolerant and/or resistant to HLB. Another project led by Assistant Professor Fernando Alferez of the UF/IFAS Southwest Florida Research and Education Center, will focus on how to use a combination of individual protective covers (IPC) and brassinosteroids (BRs) — a plant growth regulator — for enhanced protection of citrus trees against HLB after individual protective covers are removed.
UF/ IFAS entomologists Kirsten Pelz-Stelinski and Lukasz Stelinski will work on two separate projects.
Pelz-Stelinki will work to provide a reliable, insect cell culture-based method for culturing of CLas bacteria, using a novel insect cell line approach. Upon completion of this project, Pelz-Stelinski hopes to have a culture system that serves as an essential research tool for increased understanding of CLas biology and for effective, rapid screening of antimicrobial agents against CLas.
Lukasz Stelinski’s project aims to create an artificial intelligence-driven release device for chemicals that attract Asian citrus psyllids to a killing device. The hope is that a dynamic complex smell generation approach can make the attract-and-kill strategy for psyllids feasible and practical.
Another project tackles the challenging task of gathering and organizing the vast amounts of research findings available to growers by developing tools for citrus industry stakeholders, the HLB-research community and research organization administrators. Outcomes of this national project led by Associate Professor Megan Dewdney of the UF/IFAS Citrus Research and Education Center, will be a database for stakeholders to access critical information about research findings on HLB and high-quality Extension products from existing and future knowledge sources.
Below is a list of received grants.
Development, evaluation, and delivery of citrus HLB management approaches by targeting its nature as a pathogen-triggered immune disease $8,589,573 million; 5 years
Principal Investigator: Nian Wang
Citrus HLB is a pathogen-triggered immune disease similar to sepsis in humans. Recent work demonstrated that CLas, the bacterium that cause HLB, stimulates a systemic and chronic immune response in the citrus phloem including reactive oxygen species (ROS) production, which causes systemic phloem cell death and subsequent HLB disease symptoms. The central hypothesis of this project is that HLB can be controlled by managing CLas-triggered ROS. Researchers propose that they will control HLB with three approaches:1) mitigating the production of ROS in HLB-affected groves with integrated horticultural measures; 2) protecting citrus plants from CLas-triggered ROS via CTV-mediated expression of antioxidant enzymes and silencing of key genes involved in CLas-triggered ROS production; and 3) generating non-transgenic HLB resistant/tolerant citrus varieties. They expect the interdisciplinary approach will increase production efficiency in existing HLB-affected groves and protect citrus production in all growing regions in the country by developing the first non-transgenic gene-edited citrus varieties resistant/tolerant to HLB.
A coordinated network for the improvement of HLB research and extension outputs $2,000,000; 2.5 years
Principal Investigator: Megan Dewdney
Until HLB was discovered in the United States, there had been relatively little research done on the system leaving huge knowledge gaps in areas like disease epidemiology, vector ecology, and physiological effects on citrus trees. Most of these gaps have been partially filled with the concentrated research effort of the last 15 years but the large amount of data produced has not always been synthesized and relayed back to the citrus industry. Citrus industry stakeholders are desperate for information but feel that relatively little has been provided compared with the immense research effort. There is a plethora of research studies being published but it is very difficult for researchers to easily find specific information. Ways to easily search or access HLB research are needed to address grower concerns. To complicate matters, the HLB community has non-standard ways of referring to data or apply terminology in multiple, conflicting ways. These can lead to confusion in the literature and when comparing findings when comparing work. This project will address these problems and assist USDA NIFA to refine research priorities for the future by developing science-based tools for citrus industry stakeholders and NIFA administrators.
HLB-resistant rootstock candidates for the citrus industry: Validating and understanding disease resistance $1,163,487; 2 years
Principal Investigator: John Chater
Huanglongbing (HLB) is a devastating disease in Florida and threatens the US citrus industry. The currently applied strategies for mitigating HLB are costly and unsustainable. The development and commercialization of HLB-resistant rootstocks proposed in this project is a possible solution for sustainable citriculture in the US. This project will investigate rootstocks that do not test positive for CLas and/or appear resistant to citrus greening. Experiments will be conducted on inoculated plants in the greenhouse and inarching experiments will be conducted in an infected grove. The long-term goals of this project are: 1) to distribute HLB-resistant rootstocks to stakeholders; and 2) to understand the biological mechanisms behind HLB-tolerance and resistance for breeding HLB resistant scions and gene editing purposes. The objectives of this project are: 1) verify HLB resistance of rootstock candidates and determine their impacts on scion performance; 2) transfer rootstock resistant germplasm material; 3) delivery of findings to stakeholders; and 4) study underlying mechanisms of HLB resistance in rootstocks. This project will gather horticulture data and multi-omic data to gain knowledge of the multidimensional underpinnings of HLB tolerance and resistance.
Targeted production of non-transgenic HLB-tolerant trees through complementary approaches $1,499,888; 2 years
Principal Investigator: Zhonglin Mou
On-going research projects have identified several highly promising targets for gene editing and/or gene silencing to create robust tolerance to Huanglongbing (HLB). The first goal here is to produce non-genetically modified (non-GM) HLB-tolerant citrus trees by editing or silencing the promising target genes. We will use a base co-editing strategy in which editing in the target gene can be selected by co-editing a citrus gene that leads to herbicide resistance. We will also employ an improved protocol to directly deliver Cas9/gRNA ribonucleoprotein particles into citrus protoplasts for gene editing. These strategies significantly increase the rate of transgene-free editing events. Since gene editing is highly challenging in some elite citrus cultivars due to their recalcitrance to both genetic transformation and protoplast regeneration, we will also use a citrus DNA-based (intragenic) vector to express microRNA in the rootstock. Because microRNA is capable of root-to-shoot long-distance movement, the expression of microRNA in the rootstock can silence target genes and induce HLB tolerance in the scion. The resulting intragenic rootstocks can readily be grafted with various scions. Our second goal is to educate the citrus communities about the non-GM base co-editing and intragenic microRNA technologies and their benefits to the citrus industry and consumers. This will help establish market acceptance for citrus products from the trees created using these new technologies.
Combining Individual Protective Covers (IPCs) and brassinosteroids to prolong health and improve fruit yield and quality in newly planted trees under HLB $800,000; 2 years
Principal Investigator: Fernando Alferez
Currently, Huanglongbing (HLB) is the main threat the citrus industry in the USA is facing and there is no cure available. For this reason, preserving health of newly planted trees is of paramount importance. The risk of new plantings to become infected is especially high for young trees, which flush frequently and attract more psyllids, thereby becoming more vulnerable to infection than mature trees. Individual Protective Covers (IPCs) are now being increasingly adopted to protect newly planted citrus trees from psyllid colonization. Even though IPCs effectively protect young trees from HLB, it is only for the first 2-3 years after planting, and eventually covers must be removed, exposing the trees to the disease vector. Under these circumstances, it would be advantageous to prolong tree health by means affordable to growers, so trees have several years of high-quality fruit production and increased yield. Brassinosteroids (Brs) are a natural group of hormones approved for use commercially in citrus in Florida. Brs have shown promise in reducing pests and diseases. In citrus, researchers have shown that HLB progression is delayed in young newly planted trees, psyllid colonization and egg deposition are reduced in new flushes, and rust mite incidence is reduced as well. There is evidence that in Valencia oranges, application of Brs can advance internal maturation by almost 1 month. This project will use a combination of IPCs and Brs for enhanced protection of citrus trees against HLB as well as other diseases that limit citrus production in Florida. The hypothesis is that Brs will prolong health in young trees after IPC removal. To test this hypothesis, and to develop an outreach program that conveys the results in a timely manner to stakeholders. The main objectives are to: 1) study the effects of IPCs, Brs, and their combined use on tree physiology, fruit production, and yield and quality, HLB incidence, and disease progression; 2) study the effects of Brs on the biology and behavior of the HLB vector (ACP) and on reducing incidence and pathogen transmission; 3) assess the effects of the combined system (IPC+Brs) on the incidence of other pests and diseases; and 4) effectively communicate project progress and results with stakeholders.
Toward a reliable, insect cell culture-based technique for culturing CLas bacteria $793,286; 2 years
Principal Investigator: Kirsten Pelz-Stelinski
Candidatus Liberibacter asiaticus (CLas), is the presumed causative agent of citrus greening, which has devastated citrus production in Florida and now threatens all citrus growing regions in the U.S., including Texas and California. There is currently no cure or durable remedy to combat citrus greening. While the ability to culture CLas in vitro would provide huge benefits for analysis of CLas biology and for fulfillment of Koch’s postulates to confirm that CLas causes citrus greening, attempts to culture CLas in the absence of other bacteria have failed. Our long-term goal is to provide a reliable, insect cell culture-based method for culturing of CLas bacteria. The goal of this proposal is to identify an ACP cell culture system for in vitro culture of CLas. Our objectives are: 1) test for CLas replication in axenic culture using optimized insect cell culture media; 2) assess hemipteran insect cell culture systems for CLas replication; and 3) establish cell lines from CLas-positive psyllids. Once a CLas culture is established, we will test for ACP transmission of cultured CLas to healthy citrus, and whether inoculation of healthy citrus results in citrus greening disease. Upon completion of this project, we will have a culture system that serves as an essential research tool for increased understanding of CLas biology and for effective, rapid screening of antimicrobial agents against CLas.
Accelerating the delivery of conventionally developed HLB tolerant citrus scions and rootstocks as pathogen-free budlines for replicated multi-site testing $535,957; 2 years
Principal Investigator: Jude Grosser
The project is focused on accelerating the delivery of conventionally developed HLB-tolerant citrus scions and rootstocks as pathogen-free budlines for replicated multi-site testing compared with the existing system. Commercial use of pathogen-free, high-quality and unique HLB-tolerant scions and rootstocks will provide Florida growers with more sustainable and profitable alternatives to replace the hundreds of acres currently going out of production due to HLB. Future plantings will be less susceptible to tree decline and crop losses, creating a more stable and expanded citrus industry. Growers, processors, packers, and marketers will all benefit, and the project will also accelerate the delivery of new, unusual, and lucrative niche-market varieties. By using pathogen-free HLB-tolerant scion/rootstock combinations, fruit production costs including pesticide and fertilizer applications could be reduced which would have positive impacts on the environment. In addition, pathogen-free material generated from this project will include new selections that might allow for an expansion of citrus production into areas where citrus is not currently planted at a commercial level (i.e., new cold-hardy selections for north Florida) and other states. The project will utilize extensive collections of novel citrus germplasm being generated by the UF/IFAS Citrus Research and Education Center breeding team, including many that have the potential to create new niche/specialty marketing opportunities for national citrus growers. The development of a reliable rapid protocol for generating pathogen-free material with no rejuvenation is necessary for the timely delivery of these new scions and rootstocks to our citrus industry. The successful adoption of this technology will reduce the current clean-up time by 1.5-2 years, and significantly increase the number of selections that can be processed.
A method for generating an optimally attractive scent for Asian Citrus Psyllid (ACP) biocontrol $1,000,000; 2 years
Alexander Aksonov is lead investigator at University of Connecticut with Lukasz Stelinski from University of Florida as co-investigator; UF/IFAS to receive $168,734.
Huanglongbing (HLB) disease associated with the bacterium Candidatus Liberibacter asiaticus (CLas) is wiping out citrus worldwide. A primary strategy to combat HLB is intense insecticide sprays to suppress populations of the CLas’ invasive insect vector, the Asian citrus psyllid (ACP). A far better strategy would be to use chemical lures to capture and kill the insects. Unfortunately, pheromones do not appear to attract ACP. Mixtures of volatile organic compounds (VOCs) that mimic volatiles emitted from plants have been shown to hold great promise as an attractant, but these mixtures need to be highly complex and insect behavior changes with minor shifts in specific compounds’ concentrations. Consequently, prior efforts to develop effective VOC-based lures have not yet delivered a practical solution. This is mainly due to technological limitations, as it is challenging to disperse more than a few compounds in a controlled fashion. However, simple mixes, unlike natural VOC profiles, do not elicit sufficient attraction of ACP. To address this challenge, this project will dynamically optimize complex attractant VOC mixtures to achieve a potent effect on ACP. This is made possible by the breakthrough in development of graphene-based sorbent materials that will form the basis of VOC dispersion device. The new material enables delivering flexible dynamically controlled complex mixtures of VOCs for attractiveness optimization. The same material can be used for dispersion of ACP attractants in citrus orchards. In conjunction with AI-driven optimization strategies, the dynamic complex smell generation approach can make the attract-and-kill approach for ACP both feasible and practical.
–Ruth Borger, UF/IFAS