Location: Subtropical Insects and Horticulture Research
Project Number: 6034-22320-006-004-S
Project Type: Non-Assistance Cooperative Agreement
Start Date: Sep 19, 2025
End Date: Sep 17, 2027
Objective:
This research represents the final stage of developing commercializable methods to deliver biologically-based therapeutic molecules to control citrus greening disease (HLB) that can be deployed to trees in the field. We are very excited about the Symbiont technology that we use to develop a localized group of host plant cells that are modified to produce natural biological molecules that can either directly defend against the development of disease symptoms or induce natural plant defenses that block disease symptom development. This new strategy can be delivered to either treat HLB-symptomatic citrus trees or prevent development of HLB symptoms and is based on a novel way of deploying Agrobacterium tumefaciens as a beneficial strategy to protect crop plants. We are also excited because the proposed research requires the multi- and trans-disciplinary merging of biology, molecular biology and materials fabrication to develop a cost effective and rapid inoculum delivery method, and we do this by working closely with the Indian River State College Advanced Manufacturing Hub and private industry with expertise in chemical delivery to trees (ArborSystems, Inc.).
Approach:
Develop symbiont inoculation/containment devices for economic and effective commercial deployment of Symbionts. We have demonstrated the ability to form Symbionts on citrus, but the current methods are not economically viable for commercial use, so we will adapt the previously described EZ-Ject Lance for this purpose and the President of the company that manufactures and sells this device is a collaborator on this proposal. The lance is optimized for delivering small volumes of herbicide using a .22 caliber brass shell casing as the delivery cell. We will optimize this delivery cell to support containment and delivery of the Symbiont inoculum through development of design and fabrication teams composed of biology and Advanced Manufacturing hub students from the Indian River State College (IRSC). PI and other USDA, ARS scientists have routinely worked with IRSC biology/biotechnology students as interns on agriculturally related projects including the previous NIFA ECDRE grant through collaboration with Indian River College. Through collaboration with IRSC Advanced Manufacturing Hub and PI and Director of the Center for Laser and Fiber Optics Education, we will develop transdisciplinary teams of biologists and fabrication students to design and manufacture variants of the delivery cells for inoculum delivery. The manufacturing students will provide capability in areas of CNC milling and lathing, 3D printing and general fabrication. The student teams will jointly develop prototype delivery cells that will be tested at the USDA ARS USHRL research farm for delivery to field citrus trees. Agrobacterium packaging: To determine the best way to deliver Agrobacterium we will evaluate two main strategies: 1) lyophilization coupled with double chamber delivery method where the Agrobacteria are present in a lyophilized state in one chamber and the activation medium is present in a second chamber. This system will be designed so that introduction of the delivery cell into the tree will break a membrane between the two chambers and allow the bacteria to flow with the media into the area of the damaged cambial cells. 2) The second method will be to imbed the Agrobacterium in a hydrogel formulation containing the desired media components as described for lyophilization. Testing of the devices will be initially performed by PI at the USDA, ARS USHRL research farm on field trees using wild-type Agrobacterium.