A Solution for the devastating bacteria causing citrus huanglongbing?
UC Riverside scientists have found the first substance capable of controlling Citrus Greening Disease, which has devastated citrus farms in Florida and also threatens California.
Oranges afflicted with Citrus Greening Disease. (UCR)
The new treatment effectively kills the bacterium causing the disease with a naturally occurring molecule found in wild citrus relatives. This molecule, an antimicrobial peptide, offers numerous advantages over the antibiotics currently used to treat the disease.
UCR geneticist Hailing Jin, who discovered the cure after a five-year search, explained that unlike antibiotic sprays, the peptide is stable even when used outdoors in high heat, easy to manufacture, and safe for humans.
“This peptide is found in the fruit of greening-tolerant Australian finger limes, which has been consumed for hundreds of years,” Jin said. “It is much safer to use this natural plant product on agricultural crops than other synthetic chemicals.”
Currently, some growers in Florida are spraying antibiotics and pesticides in an attempt to save trees from the CLas bacterium that causes citrus greening, also known as Huanglongbing or HLB.
The Asian citrus psyllid, pictured here, spreads the bacterium that causes Citrus Greening Disease. (Mike Lewis/UCR)
“Most antibiotics are temperature sensitive, so their effects are largely reduced when applied in the hot weather,” Jin said. “By contrast, this peptide is stable even when used in 130-degree heat.”
Jin found the peptide by examining plants such as the Australian finger lime known to possess natural tolerance for the bacteria that causes Citrus Greening Disease, and she isolated the genes that contribute to this innate immunity. One of these genes produces the peptide, which she then tested over the course of two years. Improvement was soon visible.
“You can see the bacteria drastically reduced, and the leaves appear healthy again only a few months after treatment,” Jin said.
Because the peptide only needs to be reapplied a few times per year, it is highly cost effective for growers. This peptide can also be developed into a vaccine-like solution to protect young healthy plants from infection, as it is able to induce the plant's innate immunity to the bacteria.
Jin's peptide can be applied by injection or foliage spray, and it moves systemically through plants and remains stable, which makes the effect of the treatment stronger.
The treatment will be further enhanced with proprietary injection technology made by Invaio Sciences. UC Riverside has entered into an exclusive, worldwide license agreement with Invaio, ensuring this new treatment goes exactly where it's needed in plants.
“Invaio is enthusiastic to partner with UC Riverside and advance this innovative technology for combating the disease known as Citrus Greening or Huanglongbing,” said Invaio Chief Science Officer Gerardo Ramos. “The prospect of addressing this previously incurable and devastating crop disease, helping agricultural communities and improving the environmental impact of production is exciting and rewarding,” he said. “This is crop protection in harmony with nature.”
The need for an HLB cure is a global problem, but hits especially close to home as California produces 80 percent of all the fresh citrus in the United States, said Brian Suh, director of technology commercialization in UCR's Office of Technology Partnerships, which helps bring university technology to market for the benefit of society through licenses, partnerships, and startup companies.
“This license to Invaio opens up the opportunity for a product to get to market faster,” Suh said. “Cutting edge research from UCR, like the peptide identified by Dr. Jin, has a tremendous amount of commercial potential and can transform the trajectory of real-world problems with these innovative solutions.”
UV Riverside news release:
Jules L Bernstein
Senior Public Information Officer
An Automated Delivery System for Therapeutic Materials to Treat HLB Infected Citrus
Ozgur Batuman1 and Louise Ferguson2
¹Southwest Florida Research and Education Center, University of Florida, Immokalee, FL; 2UC Davis, Department of Plant Sciences, University of California Davis, Davis CA
Why is this research needed?
In 2005, a disease called Huanglongbing (HLB, citrus greening, was identified in Florida's commercial citrus groves. The disease is caused by a bacterium that affects all citrus cultivars by disrupting the flow of nutrients from the source of production, to the site of use, causing tree decline. HLB weakens the root system, increases early fruit and leaf drop, lowers tree productivity and fruit quality and ultimately kills the tree. The disease has spread to all the major production regions in Florida. Economic losses have exceeded more than $4 billion dollars. Currently, more than 95% of Florida's trees are infected. There is currently no cure for the disease.
Efforts to control HLB have been unsuccessful as the bacterium cannot be cultured, literally grown, in a petri dish, and once in the plant it proliferates within the citrus phloem. Phloem is the system that transports sugars from their site of production, the leaves, to plant parts that use sugars, the roots or flowers.Phloem transport is generally downward but can be upward as well.
Once the HLB bacterium is in a tree's phloem it has the potential to infect the entire tree. It is exceedingly difficult to introduce any control agent into the phloem with the conventional control methods of foliar spraying or soil drenching.
Thus far, no treatment preventing HLB infection, or controlling the bacterium once within the tree, has been developed. Potential chemicals are being investigated, but in order to test them, direct or indirect phloem delivery, where the bacterium proliferates, is needed. Therefore, an effective method of delivering an effective volume of theraputics into the phloem is needed to evaluate potential treatments.
What is the focus of this project?
Our project focuses on developing a method of delivering therapeutic liquid materials, bactericides, microbial metabolites, RNAi, or biologicals, into the citrus vascular tissues, both the xylem which conducts water and nutrients upward from the roots and the phloem, which conducts sugars and other metabolic products downward from the leaves. We are investigating diffusion, trunk punctures with a surrounding liquid reservoir for passive uptake and infusion, low pressure active injections. We are focusing on these methods as foliar sprays and root drenches have not been successful phloem delivery methods.
Who will be doing the research?
The project is led by plant pathologist Dr. Ozgur Batuman with colleagues at the Southwest Florida Research and Education Center (SWFREC) at University of Florida in Immokalee. This four-year project will also study the citrus vascular system with a multidisciplinary research team including UF Plant Pathologists Drs. Nabil Killiny and Amit Levy at Lake Alfred, SWFREC UF Plant Physiologist Ute Albrecht, Citrus Horticulturist Fernando Alferez, Precision Ag. Engineer Yiannis Ampatzidis, Agricultural and Natural Resources Economist Tara Wade, University of California-Davis Extension Specialist Louise Ferguson and Texas A&M-Kingsville Citrus Center Plant Pathologist Veronica Ancona as well as number of graduate students, postdocs, and Florida, Texas and California citrus industry members.
How will this research be done?
Our earlier research involving comparisons of delivery methods including foliar sprays, soil drenching and trunk injection determined Needle-Assisted Trunk Infusion (NATI) was the best potential delivery method (Figure.1). In initial experiments, using NATI, 1 ml of rhodamine (1%) dye was injected into the trunks of one-year-old citrus seedlings. A visible red color, indicative of rhodamine uptake and movement, was detected in the upper-most leaves within 30-60 min and an increase in color intensity was observed within 24 hours. Similar results were observed in two-year-old grafted Valencia plants within 48 hours. If the NATI delivery method can be automated, large numbers of trees could be treated quickly. Once the delivery method has been developed, implementation will be tested with potential treatments developed within other research projects.
Our proposed automated delivery would consist of a robotic arm with several modules at the end of the arm, installed on an ATV or tractor. One module with needles would grip and puncture the trunk, a second module would wrap a reservoir around the trunk below the punctures and third module would fill the reservoir. (Figure 2). Hopefully, a robotic arm plus automated system will be inexpensive enough for growers to purchase and simple enough to use.
Another approach is disease prevention; application pf prophylactic chemicals that prevent infection. In this scenario our system would be used treat healthy young trees with bactericides or boost their immune system. When infected by the ACP the bacterium would either be killed or suppressed, perhaps below the level that harms tree growth and productivity. This option is analogous to the vaccinations that prevent diseases in humans and animals.
What are the greatest challenges and opportunities.
The greatest challenge is successful phloem delivery. The greatest opportunity is that, if successful, we will have developed a method that will allow much more precise deliver of theraputics to citrus trees. For example, if an effective phloem delivery method is developed, it could be used to control insects that feed on citrus plant parts. Or, it could be used to deliver growth regulators, perhaps nutrients and carbohydrates, to roots and fruits to increase growth, development and fruit quality; much like an intravenous injection functions in an animal.
Among the questions we hope to investigate are:
- When, what kind of, and what amount of therapeutics can be applied by NATI?
- At what frequency?
- What type of citrus tree: cultivar, age, infected, healthy is the best for treatment by NATI?
- Can we kill the bacterium? How and when to assess a change in bacteria titer after treatment?
- When will become available and be economically feasible for growers?
Figure 1. Distribution of rhodamine (red dye; 1%) applied by NATI in various tissues (left) of grafted and non-grafted young citrus plants grown in the greenhouse (right). Photos taken 2 weeks after the treatments. Treatments and tissues observed are indicated. Yo = year-old.
Figure 2. Projected automated delivery system (ADS); an ATV with extendable arm with NATI and the cover placement systems on the arm guided onto the tree trunk (upper panel), and closeup of NATI and cover placement system (panel below).
The United States Department of Agriculture National Institute of Food and Agriculture (USDA-NIFA) Grant # 2019-70016-29096.
For more information, please visit this project's dedicated website:
Citrus greening disease, also called huanglongbing (HLB), is a bacterial infection of citrus trees that results in small, misshapen and sour fruits that are unsuitable for consumption. The disease ultimately kills the tree.
Because there is no cure, HLB is a major threat to the $10 billion citrus industry in Florida, where it was first detected in 2005,and to the $7 billion industry in California, where it appeared last year.
Researchers from the Boyce Thompson Institute (BTI), the U.S. Department of Agriculture Agricultural Research Service (USDA-ARS) and the University of Washington investigated a seemingly unlikely source of biocontrols for HLB: neuropeptides found in Asian citrus psyllids, the insect that carries the disease-causing bacterium Candidatus Liberibacter asiaticus (CLas), which it spreads while feeding on a tree's leaves and stems.
The research team, led by BTI faculty member Michelle Heck, published its findings on Feb. 10 in the Journal of Proteome Research.
Laura Fleites, a research associate in Heck's group at BTI, focused on neuropeptides because they function as hormones in hemipteran insects – a class that includes psyllids, aphids, whiteflies, shield bugs and other crop-plaguing species – to regulate growth, development and other biological functions.
“If we could develop an insecticide that is specific for Asian citrus psyllids based on one of the insect's own neuropeptides, then we could protect citrus trees from the insect that spreads CLas,” said Heck, a USDA-ARS research molecular biologist and an adjunct associate professor in the School of Integrative Plant Science, in the College of Agriculture and Life Sciences. “Citrus greening disease is devastating our citrus industry, and we need to develop new ways for our citrus growers to control it.”
Fleites and the team characterized the full array of peptides found in the psyllids and identified 122 potential neuropeptides. While promising, the findings offer only potential starting points for combatting HLB, because unmodified insect-derived neuropeptides are not suited for use as insecticides in the citrus grove.
To turn the findings into a usable product, the team is now part of a collaboration aimed at identifying the best psyllid-derived neuropeptide for development. The team will then stabilize the peptide and decide the optimum method for delivering the insecticidal molecule to citrus trees – whether as a spray, by engineering trees to make the peptides themselves, or by some other method.
For the study, Fleites developed new extraction and analysis methods other researchers could use in their investigations of insect peptides.
“Thanks to USDA support, I was able to develop techniques that enable the identification of small, functional insect peptides separately from their larger, inactive precursors,” Fleites said. “Because these techniques aren't specific to psyllids, they may be useful for identifying neuropeptides in other hemipteran insects to protect crop plants.”
Heck said this work shows how USDA grant programs can be used to fund everything from discovery to product development. The neuropeptide discovery was done in a study funded by the National Institute of Food and Agriculture; the translational research is being done under a grant from the Animal and Plant Health Inspection Service.
Funding for this work came from a USDA Specialty Crops Grant and from the USDA-ARS.
Michael J. Haas is a freelance writer for the Boyce Thompson Institute.
Asian citrus psyllids feed on a citrus tree. The psyllids deposit a bacterium in the sap that causes citrus greening disease, a scourge to the citrus industries in Florida and California, worth a combined $17 billion.
New technique has potential
to protect citrus from HLB
Citrus greening, also called Huanglongbing (HLB), is devastating the citrus industry. Florida alone has experienced a 50 to 75 percent reduction in citrus production. There are no resistant varieties of citrus available and limited disease control measures.
Some scientists think it is possible that orange juice could one day become as expensive and rare as caviar. In an effort to prevent this, three plant pathologists at the University of California-Berkeley and United States Department of Agriculture conducted research into ways to boost citrus immunity and protect the valuable fruit against citrus greening.
Because the bacteria that causes citrus greening cannot be grown in a lab, scientists have to find novel ways to conduct experiments. The University of California-Berkeley/USDA team looked at many different strains of the bacteria that cause citrus greening to see if they could identify peptides (a compound of two or more amino acids) that would trigger immune responses.
"This was a long list, so we narrowed it down by selecting small peptides that were a bit different in their peptide sequence, which might imply that the bacterium had made those sequence changes so that they wouldn't be recognized by the plant immune system," explained Jennifer D. Lewis, group leader of the research team. "Then we further narrowed that list to peptides from strains that caused disease in citrus."
Through this research, they showed that two peptides could trigger immune responses in multiple plant species, including citrus. These peptides may play a role in preventing or reducing yield loss from citrus greening.
According to Lewis, "We thought it was particularly interesting that some of the peptides predicted to elicit a response, could actually trigger immune responses in multiple plant species. This suggests that the immune response to these peptides is conserved across species."
IMMOKALEE, Fla. — University of Florida scientists are working toward establishing a new biological method that may help farmers control the insect that transmits the deadly greening disease into citrus trees. Greening is present in about 95 percent of the citrus trees in Florida, so by using a virus that may kill the insect, growers may be able to reduce the need for pesticides.
Instead of spraying insecticides, scientists hope to harness the natural enemies of the tiny Asian citrus psyllid to manage the invasive pest, said Ozgur Batuman, a plant pathologist at the UF/IFAS Southwest Florida Research and Education Center in Immokalee.
Batuman is leading a UF/IFAS research team investigating viruses that live in the digestive system of the psyllid. They want to see whether any of them can weaken or kill the insect.
The psyllid transmits the bacterium responsible for greening by feeding on a citrus tree's new shoots and leaves.
UF/IFAS scientists are getting promising early results with this research. Their latest research suggests a natural process that would kill the insect or at least prevent greening from being transmitted to citrus trees.
“This invasive pest is now established throughout Florida's commercial groves, so growers use insecticide sprays that are potentially harmful for the environment as one of their primary tactics for fighting citrus greening,” Batuman said. “By reducing psyllid populations in their groves, growers hope to increase the quality of fruits and the productive lives of their trees by minimizing the number of times a tree is exposed to the greening pathogen.”
To arrive at their results, Batuman and his team spent two years collecting psyllids from commercial citrus groves in 22 central and south Florida counties – from Lake and Orange counties in the north to Martin County in the southeast to Collier County in the southwest.
They identified viruses within the psyllids' bodies.
By using a test that amplifies DNA, scientists found five viruses in the guts of the psyllids they collected. They believe they can use those viruses to control the psyllids that live in the very groves from which scientists collected them.
With these experiments, for the first time, scientists have taken big strides toward identifying viruses associated with the Florida psyllids. They also now better understand the presence of those viruses in the psyllids. Asian citrus psyllids first arrived in southeast Florida in 1998.
“Future experiments will investigate how these viruses can be manipulated so that they may alter the pysllid's biology,” said Batuman. “We also need to find out how the viruses affect survival and transmission of the bacterium that causes citrus greening disease, also known as Huanglongbing, or HLB.”
“By understanding how the viruses interact with the psyllid biology, we may be able to understand how to better control the psyllids,” he said.
By: Brad Buck, 813-757-2224, firstname.lastname@example.org, 352-875-2641 (cell)