Towards realizing nano-enabled precision delivery in plants.

Nanocarriers (NCs) that can precisely deliver active agents, nutrients and genetic materials into plants will make crop agriculture more resilient to climate change and sustainable. As a research field, nano-agriculture is still developing, with significant scientific and societal barriers to overcome. In this Review, we argue that lessons can be learned from mammalian nanomedicine. In particular, it may be possible to enhance efficiency and efficacy by improving our understanding of how NC properties affect their interactions with plant surfaces and biomolecules, and their ability to carry and deliver cargo to specific locations. New tools are required to rapidly assess NC-plant interactions and to explore and verify the range of viable targeting approaches in plants. Elucidating these interactions can lead to the creation of computer-generated in silico models (digital twins) to predict the impact of different NC and plant properties, biological responses, and environmental conditions on the efficiency and efficacy of nanotechnology approaches. Finally, we highlight the need for nano-agriculture researchers and social scientists to converge in order to develop sustainable, safe and socially acceptable NCs.

Design and Development of a Polymeric-Based Curcumin Nanoparticle for Drug Delivery Enhancement and Potential Incorporation into Nerve Conduits.

Peripheral nerve injuries (PNI) impact millions of individuals in the United States, prompting thousands of nerve repair procedures annually. Nerve conduits (NC) are commonly utilized to treat nerve injuries under 3 cm but larger gaps still pose a challenge for successful peripheral nerve regeneration (PNR) and functional recovery. This is partly attributed to the absence of bioactive agents such as stem cells or growth factors in FDA-approved conduits due to safety, harvesting, and reproducibility concerns. Therefore, curcumin, a bioactive phytochemical, has emerged as a promising alternative bioactive agent due to its ability to enhance PNR and overcome said challenges. However, its hydrophobicity and rapid degradation in aqueous solutions are considerable limitations. In this work, a nanoscale delivery platform with tannic acid (TA) and polyvinylpyrrolidone (PVP) was developed to encapsulate curcumin for increased colloidal and chemical stability. The curcumin nanoparticles (CurNPs) demonstrate significantly improved stability in water, reduced degradation rates, and controlled release kinetics when compared to free curcumin. Further, cell studies show that the CurNP is biocompatible when introduced to neuronal cells (SH-SY5Y), rat Schwann cells (RSC-S16), and murine macrophages (J774 A.1) at 5 µM, 5 µM, and 10 µM of curcumin, respectively. As a result of these improved physicochemical properties, confocal fluorescence microscopy revealed superior delivery of curcumin into these cells when in the form of CurNPs compared to its free form. A hydrogen peroxide-based oxidative stress study also demonstrated the CurNP's potential to protect J774 A.1 cells against excessive oxidative stress. Overall, this study provides evidence for the suitability of CurNPs to be used as a bioactive agent in NC applications.

BacteSign: Building a Findable, Accessible, Interoperable, and Reusable (FAIR) Database for Universal Bacterial Identification.

With the increasing incidence of diverse global bacterial outbreaks, it is important to build an immutable decentralized database that can capture regional changes in bacterial resistance with time. Herein, we investigate the use of a rapid 3D printed µbiochamber with a laser-ablated interdigitated electrode developed for biofilm analysis of Pseudomonas aeruginosa, Acinetobacter baumannii and Bacillus subtilis using electrochemical biological impedance spectroscopy (EBIS) across a 48 h spectrum, along with novel ladder-based minimum inhibitory concentration (MIC) stencil tests against oxytetracycline, kanamycin, penicillin G and streptomycin. Furthermore, in this investigation, a search query database has been built demonstrating the deterministic nature of the bacterial strains with real and imaginary impedance, phase, and capacitance, showing increased bacterial specification selectivity in the 9772.37 Hz range.

Chitosan coated selenium: A versatile nano-delivery system for molecular cargoes.

The use of nanoscale delivery platforms holds tremendous potential to overcome the current limitations associated with the conventional delivery of genetic materials and hydrophobic compounds. Therefore, there is an imperative need to develop a suitable alternative nano-enabled delivery platform to overcome these limitations. This work reports the first one-step hydrothermal synthesis of chitosan functionalized selenium nanoparticles (Selenium-chitosan, SeNP) that are capable of serving as a versatile nanodelivery platform for different types of active ingredients. The chitosan functionalization modified the surface charge to allow the loading of active ingredients and improve biocompatibility. The effective loading of the SeNP was demonstrated using genetic material, a hydrophobic small molecule, and an antibiotic. Furthermore, the loading of active ingredients showed no detrimental effect on the specific properties (fluorescence and bactericidal) of the studied active ingredients. In vitro antimicrobial inhibitory studies exhibited good compatibility between the SeNP delivery platform and Penicillin G (Pen), resulting in a reduction of the minimum inhibitory concentration (MIC) from 32 to 16 ppm. Confocal microscopy images showed the uptake of the SeNP by a macrophage cell line (J774A.1), demonstrating trackability and intracellular delivery of an active ingredient. In summary, the present work demonstrates the potential of SeNP as a suitable delivery platform for biomedical and agricultural applications.

Novel magnesium-copper hybrid nanomaterials for management of bacterial spot of tomato.

Bacterial spot of tomato (BST), predominantly caused by Xanthomonas perforans (Xp) in Florida, is one of the most devastating diseases in hot, humid environments. Bacterial resistance to copper-based bactericides and antibiotics makes disease management extremely challenging. This necessitates alternative solutions to manage the disease. In this study, we used two novel hybrid copper and magnesium nanomaterials noted as magnesium double-coated (Mg-Db) and magnesium-copper (Mg-Cu), to manage BST. In in vitro experiments, no viable cells were recovered following 4 h exposure to 500 µg/ml of both Mg-Db and Mg-Cu, while 100 and 200 µg/ml required 24 h of exposure for complete inhibition. In viability assay using live/dead cell straining method and epifluorescence microscopy, copper tolerant Xp cells were killed within 4 h by both Mg-Cu and Mg-Db nanomaterials at 500 µg/ml, but not by copper hydroxide (Kocide 3000). In the greenhouse, Mg-Db and Mg-Cu at 100-500 µg/ml significantly reduced BST severity compared to micron-sized commercial Cu bactericide Kocide 3000 and the growers' standard (copper hydroxide + mancozeb) (P < 0.05). In field studies, Mg-Db and Mg-Cu nanomaterials significantly reduced disease severity in two out for field trials. Mg-Db at 500 µg/ml reduced BST severity by 34% compared to the non-treated control without affecting yield in Fall, 2020. The use of hybrid nanomaterials at the highest concentrations (500 µg/ml) used in the field experiments can reduce copper use by 90% compared to the growers' standard. In addition, there was no phytotoxicity observed with the use of hybrid nanomaterials in the field. These results suggest the potential of novel magnesium-copper based hybrid nanomaterials to manage copper-tolerant bacterial pathogens.

Multi-Functional Chitosan Nanovesicles Loaded with Bioactive Manganese for Potential Wound Healing Applications.

Chronic skin wound is a chronic illness that possesses a risk of infection and sepsis. In particular, infections associated with antibiotic-resistant bacterial strains are challenging to treat. To combat this challenge, a suitable alternative that is complementary to antibiotics is desired for wound healing. In this work, we report multi-functional nanoscale chitosan vesicles loaded with manganese (Chi-Mn) that has potential to serve as a new tool to augment traditional antibiotic treatment for skin wound healing. Chi-Mn showed antioxidant activity increase over time as well as antimicrobial activity against E. coli and P. aeruginosa PA01. The modified motility assay that mimicked a skin wound before bacterial colonization showed inhibition of bacterial growth with Chi-Mn treatment at a low area density of 0.04 µg of Mn per cm2. Furthermore, this study demonstrated the compatibility of Chi-Mn with a commercial antibiotic showing no loss of antimicrobial potency. In vitro cytotoxicity of Chi-Mn was assessed with macrophages and dermal cell lines (J774A.1 and HDF) elucidating biocompatibility at a wide range (2 ppm-256 ppm). A scratch wound assay involving human dermal fibroblast (HDF) cells was performed to assess any negative effect of Chi-Mn on cell migration. Confocal microscopy study confirmed that Chi-Mn tested at the MIC (16 ppm Mn) has no effect on cell migration with respect to control. Overall, this study demonstrated the potential of Chi-Mn nanovesicles for wound healing applications.

Carbon nanomaterials are a superior soil amendment for sandy soils than biochar based on impacts on lettuce growth, physiology and soil biochemical quality.

A significant bottleneck of current agricultural systems remains the very low agronomic efficiency of conventional agrochemicals, particularly in sandy soils. Carbon nanomaterials (CNMs) have been proposed to address this inefficiency in sandy soils, which could potentially improve soil fertility and enhance crop growth and physiological processes. However, the effects of different rates of CNMs on crop physiological and soil biochemical quality in sandy soils must be compared to other carbon sources (e.g., biochar) before CNMs can be broadly used. To address this, a 70-day pot experiment was set up, growing lettuce under ten treatments: a negative control with no CNMs, biochar or fertilizer; a fertilizer-only control; three CNMs-only unfertilized treatments (CNMs at 200, 400 and 800 mg kg-1 soil); two biochar treatments with fertilizer (biochar at 0.5% and 1% by soil mass + fertilizer); and three CNMs treatments with fertilizer (CNMs at 200, 400 and 800 mg kg-1 soil + fertilizer). A novel amorphous, water-dispersible, and carboxyl-functionalized CNMs with pH of 5.5, zeta potential of -40.6 mV and primary particle diameter of 30-60 nm was used for this experiment. Compared to the fertilizer-only control, CNMs applied at low to medium levels (200-400 mg kg-1) significantly increased lettuce shoot biomass (20-21%), total chlorophyll (23-27%), and fluorescence and photosynthetic activities (4-10%), which was associated with greater soil nutrient availability (N: 24-58%, K: 68-111%) and higher leaf tissue accumulation (N: 25-27%; K: 66%). Low to medium levels of CNMs also significantly increased soil biochemical properties, such as higher soil microbial biomass carbon (27-29%) and urease enzyme activity (34-44%) relative to fertilizer-only applications. In contrast, biochar (0.5%) increased lettuce biomass relative to fertilizer-only but had no significant effect on soil fertility and biological properties. These results suggest that CNMs at low to medium application rates are a superior carbon-based amendment relative to biochar in sandy soils.

Improved Persistence of Bacteriophage Formulation with Nano N-Acetylcysteine-Zinc Sulfide and Tomato Bacterial Spot Disease Control.

Bacteriophages are biocontrol agents used to manage bacterial diseases. They have long been used against plant pathogenic bacteria; however, several factors impede their use as a reliable disease management strategy. Short-lived persistence on plant surfaces under field conditions results mainly from rapid degradation by exposure to ultraviolet (UV) light. Currently, there are no effective commercial formulations that protect phages from UV. The phage ΦXp06-02-1, which lyses strains of the tomato bacterial spot pathogen Xanthomonas perforans, was mixed with different concentrations of the nanomaterial N-acetylcysteine surface-coated manganese-doped zinc sulfide (NAC-ZnS; 3.5 nm). In vitro, NAC-ZnS at 10,000 µg/ml formulated phage, when exposed to UV for 1 min, provided statistically equivalent plaque-forming unit (PFU) recovery as phages that were not exposed to UV. NAC-ZnS had no negative effect on the phage's ability to lyse bacterial cells under in vitro conditions. NAC-ZnS reduced phage degradation over time in comparison with the nontreated control, whereas N-acetylcysteine-zinc oxide (NAC-ZnO) had no effect. In fluorescent light, without UV exposure, NAC-ZnO-formulated phages were more infective than NAC-ZnS-formulated phages. The nanomaterial-phage mixture did not cause any phytotoxicity when applied to tomato plants. Following exposure to sunlight, the NAC-ZnS formulation improved phage persistence in the phyllosphere by 15 times compared with nonformulated phages. NAC-ZnO-formulated phage populations were undetectable within 32 h, whereas NAC-ZnS-formulated phage populations were detected at 103 PFU/g. At 4 h of sunlight exposure, NAC-ZnS-formulated phages at 1,000 µg/ml significantly reduced tomato bacterial spot disease severity by 16.4% compared with nonformulated phages. These results suggest that NAC-ZnS can be used to improve the efficacy of phages for bacterial diseases.

Potential of Novel Magnesium Nanomaterials to Manage Bacterial Spot Disease of Tomato in Greenhouse and Field Conditions.

Bacterial spot of tomato is among the most economically relevant diseases affecting tomato plants globally. In previous studies, non-formulated magnesium oxide nanoparticles (nano-MgOs) significantly reduced the disease severity in greenhouse and field conditions. However, the aggregation of nano-MgO in liquid suspension makes it challenging to use in field applications. Therefore, we formulated two novel MgO nanomaterials (SgMg #3 and SgMg #2.5) and one MgOH2 nanomaterial (SgMc) and evaluated their physical characteristics, antibacterial properties, and disease reduction abilities. Among the three Mg nanomaterials, SgMc showed the highest efficacy against copper-tolerant strains of Xanthomonas perforans in vitro, and provided disease reduction in the greenhouse experiments compared with commercial Cu bactericide and an untreated control. However, SgMc was not consistently effective in field conditions. To determine the cause of its inconsistent efficacy in different environments, we monitored particle size, zeta potential, morphology, and crystallinity for all three formulated materials and nano-MgOs. The MgO particle size was determined by the scanning electron microscopy (SEM) and dynamic light scattering (DLS) techniques. An X-ray diffraction (XRD) study confirmed a change in the crystallinity of MgO from a periclase to an Mg(OH)2 brucite crystal structure. As a result, the bactericidal activity correlated with the high crystallinity present in nano-MgOs and SgMc, while the inconsistent antimicrobial potency of SgMg #3 and SgMg #2.5 might have been related to loss of crystallinity. Future studies are needed to determine which specific variables impair the performance of these nanomaterials in the field compared to under greenhouse conditions. Although SgMc did not lead to significant disease severity reduction in the field, it still has the potential to act as an alternative to Cu against bacterial spot disease in tomato transplant production.

Zinc Oxide-Based Nanoformulation Zinkicide Mitigates the Xylem-Limited Pathogen Xylella fastidiosa in Tobacco and Southern Highbush Blueberry.

The xylem-limited pathogen Xylella fastidiosa causes severe economic losses worldwide, and no effective antimicrobial disease management options are available. The goal of this study was to evaluate the efficacy of a novel ZnO-based nanoparticle formulation, Zinkicide TMN110 (ZnK), against X. fastidiosa in vitro and in planta. In vitro, minimum bactericidal concentration (MBC) of ZnK analyzed in Pierce's Disease 2 medium was estimated at approximately 60 ppm. Time-kill kinetics assay showed a 100% reduction of culturable X. fastidiosa in less than 1 h after ZnK treatment. Microfluidic chambers assays showed that ZnK also inhibits X. fastidiosa cell aggregation and growth under flow conditions. Phytotoxicity assessments in the greenhouse demonstrated that ZnK can be applied as a soil drench in 50 ml at 500 ppm/plant/week up to four times to tobacco and blueberry without causing visible damage. ZnK was also evaluated for disease control in the greenhouse using tobacco infected with X. fastidiosa subsp. fastidiosa strain TemeculaL. ZnK soil drench weekly applications at concentrations of 500 followed by 1,000 ppm (500/1,000) and 500/500/1,000 ppm (in 50 ml each), reduced X. fastidiosa populations by >2 to 3 log10 units and disease severity by approximately 57 and 76%, respectively, compared with the untreated control. Similarly, when blueberry plants infected with X. fastidiosa subsp. multiplex strain AlmaEm3 were soil drenched with ZnK at concentrations 1,000/1,000 ppm and 1,000/1,000/500 ppm (in 200 ml each), the bacterial population was reduced by approximately 1 to 2 log10 units, and disease severity decreased by approximately 39 and 43%, respectively. Overall, this study shows antibacterial activity of ZnK against X. fastidiosa and its effectiveness in plants to reduce disease symptoms under controlled conditions.

Plotter Cut Stencil Masks for the Deposition of Organic and Inorganic Materials and a New Rapid, Cost Effective Technique for Antimicrobial Evaluations.

Plotter cutters in stencil mask prototyping are underutilized but have several advantages over traditional MEMS techniques. In this paper we investigate the use of a conventional plotter cutter as a highly effective benchtop tool for the rapid prototyping of stencil masks in the sub-250 µm range and characterize patterned layers of organic/inorganic materials. Furthermore, we show a new diagnostic monitoring application for use in healthcare, and a potential replacement of the Standard Kirby-Bauer Diffusion Antibiotic Resistance tests was developed and tested on both Escherichia coli and Xanthomonas alfalfae as pathogens with Oxytetracycline, Streptomycin and Kanamycin. We show that the reduction in area required for the minimum inhibitory concentration tests; allow for three times the number of tests to be performed within the same nutrient agar Petri dish, demonstrated both theoretically and experimentally resulting in correlations of R ≈ 0.96 and 0.985, respectively for both pathogens.

Huanglongbing Pandemic: Current Challenges and Emerging Management Strategies.

Huanglongbing (HLB, aka citrus greening), one of the most devastating diseases of citrus, has wreaked havoc on the global citrus industry in recent decades. The culprit behind such a gloomy scenario is the phloem-limited bacteria "Candidatus Liberibacter asiaticus" (CLas), which are transmitted via psyllid. To date, there are no effective long-termcommercialized control measures for HLB, making it increasingly difficult to prevent the disease spread. To combat HLB effectively, introduction of multipronged management strategies towards controlling CLas population within the phloem system is deemed necessary. This article presents a comprehensive review of up-to-date scientific information about HLB, including currently available management practices and unprecedented challenges associated with the disease control. Additionally, a triangular disease management approach has been introduced targeting pathogen, host, and vector. Pathogen-targeting approaches include (i) inhibition of important proteins of CLas, (ii) use of the most efficient antimicrobial or immunity-inducing compounds to suppress the growth of CLas, and (iii) use of tools to suppress or kill the CLas. Approaches for targeting the host include (i) improvement of the host immune system, (ii) effective use of transgenic variety to build the host's resistance against CLas, and (iii) induction of systemic acquired resistance. Strategies for targeting the vector include (i) chemical and biological control and (ii) eradication of HLB-affected trees. Finally, a hypothetical model for integrated disease management has been discussed to mitigate the HLB pandemic.

Multifunctional Surface, Subsurface, and Systemic Therapeutic (MS3T) Formulation for the Control of Citrus Canker.

A multifunctional surface, subsurface and systemic therapeutic (MS3T) formulation comprised of two bactericides, both didecyldimethylammonium chloride (DDAC) and a zinc (Zn)-chelate, was developed as an alternative to copper pesticides for crop protection. Agricultural grade chemicals were used to prepare MS3T formulations. Minimal inhibitory concentration (MIC) was determined to be tested in vitro against Xanthomonas alfalfae subsp. citrumelonis (herein called Xa), Escherichia coli (E. coli), and Pseudomonas syringae (Ps). Assessment of the phytotoxic potential was carried out on tomato under greenhouse conditions. Moreover, field trials were conducted during three consecutive years on grapefruit (Chrysopelea paradise) groves to evaluate efficacy against citrus canker (Xanthomonas citri subsp. citri), scab (Elsinoe fawcetti), and melanose (Diaporthe citri). In addition to disease control, improvements to both fruit yield and quality were observed likely due to the nutritional activity of MS3T via the sustained release of plant nutrients (Zn and nitrogen). Zn residues of leaf tissues were analyzed via atomic absorption spectroscopy (AAS) at various time points before and after MS3T foliar applications throughout the duration of the 2018 field trial. Field trial results demonstrated MS3T to be an effective alternative to copper (Cu)-based formulations for the control of citrus canker.

Tracking of fluorescent antibiotic conjugate in planta utilizing fluorescence lifetime imaging.

MAIN CONCLUSION: Excited state lifetime-based separation of fluorophore-tagged antibiotic conjugate emission from the spectrally broad plant autofluorescence enables in planta tracking of the translocation of systemic cargo such as antibiotics via fluorescence lifetime imaging. The efficacy of antibiotic treatments in citrus crops is uncertain due to mixed results from in-field experiments and a lack of study on their systemic movement. As of yet there has been an inability to track treatments using traditional fluorescence microscopy due to treatments having little fluorescence characteristics, and signal convolution due to plant autofluorescence. In this study, we used streptomycin sulfate, a commercially available antibiotic, and conjugated it to a modified tris(bipyridine) ruthenium (II) chloride, a dye with an excited state lifetime magnitudes higher than other commonly used organic fluorescent probes. The resultant is a fluorescence lifetime imaging (FLIM) trackable antibiotic conjugate, covalently attached via an amide linkage that is uniquely distinguishable from plant autofluorescence. Characterization of the fluorescent antibiotic conjugate showed no mitigation of excited state lifetime, and a distinct IR peak not found in any synthetic components. Subsequent tracking using FLIM in citrus tissue was achieved, with identification of movement through citrus plant vasculature via tissue localization in xylem and phloem. Results indicated upwards systemic movement of the conjugate in both xylem and phloem after 48 h of incubation. However, the conjugate failed to move down towards the root system of the plant by 168 h. Mechanistically, it is likely that xylem contributes heavily in the translocation of the conjugate upwards; however, phloem led flow due to growth changes could act as a contributor. This proof-of-concept sets groundwork for subsequent studies regarding antibiotic localization and movement in citrus.

Copper-fixed quat: a hybrid nanoparticle for application as a locally systemic pesticide (LSP) to manage bacterial spot disease of tomato.

The development of bacterial tolerance against pesticides poses a serious threat to the sustainability of food production. Widespread use of copper (Cu)-based products for plant disease management has led to the emergence of copper-tolerant pathogens such as Xanthomonas perforans (X. perforans) strains in Florida, which is very destructive to the tomato (Solanum lycopersicum) industry. In this study, we report a hybrid nanoparticle (NP)-based system, coined Locally Systemic Pesticide (LSP), which has been designed for improved efficacy compared to conventional Cu-based bactericides against Cu-tolerant X. perforans. The silica core-shell structure of LSP particles makes it possible to host ultra-small Cu NPs (<10 nm) and quaternary ammonium (Quat) molecules on the shell. The morphology, release of Cu and Quat, and subsequent in vitro antimicrobial properties were characterized for LSP NPs with core diameters from 50 to 600 nm. A concentration of 4 µg mL-1 (Cu): 1 µg mL-1 (Quat) was found to be sufficient to inhibit the growth of Cu-tolerant X. perforans compared to 100 µg mL-1 (metallic Cu) required with standard Kocide 3000. Wetting properties of LSP exhibited contact angles below 60°, which constitutes a significant improvement from the 90° and 85° observed with water and Kocide 3000, respectively. The design was also found to provide slow Cu release to the leaves upon water washes, and to mitigate the phytotoxicity of water-soluble Cu and Quat agents. With Cu and Quat bound to the LSP silica core-shell structure, no sign of phytotoxicity was observed even at 1000 µg mL-1 (Cu). In greenhouse and field experiments, LSP formulations significantly reduced the severity of bacterial spot disease compared to the water control. Overall, the study highlights the potential of using LSP particles as a candidate for managing tomato bacterial spot disease and beyond.

Zinkicide Is a ZnO-Based Nanoformulation with Bactericidal Activity against Liberibacter crescens in Batch Cultures and in Microfluidic Chambers Simulating Plant Vascular Systems.

Phloem-limited bacterial "Candidatus Liberibacter" species are associated with incurable plant diseases worldwide. Antimicrobial treatments for these pathogens are challenging due to the difficulty of reaching the vascular tissue they occupy at bactericidal concentrations. Here, in vitro antimicrobial mechanisms of Zinkicide TMN110 (ZnK), a nonphytotoxic zinc oxide (ZnO)-based nanoformulation, were compared to those of bulk ZnO (b-ZnO) using as a model the only culturable species of the genus, Liberibacter crescens Minimum bactericidal concentration (MBC) determination and time-kill assays showed that ZnK has a bactericidal effect against L. crescens, whereas b-ZnO is bacteriostatic. When ZnK was used at the MBC (150 ppm), its antimicrobial mechanisms included an increase in Zn solubility, generation of intracellular reactive oxygen species, lipid peroxidation, and cell membrane disruption; all of these were of greater intensity than those of b-ZnO. Inhibition of biofilms, which are important during insect vector colonization, was stronger by ZnK than by b-ZnO at concentrations between 2.5 and 10 ppm in batch cultures; however, neither ZnK nor b-ZnO removed L. crescens preformed biofilms when applied between 100 and 400 ppm. In microfluidic chambers simulating source-to-sink phloem movement, ZnK significantly outperformed b-ZnO in Zn mobilization and bactericidal activity against L. crescens planktonic cells in sink reservoirs. In microfluidic chamber assays assessing antibiofilm activity, ZnK displayed a significantly enhanced bactericidal activity against L. crescens individual attached cells as well as preformed biofilms compared to that of b-ZnO. The superior mobility and antimicrobial activity of ZnK in microenvironments make this formulation a promising product to control plant diseases caused by "Candidatus Liberibacter" species and other plant vascular pathogens.IMPORTANCE "Candidatus Liberibacter" species are associated with incurable plant diseases that have caused billions of dollars of losses for United States and world agriculture. Chemical control of these pathogens is complicated, because their life cycle combines intracellular vascular stages in plant hosts with transmission by highly mobile insect vectors. To date, "Candidatus Liberibacter" species are mostly unculturable, except for Liberibacter crescens, a member of the genus that has been used as a model for in vitro assays. Here, we evaluated the potential of Zinkicide (ZnK) as an antimicrobial against "Candidatus Liberibacter" species in batch cultures and under flow conditions, using L. crescens as a biological model. ZnK displayed bactericidal activity against L. crescens in batch cultures and showed increased mobility and bactericidal activity in microfluidic devices resembling "Candidatus Liberibacter" species natural habitats. ZnK performance observed here against L. crescens makes this compound a promising candidate to control plant diseases caused by vascular pathogens.

SDS-PAGE for Monitoring the Dissolution of Zinc Oxide Bactericidal Nanoparticles (Zinkicide) in Aqueous Solutions.

Zinkicide is a systemic bactericidal formulation containing protein-size fluorescent zinc oxide-based nanoparticles (nano-ZnO). Previous studies have shown that Zinkicide is effective in controlling citrus diseases. Its field performance as an antimicrobial agent has been linked to the bioavailability of zinc ions (Zn2+) at the target site. It is therefore important to monitor Zn2+ release from Zinkicide so that application rates and frequency can be estimated. In this study, we present a simplistic approach designed to monitor Zinkicide nanoparticle dissolution rates in water and acidic buffer solutions using traditional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The evolution of nano-ZnO in the polyacrylamide gel scaffolds was studied by exciting the sample with UV light and detecting the fluorescence of nano-ZnO. Fluorescence intensities measured with this assay allowed for quantitative analysis of molecular weight changes of nano-ZnO in citrate buffer, a surrogate of citrus juice. Our results demonstrated that citrate buffer induced the greatest degradation of Zinkicide. Fluorescence intensity fluctuations were observed over time, indicating interactions of citrate with the surface of nano-ZnO. These findings provide a new approach to quantify the dissolution of nanoparticles in simulated environments, even when other analytical methods lack sensitivity because of the small size of the system (≈4 nm).

Advances in Translational Nanotechnology: Challenges and Opportunities.

The burgeoning field of nanotechnology aims to create and deploy nanoscale structures, devices, and systems with novel, size-dependent properties and functions. The nanotechnology revolution has sparked radically new technologies and strategies across all scientific disciplines, with nanotechnology now applied to virtually every area of research and development in the US and globally. NanoFlorida was founded to create a forum for scientific exchange, promote networking among nanoscientists, encourage collaborative research efforts across institutions, forge strong industry-academia partnerships in nanoscience, and showcase the contributions of students and trainees in nanotechnology fields. The 2019 NanoFlorida International Conference expanded this vision to emphasize national and international participation, with a focus on advances made in translating nanotechnology. This review highlights notable research in the areas of engineering especially in optics, photonics and plasmonics and electronics; biomedical devices, nano-biotechnology, nanotherapeutics including both experimental nanotherapies and nanovaccines; nano-diagnostics and -theranostics; nano-enabled drug discovery platforms; tissue engineering, bioprinting, and environmental nanotechnology, as well as challenges and directions for future research.

Control of Citrus Canker in Greenhouse and Field with a Zinc, Urea, and Peroxide Ternary Solution.

Accumulation of toxic copper in soil and development of copper-resistant pests are emerging challenges currently faced by the agricultural community worldwide. As an alternative, we have developed a ternary zinc chelate solution (TSOL) pesticide where zinc ions are the primary active ingredient. The material is composed of zinc, urea, and hydrogen peroxide. Urea was chosen as it is widely used as a plant fertilizer and can also bind to both zinc and hydrogen peroxide. No phytotoxicity was observed with TSOL on Meyer lemon (Citrus × meyeri) seedlings at a field spray rate of 800 µg/mL Zn metal concentration. Antimicrobial studies showed that TSOL exhibited improved killing efficacy against Escherichia coli and Xanthomonas alfalfae compared to Zn ions alone. Citrus canker field trials in a grapefruit (Chrysopelea paradisi) grove over three years showed that TSOL provided comparable disease protection to copper products at an equivalent or lower metal content.

Precision Vascular Delivery of Agrochemicals with Micromilled Microneedles (µMMNs).

We demonstrate use of makerspace techniques involving subtractive microtechnologies to fabricate micromilled microneedles (µMMNs) of stainless steel (SS) for precise delivery of agrochemicals into vascular bundles of plant tissue. Precision delivery is of immense importance for systemic pathogen control in specific areas of plant tissue. Optimization of the micromilling allows for selective removal of SS at the microscale and the microfabrication of a 5 × 5 array of µMMNs having both base width and height of 500 µm to enable precise puncture into the stem of citrus saplings. Atomic Absorption Spectroscopy reveals up to 7.5× increase in the uptake of a therapeutic cargo while Scanning Electron Microscopy reveals that specific sites of the vascular bundle; either xylem or the phloem can be uniquely targeted with customized µMMNs. Such rapid and cost-effective customization with intricate designs along with scalability is enabled by makerspace microfabrication. Additionally, a 19 × 20 array of micromilled mesoneedles has been fabricated and affixed to a paint roller as an applicator system for real-world field testing outside the laboratory. Initial results indicate reliable behavior of the applicator system and the technique can be applied to the systemic delivery of agrochemicals while conserving the loss of the agrochemical with increased application efficiency.