1) As in previous years, ACP egg lays decreased substantially in cooler weather resulting in fewer embryos for injection, a fewer adults to maintain the colony. 2) During this quarter 524 Asian citrus psyllid (ACP) control uninjected eggs were set-up yielding a 36.3% nymphal hatch rate, resulting in an approx. increase of 11% above the previous quarter. 1,314 eggs were micro-injected with transformation vector/helper plasmids having a 4.2% hatch frequency, approx. 1% greater the previous quarter. Of the 55 newly hatched nymphs from injected eggs, 22 survived to adulthood and were backcrossed to wild (uninjected) ACP adults. Though none of the 63 G1 offspring exhibited DsRed fluorescence, a significant improvement in ACP viability post-injection from nymph to adulthood was achieved. Further improvement may be possible after completion of the environmental chamber retro-fit for temperature, humidity and lighting control. 3) To further improve embryo injections, nymphal hatching and survival to adulthood tests have been initiated to determine optimal heat shock temperature and duration. 722 eggs were collected and placed on tape, but not injected. After 18 hours 505 were heat shocked at 37 C for 45 min of which 20.2% hatched as nymphs, while 217 eggs that were not heat shocked (maintained at 25 C) had a hatch rate of 18.4% indicating that typical heat shock conditions did not have a negative effect on nymphal hatching.. 4) Reverse-transcriptase PCR (RT-PCR) was continued to test DsRed marker expression in ACP injected with vector plasmid (containing the IE1-DsRed marker) to determine if the IE1 baculovirus promoter functions normally in this species (as it does in caribfly, A. suspensa). For the A. suspensa controls and ACP samples, 400 embryos from each species were injected with pBac[IE1hr5-DsRed] vector plasmid and incubated for 24 hr before RNA extraction, and subsequent RT for cDNA production. cDNA from each sample and uninjected ACP and a water control were then subjected to PCR using DsRed primers which yielded positive DsRed expression in both A. suspensa and ACP, though approximately 2-fold higher in A. suspensa. This indicates IE1 promoter does function in ACP, though possibly less functional compared to A. suspensa. Tests will be continued to verify and quantify this activity, and similar tests will be performed to quantify piggyBac transposase expression in ACP.
Host plant resistance can be expected to provide the foundation for successful management of Asian citrus psyllid (ACP) in the future. We performed choice assays to assess antixenosis and no-choice assays to identify antibiosis effects of Poncirus trifoliata on host selection behavior of ACP to preferred and non-preferred hosts and studied how the chemical constituents of host plants affect oviposition and nymphal development. Some trifoliate accessions exhibited reduced larval and adult development (antibiosis) and reduced colonization (antixenosis). Experiments were performed to explore these aspects of decreased oviposition and nymphal development in trifoliates and the concurrent reduced feeding by ACP. Electrical penetration graph (EPG) studies were performed on resistant and susceptible accessions to characterize feeding from xylem and phloem and to provide insight into physical barriers in plant vascular elements that reduce phloem ingestion. EPG is used to characterize feeding behavior of piercing-sucking insects such as aphids, psyllids, whiteflies etc. While the insect probes, ingests, and salivates within the food source, characteristic voltage waveforms are produced that, in conjunction with histological studies, allow researchers to determine feeding patterns associated with acquisition and inoculation of pathogens. A primary limitation to this method is data analysis. Recordings of multiple insects for >3 hours generate gigabytes of data. Classification of data into insect feeding states representing ingestion, salivation, or other activities is typically accomplished through visual inspection of waveforms and manual annotation by comparison to published standards. Analysis is time consuming, requires expert training and manual annotation that precludes high-throughput analysis. We removed the data analysis bottleneck through application of machine learning algorithms designed to teach a computer to recognize and learn from minimal human coding. Machine learning algorithms can transform raw EPG data to feeding states with little or no human input. We used supervised and semi-supervised machine learning models to annotate ACP feeding waveforms and discovered previously unrecognized feeding states. With minimal (5%) human annotation of EPG recordings, machine learning models classified the remainder with >95% accuracy. A manuscript has been submitted for publication. Further analysis of feeding states should provide insight into the nature of pathogen transmission and allow identification of characteristics that render certain plant varieties more resistant to pathogen infection.
Volatile compounds often serve as cues for orientation by phytophagous insects to their host plants. However, all citrus volatiles studied to date in Y-tube olfactometer tests or field tests have elicited little to no orientation from the Asian citrus psyllid (ACP). We devised a probing choice assay to measure short range orientation and subsequent probing behavior by ACP on a wax substrate containing odorants. ACP probing increased with addition of a blend of formic and acetic acids compared with a control. A 2:1 blend of formic:acetic acids received the highest number of probes compared to single odorants including formic acid, acetic acid, ocimene and citral. Our results demonstrated that acetic and formic acids (and perhaps others compounds) act as phagostimulants for ACP. These novel results have been accepted for publication in the journal Chemical Senses and will be more fully described in the next report. We also studied the effect of incorporating additional compounds into multiple component blends to increase the stimulatory effect of acetic and formic acids on probing behavior of adult ACP. Compounds identified as stimulatory to olfactory receptors by single-cell recordings from neurons in ACP antennae were selected as candidate compounds. Experiments were performed using geometric mixture designs and response surface methods to identify primary drivers in 5- and 3-component mixtures to identify an optimal blend of odorants to maximize probing by ACP. We also studied the interaction between visual attraction to yellow and olfaction or gustation. Test compounds were incorporated into a slow-release wax matrix (SPLAT , ISCA Technologies Inc., Riverside, CA) and offered to caged ACP adults. Treatments (white or yellow SPLAT with or without odorants) were applied as 1 ml wide strips or beads to glass cover slips and arrayed on the floor of a cubical cage. Cohorts of ACP adults were starved for 6 h and released into each cage and allowed to move freely and probe on beads for 21 h. The beads were stained with Coomassie blue dye to visualize stylet sheaths produced by feeding attempts on the wax beads. There was a strong interaction between color and odorant. The yellow beads always received more probes compared with white beads. Addition of 1 (formic acid), 2 (formic and acetic acids) or 3 (formic acid, acetic acid and para-cymene) resulted in progressively more probing behavior on yellow beads, but not on white beads. Ethyl-butyrate and myrcene had no effect or a negative effect on probing behavior. The optimal 3-component blend will be reported as a phagostimulant blend in a manuscript under preparation. This blend will be further tested in greenhouse and field trials. This phagostimulant may be used in an attract-and-kill strategy for ACP control.
The large-scale validation of citrus leafminer (CLM) disruption with the ISCA DCEPT CLM technology came to an end for the 2015 season and we are analyzing and processing data currently. We are currently in the process of facilitating data analysis to determine how much better the technology worked on a larger scale as compared with previous seasons. We have continued to collaborate and support Dr. Stephen Lapointe’s laboratory for this research. This has required counting traps, measuring damage levels by CLM, and also performing advanced statistical analyses to determine efficacy of the products that we were are testing. One of the factors we have continued to investigate was dormant season applications of pheromones for management of CLM. After winter treatment, dispensers provided >85% disruption of male moth catch in traps for 37 weeks, and after spring treatment >92% for 26 weeks, but there was only a 12% reduction in leaf infestation in spring. Two applications were not better than only a single application in spring. Disruption of moth catch was weaker in treated plots where traps were placed high (3.1 m) rather than low (1.6 m) in the tree canopy. Dispensers provided effective and persistent disruption of male catch in pheromone-baited monitoring traps but were minimally effective in reducing leaf infestation by ACP. Winter application of pheromone did not reduce leaf mining in spring compared with spring application alone. Tops of trees may have provided a refuge for mating. Mining of leaves in treated plots may have derived from inadequate disruption of mating in the upper canopy of trees or influx of mated females from neighboring areas, or because of dissipation of pheromone near edges of treated areas. Border row application of pheromone requires further investigation, as well as, other application methods of the pheromone. The possibility of fly-in mated females requires significantly more investigation as well, as we have begun this research. Inadequate disruption in tops of trees may be fixed by placing pheromone dispensers at the very top of tree canopies as is for other lepidopteran species. We have also begun investigating this hypothesis. Dissipation of pheromone due to edge effects may be decreased by increasing size of the treated area even beyond our current efforts or by increasing density of dispensers near borders of treated areas. Insecticide applications near borders might also improve control of this species within pheromone-treated areas. Our results corroborate previous investigations and suggest that management of CLM with mating disruption on a small scale may be ineffectual, despite the fact that this was the largest trial ever realized for CLM. It appears that the size should be of ever larger scale. Our results point to the likely need for even larger-scale area wide treatments for effective mating disruption of ACP than we had previously expected. However, we were able to effectively reduce CLM damage with these treatments.
Although harvests are not complete for the Hamlin tests at all three of the sites, leaf drop and fruit drop counts are completed through different dates in December for each site. Leaf drop was not greatly different between the sites, nor were large differences found between the low concentration treatment and the control trees. Total leaf drop for Treated vs Control at each site was 370 vs 340; 243 vs 254, and 231 vs 233 total leaves for the fall count period. There was very little difference between 1 rated and 3 rated (decline) trees. Leaf drop was much heavier during September and early October than later. In one case almost 70 % of the leaf loss occurred by October 8th. This late summer, early fall leaf drop did not occur prior to HLB. Fruit drop was more uniform from late September until December. Per site drop rates for Treated vs Control trees was 19 vs 15.5, 18.8 vs 19.4 and 24.5 vs 25.1. There was usually only 1 or 2 % difference between 1 and 3 rated trees. Although fruit drop was reduced by the low concentration sprays of PGRs a year ago, there does not appear to be any effect on this year’s drop in the Hamlin cultivar. Valencia data is now being collected.
As previously reported, trees were steam-treated as described in our research proposal during July 2015. Adults and nymphs were enclosed in mesh sleeves on trees for acquisition feeding approximately 5 weeks following treatments. Following acquisition feeding, insect and leaf samples were collected (45 d post-treatment) from trees and taken to the lab for subsequent nucleic acid extraction and analysis. Acquisition feeding assays were repeated approximately two months later, with samples collections beginning 114 d post-treatment. Nymphs were collected from plants after adult emergence, until no psyllids remained in the mesh sleeves. The titer of Candidatus Liberibacter asiaticus (CLas) in trees receiving steam treatment did not significantly differ from untreated trees on days 0, 45, or 114 post-treatment (p = 0.99, 0.11, and 0.81, respectively; Tukey’s Honestly Significant Difference (HSD) test); however CLas titers in treated and untreated trees were lower at 45 d post treatment as compared to days 0 and 114 post-treatment. This is likely due to naturally-occurring seasonal decreases in CLas titers. CLas titers were significantly higher in steam-treated trees than untreated trees on day 0 as compared to day 45. CLas acquisition by adult psyllids enclosed on trees receiving thermal treatments did not differ significantly from acquisition by adult psyllids on untreated trees. Samples from CLas acquisition feeding assays with psyllid nymphs are still being processed. Based on these results, which indicated the thermal treatments applied during July 2015 did not reduce plant CLas titer or psyllid acquisition, a second thermal treatment was applied during late November 2015. In early January 2016, adults and nymphs were enclosed in mesh sleeves on trees for acquisition feeding approximately 5 weeks following treatments. Insect and leaf samples were collected after 10d of acquisition feeding or upon adult emergence to assess adult and nymph acquisition, respectively. In addition, we have initiated a complementary laboratory study to evaluate the effect of thermal therapy on acquisition of CLas under controlled conditions. Two year old Valencia trees were inoculated with CLas by enclosing plants with CLas-infected psyllids for two weeks. Currently, plants are being held in a secure, insect-free greenhouse until they are determined to be positive for CLas. At that time, a controlled environmental chamber will be used to apply heat treatments to trees for use in subsequent acquisition experiments
The goals of this project are to: 1. Continue monitoring ACP field populations for insecticide susceptibility in Florida. 2. Develop a useful tool to improve monitoring for resistance and to make such monitoring quick and easy. 3. Refine and implement effective rotation schedules based on understanding what is taking place in the field and our understanding of the fundamental resistance mechanisms in ACP. We are looking for development of an inexpensive, standardized diagnostic method to monitor insecticide resistance for all citrus grower against Asian citrus psyllid (ACP), Diaphorina citri, in Florida. This experiment used susceptible laboratory population of ACP was reared in a greenhouse at University of Florida, the Citrus Research and Education Center, Lake Alfred, FL. Tested insecticides were analytical grade and included bifenthrin (98%), dimethoate (99.5%), fenpropathrin (99.1%), imidacloprid (99.9%), cyantraniliprole (98.%), sulfoxaflor (99.5%), flupyradifurone (99.5%) and spinetoram (76.2% J-21.0% L) representing several insecticide classes. According to the manufacture recommendation, the insecticides were stored in freezer, room temperature and a cool dry place, respectively. About 0.01 or 0.1g of each chemical was weight and mixed with acetone (assay 99.5%). Various concentrations of the chemicals were prepared using the serial dilution procedure. Dilutions of technical insecticide in acetone were made to generate test concentrations ranging from 0.0001 to 10000 ng/ l. The bottle bioassay method was used to evaluate direct toxicity of insecticides against ACP adults to determine the LC50 and time to 100% knockdown rate. Each experiment was repeated three times and replicated five times. Toxicity of ACP was assessed 24 hours after treatment. Time to 100% knockdown rate was observed after 15, 30, 45, 60, 75, 90, 105 and 120 min. Results showed LC50 values of test insecticides ranged from 0.05 to 0.84 ng/ l. The 100% knockdown rate within the 1 hour period time was between 1000 to 10000 ng/ l with the exception of sulfoxaflor, cyantraniliprole and spinetoram. This study showed that a bottle bioassay is suitable for assaying insecticide resistance against adult ACP in the laboratory and should be useful to monitor insecticide resistance in the field. It should be a feasible and flexible tool for rapid testing of insecticide resistance.
The goals of this project are to: 1: Determine the fight initiation thresholds of ACP depending on temperature and humidity. 2: Determine the effect of wind speed on flight and the direction of psyllid flight with respect to wind. 3: Determine the effects of barometric pressure changes on psyllid dispersal. 4: Measure how psyllid dispersal is affected by abiotic factors in the field. 5: Establish a model to predict the risk of ACP dispersal/invasion based on prevailing abiotic conditions. Deliver this model as an online tool for growers. We are continuing experiments to study the effect of ambient temperature and relative humidity on the dispersal behavior of the Asian citrus psyllid (ACP). The experiments are set up in a climatic chamber where temperature and relative humidity are controlled precisely. Humidity and temperature are varied in a range that is in accordance with the conditions observed in Florida during spring and summer. The temperature treatments tested in the chamber were 15, 18, 21, 25, 30, 35 C and the humidity treatments 35%, and 75% and 95% RH. We obtained the highest percentage of dispersing individuals (67.80% after 3 days) at 30 C and observed slightly decreased dispersal between 33 C (63.00% after 3 days) indicating that the optimal temperature for short range movement in ACP is comprised between 30 and 33 C. Humidity did not affect the dispersal behavior of ACP except at low temperatures. Whereas 5.8% of ACP dispersed after 3 days at 18 C with 70% RH, 17.00% dispersed after 3 days at 18 C with 35% RH. The dispersion of ACP was the same at any humidity level after 21 C. In summary, our experiments indicate that the minimal threshold for psyllid movement is comprise between 15 and 18 C at 35% RH and is close to 18 C at 70% RH. We finished our automatized flight mill and are now capable to record the flight of four psyllids at the same time and for a longer time than previously. We will use this flight mill to investigate the effect of temperature and relative humidity on long-range flights. We also built a new wind tunnel adapted for Asian citrus psyllid; this wind tunnel has been controlled for regular air flow and absence of air disturbance. We will use this wind tunnel in the next weeks to determine how psyllids move depending of wind direction, and the maximum and minimal thresholds for wind speed regarding flight initiation. Finally we have developed a pressure chamber for psyllid behavior where we will be able to measure psyllid dispersion depending of controlled barometric pressure changes.
Currently, objective 1, investigating the specificity and efficacy of immune priming response in ACP, is underway. The objective of this experiment is to identify if ACP will survive a lethal dose of pathogenic bacteria, Serratia marcescens, if they have had prior immune challenge by a sublethal dose of the same or other species of bacteria. We are currently investigating the effect of: a) pathogen and b) non-pathogen to determine whether immune priming regulates the response of ACP to subsequent infections. Fifty experimental 3 day old ACP adults were collected and starved for 3 h. Half of the insects were orally primed by feeing on an artificial diet solution containing a sublethal dose priming treatment, corresponding to a lethal dose response of 1% (LD1) of the insect pathogen S. marcescens, while they other half were fed a control artificial diet solution containing bacterial media only (Tidbury et al. 2011) (Table I, Objective 1a; Fig 2A). The diet solution consist edof 15% (w:v) sucrose and 1/10 (v:v) green food coloring (McCormick & Co.) sandwiched between two parafilm layers, as described in Hall et al. (2010). After 24 hours, each treatment group was enclosed on separate branches of a potted citrus plant (var. ‘Swingle’). Seven days after priming, insects were removed from plants, starved for 3 h, and either injected (experiment 1) or orally inoculated (experiment 2) on a diet solution containing a lethal dose pathogen challenge or control treatment. After feeding or injection, the ACP were returned to the ‘Swingle’ plants for 14 days or 100% mortality. Mortality throughout the experiment was recorded every 24 hours. After 14 d, ACP were examined for the presence of S. marcescens by plating crushed insects on Petri dishes containing LB nutrient agar. Samples of colonies formed after 3 d were collected for DNA extraction and confirmation of bacterial identity using conventional PCR. Each experiment was replicated six times. We are currently analyzing the results of these experiments. Subsequent experiments to address the effects of non-pathogen immune priming to pathogenic bacteria (objective 1b) are still underway. Additionally, we have initiated the second project objective, to determine the specificity of RNAi immune priming in D. citri. We have begun construct dsRNA using a GFP sequence. GFP is not naturally present in ACP; therefore, introduction of the dsRNA for this target should activate the RNAi response without inducting psyllid mortality. Additional targets will be constructed during the current project quarter.
The objective of this project is first to identify a Bacillus thuringiensis (Bt) crystal toxin with basal toxicity against Asian citrus psyllid (ACP). The toxicity of the selected toxin will then be enhanced by addition of a peptide that binds to the gut of ACP. This peptide addition to the toxin is expected to enhance both binding and toxicity against ACP. Seven Bt strains showed toxicity against ACP with significant ACP mortality at 500ug/ml relative to control treatments. One Bt strain was selected for further identification of individual toxins by LC-MS/MS analysis. Based on the results, three specific Cry toxins were identified. The identity of these toxins was confirmed by comparison with the Bt Cry toxin holotype list, using ClustalW2-nucleotide. Further ACP bioassays with two of the toxins from the selected strain indicated that both toxins show toxicity against ACP at 500ug/ml. One of these toxins was selected for modification with the gut binding peptide. A phage disulfide-constrained heptapeptide library was screened and four ACP gut binding peptides were isolated. These peptides were expressed as peptide-mCherry fusion proteins as described in the previous report. Pull down assays were used to assess the binding of peptide-mCherry fusion proteins to ACP brush border membrane vesicles (BBMV) to confirm binding and to assess the relative strength of binding of the different peptides. mCherry alone and a random sequence peptide-mCherry fusion protein were used as negative controls for these assays. Specific binding of peptides to BBMV proteins was supported by the results of binding competition assays with mCherry or with a competitive synthetic peptide. Two dimensional gel electrophoresis coupled with ligand blot analysis showed that four peptide-mCherry fusion proteins bind to 50kDa, 37kDa and 25kDa BBMV proteins all with the same pI (~9). The negative control, mCherry showed non specific binding to an abundant 50kDa protein with a pI ~5. Analysis of peptide binding to the psyllid gut in vivo using fluorescence microscopy indicated that peptide 15-mCherry and peptide 18-mCherry bind the ACP gut, with minimal background fluorescence from the mCherry negative control.
Two citrus groves, one – 20 year-old Hamlin sweet orange trees predominately on Swingle rootstock and a second consisting of three year old Hamlin sweet orange trees on Swingle rootstock have received acid injection to selected blocks with and without sulfur applications for fifteen months. Irrigation water was acidified at one of four target water pH (7.5, 6.0, 5.0, and 4.0). A controlled release form of elemental sulfur was applied to half of the trees in each pH treatment (main effect) including the non-acidified control (pH~7.5). A controlled released form of elemental sulfur (Tiger 90) was allied at a rate of 500 pounds per treated acre to plots receiving either acidified irrigation water or control plots receiving irrigation water that was not acidified in June. Soil samples collected in August indicate that soil pH in the plots recieving sulfur applications remained near target pH level at both sites. However, plots not receiving sulfur applications increased in soil pH levels by an average of 0.75 pH units with plots to have pH between 6.0 and 7.0 returning near the pH of the irrigation water . This result is similar to data collected in 2014. The cause of soil pH is the low amount of irrigation required during the summer months. At both the mature and young tree site, no significant difference in root density was found in samples collected in August 2015. Significant increases in nutrient concentrations of leaves collected in June 2015 were been found in plots at both sites with reduced water and/or soil pH. The greatest increase in leaf nutrient concentrations were found for Mg, Mn, Zn, and B. These results may indicate increased nutrient uptake from soils with soil solutions below 6.5. Nutrient deficient symptoms consistent with HLB positive trees have lower nutrient concentration in their woody tissues and thus can not provide nutrients to leaves until these reserves are replenished by higher nutrient availability presumed at lower soil pH levels (5.5 to 6.5). Average Hamlin and Valencia tree water uptake under greenhouse conditions were not significantly different from one another. However, water uptake by trees affected with HLB were 20%-25% lower than healthy trees. These data have been consistent for the past year. There is increasing evidence of reduced water uptake for trees receiving water supplemented with calcium bicarbonate. The cause of reduced water uptake appears to be lower but non-significant reductions in root density and soil pH increases in soil irrigated with higher concentrations of calcium carbonate. Reduced water uptake by trees receiving calcium carbonate in irrigation water would account for reduced leaf area and trunk diameter.
Two citrus groves, one – 20 year-old Hamlin sweet orange trees predominately on Swingle rootstock and a second consisting of three year old Hamlin sweet orange trees on Swingle rootstock have received acid injection to selected blocks with and without sulfur applications for fifteen months. Irrigation water was acidified at one of four target water pH (7.5, 6.0, 5.0, and 4.0). A controlled released form of elemental sulfur (Tiger 90) was allied at a rate of 500 pounds per treated acre to plots receiving either acidified irrigation water or control plots receiving irrigation water that was not acidified in December. Significant increases in nutrient concentrations of leaves collected in November 2015 were been found in plots at both sites with reduced water and/or soil pH despite increases in soil pH during the summer months (reported in September 2015 report). The greatest increase in leaf nutrient concentrations were found for Mn, and Zn. These results may indicate increased nutrient uptake from soils with soil solutions below 6.5 and is presumed to be because of lower soil pH levels (5.5 to 6.5). Average water uptake by trees affected with HLB continue to be 20%-25% lower than healthy trees. Water uptake for trees receiving water supplemented with calcium bicarbonate was significantly reduced compared with health trees. Despite higher soil pH, tree root densities were not significantly different for HLB affected trees irrigated with water supplemented with calcium carbonate when compared with healthy trees irrigated without supplemental calcium carbonate. Tree size remain similar for all treatments.
The objective of this research project is to investigate and develop a potential non-phytotoxic, environmentally-friendly film-forming ACP repellent solution for preventing HLB infection. In the last reporting period, OS-SG 11, 12 and 13 were studied for plant safety and rainfastness and these formulations were delivered to our collaborators for ACP infection trials. In this reporting period, a new series of formulations (OS-SG 14-A,B,C,D) were developed using a new EPA approved polymer and environmentally friendly silica source. Optimization process involved different ratios of polymer to silica. The objective was to have the best combination for formulation stability and rainfastness. A clay based commercial control (Surround WP) was used for comparison purposes and this product is available to growers. Safety analysis of OS-SG 14 series was conducted using sour orange citrus as a model plant. Phytotoxicity studies were conducted using a Panasonic Environmental Test Chamber (Model MLR- 352H) to control light intensity, humidity and temperature cycling to simulate summer conditions (85% RH, 34oC). OS-SG 14 series did not cause any plant tissue damage at the applied rates, neither the Surround WP control. Temperature testing was conducted to determine if the presence of polymer posed a risk of increasing the heat build up on the plant surface. The OS-SG 14 series did not exhibit significant increases in temperature over commercial control. The composition and interactions between the components were confirmed using Fourier transform infrared spectroscopy (FT-IR), which confirmed the presence of silica from Si-O stretching, SiO-H stretching, Si-H bending and the polymer was detected from C=O stretching. The morphology of the new silica core used in OS-SG 14 series was observed using Scanning Electron Microscopy (SEM) and revealed large micron and sub-micron size irregular shaped particulates. The crystallinity of the silica core was studied using X-ray diffraction and it was found to be mostly amorphous with some crystalline structures. The commercial control material was amorphous in nature. The most optimized version from the OS-SG 14 series will undergo further studies including additional SEM, XRD and NMR characterizations. Film adherence (rainfastness) will be studied using AAS and atomic force microscopy (AFM). One or two best performing formulations will be selected from the from the OS-SG 14 series for future trials.
This project was initiated in 2014 and is focused on understanding the effect of nutrients applied through foliar fertilization programs (FFP) on HLB-affected trees in the Indian River marketing district. Two research trials have been established in commercial mature grapefruit groves in St. Lucie County and a young tree trial is being conducted at the UF-IRREC grove. Grove 1 has ~25 years old of Flame grapefruit on Swingle rootstock. Grove 2 utilizes ~7-year-old Ruby Red on Sour orange trees. Trial 3 is looking at the effect(s) that foliar fertilizers have on young tree growth and their ability to protract HLB disease symptoms in 2-year-old Ray Ruby grapefruit on Kuharske rootstock. Combinations of macro and micronutrient treatments initiated on all three trials in February 2014 and applications have been made quarterly since. The populations of the psyllids have been stable in both groves where the experiments are being conducted. The traps located in the different treatments are no showing significant differences among them, with an average between 0.03 and 0.06 individuals per trap per month. These values are low in comparison with the initial evaluations where the averages were between 1-3 individuals per trap. Fruit drop counts taken at about 3-week intervals beginning in July in all treatments for both groves showed no significant differences with the control treatment in terms of total amount of fruit drop or average fruit drop per tree. These results follow the same pattern observed during the evaluation in the 2014/15 season. The average of fruit drop was 53.2 and 26.2 fruit per tree Grove 1 and Grove 2, respectively. The grapefruit in Grove 2 was commercially harvested in mid-December. Four trees within each plot were individually harvested and all the fruit from each tree was run through the mobile Autoline sorting machine to determine fruit count and size distribution. Treatments 2B (application of micros, phosphite and DKP) and 4A (application of phosphite and KNO3) had significantly more larger fruit (27, 23 and greater than 23) in comparison with 1A (Control) and treatment 1-Ca ( control with soil application of CaNO3). In terms of Gross Packet Value (GPV), the treatment 2B showed a higher value per tree ($127.15) relative to the control treatment ($76.68). In general the GPV was superior for all treatments in 2015 than in 2014. The amount of boxes per tree was not statistically different among treatments, however the highest value was found for treatment 2B (average of 4.62 boxes per tree) and the lowest for the control treatment (average of 2.98 boxes per tree).
This project was initiated in 2014 and is focused on understanding the effect of nutrients applied through foliar fertilization programs (FFP) on HLB-affected trees in the Indian River marketing district. Two research trials have been established in commercial mature grapefruit groves in St. Lucie County and a young tree trial is being conducted at the UF-IRREC grove. Grove 1 has ~25 years old of Flame grapefruit on Swingle rootstock. Grove 2 utilizes ~7-year-old Ruby Red on Sour orange trees. Trial 3 is looking at the effect(s) that foliar fertilizers have on young tree growth and their ability to protract HLB disease symptoms in 2-year-old Ray Ruby grapefruit on Kuharske rootstock. Combinations of macro and micronutrient treatments initiated on all three trials in February 2014 and applications have been made quarterly since. Foliar nutrient applications were made according to schedule during the quarter. Materials are sprayed with a hand gun to all trees within plots, averaging about 2.3 gallons of solution per tree. Sticky traps are changed out every month and the number of psyllids counted. No differences between treatments have been observed. Fruit drop counts began in each grove in mid-July. During each count, the number of fruit on the ground under each measurement tree in each plot were counted. After counting, fruit were raked into the water furrow or bed middles to ensure an accurate count on the next measurement date. During the four measurement dates in the quarter, fruit drop in Grove 2 was minimal, ranging from 0 to 18 fruit per tree, averaging 4.1 fruit per tree. There were no differences among treatments. In Grove 1, drop was also minimal with an average of 3.0 fruit per tree. Drop ranged from 0 to 20 fruit per tree. No differences were found with respect to treatment. Leaf samples were taken in each grove for nutrient analysis in mid-August. In Grove 1, trees in the control plots and those not receiving phosphite had lower K than other treatments. Treatments not receiving the minor element sprays had lower leaf Mg, FE, Cu, Mn, Zn, and B than trees in treatments receiving the applications. In Grove 2, most leaf N and K levels overall were in the optimum range compared to mostly below optimum in Grove 1. With the exception of Zn and Mn, the minor element concentrations were similar among treatments. Leaf Mn and Zn concentrations were lower in control trees and the treatments not receiving the minor element sprays. Soil samples were taken in all plots in July. In grove 2, pH ranged from 7.2 to 7.6, averaging 7.4. Soil P ranged from 166 to 244 lb/ac, averaging 208 lb/ac. In Grove 1, soil pH ranged from 7.4 to 7.9, averaging 7.6. Soil P ranged from 148 lb/ac to 209 lb/ac, averaging 172 lb/ac. No differences in any soil parameter were evident with respect to treatment in either of the groves.
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