Name: Attachment of the Direct Plant Infusion (DPI) Device to Citrus Plants for Compound Injection
Uploaded: 2023-06-02T14:30:00
Duration: 1 min 38 s
Description: Watch this Scientific Journal Video about Attachment of the Direct Plant Infusion DPI Device to Citrus Plants for Compound Injection at JoVE.com

The video demonstrates how to propagate small potted citrus trees and attach a 3D-printed Direct Plant Infusion (DPI) device to their trunks. It includes instructions for creating a hole in the trunk, fitting the device, and ensuring proper alignment for effective compound delivery, using specific materials like silicone tape.

attachment of the direct plant infusion (dpi) device to citrus plants for compound injection

Molekylleverans i växter med hjälp av en direkt infusionsanordning

The management of plants in commercial and landscape settings often requires the use of chemical compounds to optimize the plant growth and health. How these molecules are delivered depends on the type of molecule, the function of the molecule, the type of plant, and the management system in place. Foliar and soil applications are the easiest delivery strategies, but limitations in the uptake of some molecules necessitate direct delivery. An example of these molecules is therapeutic molecules that function best when they move systemically within the plant but cannot be effectively delivered by simple topical applications1. This is the case with Huanglongbing (HLB), also called citrus greening disease. HLB is a disease associated with a phloem-limited bacterium, Candidatus Liberibacter asiaticus (CLas), which cannot be cultured outside of the plant, or its insect vector, Diaphorina citri Kuwayama (D. citri)2.
If the putative therapeutic molecules are gene products, they can be tested by creating transgenic plants expressing these compounds. However, transgenic plant production can be time- and resource-intensive, is highly genotype-dependent, and can be inhibited by gene silencing3. In addition, even if these transgenics show promising results, regulatory and public perception constraints reduce the likelihood of their commercial acceptance4,5. The exogenous application of compounds, however, simplifies the testing of biological and synthetic molecules because it does not require the production of stable or transiently expressing transgenic plants, which reduces the time and resources for testing a molecule&#39;s effects. A method for the effective and efficient systemic plant delivery of exogenous compounds could be used for a wide variety of research and screening purposes.
One of these applications is the analysis of systemic molecule movement within a plant&#39;s vascular system, which can be done using trackable markers, whether they are fluorescent, visible, or unique chemical isotopes6,7,8,9. One commonly used fluorescent marker is 5,6-carboxyfluorescein-diacetate (CFDA), which is a membrane-permeable dye that is degraded by intracellular esterases into 5,6-carboxyfluorescein (CF) and, subsequently, becomes fluorescent and membrane-impermeable10. CFDA has been extensively used to monitor phloem transport, sink and source relationships, and vasculature patterning in plant tissue11,12.
In addition to these markers, certain compounds may directly alter the plant&#39;s physiology to increase the productivity or kill the plant in the case of herbicides. Both insecticides and antimicrobial compounds are a means of increasing plant productivity, especially in the presence of HLB. An example of an antimicrobial molecule that is used to control CLas is streptomycin. Streptomycin is an aminoglycoside antibiotic that was originally isolated from Streptomyces griseus and has been shown to inhibit bacterial growth through the inhibition of protein biosynthesis13. In terms of insecticides, the main target for HLB research is D. citri, which transmits CLas from tree to tree14. For this purpose, neonicotinoids, such as imidacloprid, are commonly utilized, as they are the gold standard for controlling insect pests15. All these varied uses are important aspects of current plant management strategies, and the development of novel products is dependent on efficient screening assays.
One method that is used for the introduction of compounds into woody plants is direct injection into the trunk. A variety of systems have been designed that vary in their needs for predrilled injection sites, and these systems utilize either pressure-based injection or passive flow16. Although pressure-based systems allow for the quick introduction of a given compound, the potential physical damage caused by forcing liquid through blocked or embolized vasculature needs to be considered17. Although the foliar or drench application of compounds is less time-intensive to implement, direct plant injection reduces waste of the compound of interest due to losses to the air or soil and can also lengthen the time that compounds are in an active state by reducing the exposure to the outside environment18. Both these aspects are important for preserving expensive reagents and ensuring consistency among replicates in research settings.
This study describes the design, construction, and use of an innovative direct plant infusion (DPI) device, which can be used to assess how compounds of interest affect a host plant. A standard 3D printer was used to manufacture both the device itself and several components associated with its construction. This in-house construction method allows researchers to modify the device and device components based on their specific experimental needs and reduces the reliance on commercially available plant injection devices. The device setup is simple and efficient, and all the auxiliary components are readily available and inexpensive. Although the system was designed for use with a variety of plant species, the examples presented here pertain to potted citrus plants. Additionally, this study demonstrates that this device is capable of efficiently delivering multiple types of compounds systemically to young citrus plants without causing lethality. The compounds tested included CFDA, which was used to assess the compound distribution in the plant, and streptomycin and imidacloprid, which were used to verify that the antimicrobial and insecticidal effects of these compounds were observed when delivered via DPI.

Författare i fokus: Direkt infusionsanordning för molekylleverans i växter  

Direktinfusionsanordning för molekylleverans i växter

Delivering a therapeutic molecule to where a pathogen resides in a plant can be challenging. Treating Candidatus Liberibacter asiaticus, or CLas, a bacteria that is limited to the phloem, poses one of the biggest challenges in delivering molecules against it, especially when using conventional methods such as spray applications. The ease of printing the direct plant infusion device and its simple implementation makes it accessible to anyone.This enables the widespread screening of molecules for their potential therapeutic effect in trees. An added benefit is that the diffusion of molecules utilizes the natural process of plant transpiration. Our research improved efficiency and precision of molecular delivery in plants, which can be used for a range of research applications.By screening hundreds of molecules on small plants, we can eliminate the need for larger trees, leading to a reduction in cost and space utilization.

Watch this Scientific Journal Video about Attachment of the Direct Plant Infusion DPI Device to Citrus Plants for Compound Injection at JoVE.com

Attachment of the Direct Plant Infusion (DPI) Device to Citrus Plants for Compound Injection  

Fäste enheten för direkt växtinfusion (DPI) på citrusväxter för sammansatt injektion

Föröka små krukväxter citrusträd från rotade vegetativa sticklingar eller frön. Odla de krukväxta citruslinjerna vid 28 grader Celsius, behåll en ordentlig dagslängd. Välj citrusväxter av lämplig storlek för experimentet.För att fästa den 3D-tryckta Direct Plant Infusion- eller DPI-enheten på växtens stam, hitta en jämn, hälsosam plats på stammen utan stötar eller knutar. Använd en borr med två millimeter diameter och borra horisontellt ett hål genom mitten av stammen i en vinkel på 90 grader mot stamytan. Kör borrkronan genom hålet flera gånger för att skapa ett fritt, slätt hål.Skapa sedan en vertikal skiva i plastisolringen på motsatt sida av den sammansatta leveranskanalen. Montera ringen runt anläggningen och rikta in leveranskanalen mot det tidigare borrade hålet. Fäst DPI-enheten på plastisolringen, se till att de sitter tätt ihop och rikta in DPI-enhetens pip med plastisolringens leveranskanal och det borrade hålet.Vik den tätt med silikontejp för att hålla apparaten på plats. Kontrollera den helt monterade och anslutna enheten för att säkerställa korrekt inriktning och orientering. Spåret runt den vertikala kanalen möjliggjorde ytterligare sammansatt upptag genom barken.När den monterades korrekt spolades plastisolringen mot DPI-enheten och pipen var i linje med ett hål borrat i trädet.

Research

JoVE Journal

Biology

Attachment of the Direct Plant Infusion (DPI) Device to Citrus Plants for Compound Injection

Direct Infusion Device for Molecule Delivery in Plants

Testing the function of therapeutic compounds in plants is an important component of agricultural research. Foliar and soil-drench methods are routine but ...