This project, in general, seeks to determine if there are any observable influences of foliar fertilizers on HLB-affected grapefruit in the Indian River marketing district. To-date, this project has established 3 separate research trials in commercial grapefruit groves and university research farms in St. Lucie county, Florida. Trial #1 is ~25 years old of ‘Flame’ grapefruit on Swingle rootstock. Trial #2 utilizes ~6 year old ‘Ruby Red’ on Sour orange trees. The first 2 trials are located in commercial groves. Trial #3 seeks to determine the effect(s) that foliar fertilizers have on young tree growth and their ability to protract HLB disease symptoms in 1 year old ‘Ray Ruby’ grapefruit on Kuharske rootstock. For trial #1, there are a total of 30 replicated experimental units; for trial #2, there are a total of 50 replicated experimental units; and, for the trial #3, there are a total of 24 replicated experimental units. Foliar fertilizer applications have been started since February 2014. Tree growth data, leaf nutrition data, and CLas titer measurements have been made. Fruit drop monitoring will begin in September 2014 in trials #1 and #2. In prosecution of these efforts, 2-part-time OPS employees have been hired to do our foliar sprays and collect data observations. As of January 2015, the first year of data collection on the commercial grove plots (trials #2 and #3) as well as the young tree trial at the IRREC (trial #1). First-year fruit yields were gathered in the commercial trials (trials #2 and #3) in December 2014. In addition, all of the foliar fertilizer applications were made (once per quarter) to all trials. Currently, data are being analyzed from the first growing season (2014).
The canker field trial of Zinkicide compared to industry standard management on grapefruit showed better control of canker on fruit than all other treatments. We also observed control of some fungal pathogens. A second year of this field trial used a half and quarter rate of Zinkicide compared to the previous year. The half rate provided equivalent canker control to copper, while the quarter rate started to show reduction in canker control. Analysis of the data from the nanoparticle and non-nanoparticle version of Zinkicide gives insight into possible systemic movement. Dr. Santra’s group has improved synthesis of Zinkicide to reduce the amount of some ingredients needed for synthesis while maintaining efficacy. This improvement is aimed at reducing cost of production. The research team continues working on commercial production and registration for canker although the time line for registration is still too preliminary to accurately predict.
This research seeks to determine whether young trees infected with CLas and displaying typical HLB symptoms can be brought to maturity and produce an economically viable yield. This will be achieved by managing a 58 acre grove of 3-year-old ‘Valencia’ / Kuharske Carrizo trees using a combination of three different foliar and three different ground applied nutritional programs. Factorial AxB treatments consist of A) ground-applied: 1) Liquid/dry+Ca (BHG standard), 2) Liquid+Ca, 3) Liquid/dry-Ca B) foliar-applied: 1) BHG standard-Ca, 2) BHG standard+Ca, 3) “Prescription”(+Ca). The prescription treatment was designed to be dynamic, customized for optimization, with feedback based on frequent leaf tissue analyses, visual symptoms, and the growth of the tree canopies and yield. There are six replications of treatments, with two being pure replications. The grove continues to grow well and also look normal, despite the nearly 100% HLB incidence. Foliation of canopies is dense, and leaf color in the summer and fall was a healthy green. There are no new measurements to report during this quarter. The next reported update will include the fruit yield and quality measurements of the 2014/15 season.
This is a project to develop novel approaches to controlling psyllids . Effective techniques to reduce the rate of Huanglongbing (HLB) spread are key to slowing its incidence, especially for new citrus plantings. RNA-interference (RNAi) is a natural regulatory and anti-viral response in eukaryotes and can be manipulated to target mRNAs/gene expression, including to control insects. Our on-going collaboration has found that RNAi inducers, expressed in citrus trees using the Citrus tristeza virus (CTV) vector, reduce the survival of adult Diaphorina citri moving onto the trees, and greatly reduce their reproduction and acquisition of Candidatus Liberibacter asiaticus by progeny. Our goal is to further improve RNAi activity such that it will help to manage D. citri and HLB, allow reduction in pesticide use and lower grower costs for U.S. citrus. Sequences of specific psyllid genes that are thought to be needed for the survival and reproduction of psyllids are cloned into the CTV vector. As the virus replicates in phloem cells, it produces large amounts of dsRNA intermediates that now also produces dsRNAs containing psyllid sequences. The normal plants RNAi defense mechanism processes the dsRNAs into small 21 nt siRNAs that target mRNA degradation. These siRNAs migrate from virus infected cells into the sieve element. As the psyllid feeds, it sucks up these siRNAs that now target the psyllid mRNAs and prevents the psyllid from making this protein. The lack of this protein has detrimental effects on the survival, reproduction, and CLas acquisition of psyllids. So far, we have seen reduction in survival of adult psyllids placed on RNAi expressing plants, but the effects on reproduction of the new generation of psyllids has been much greater. This is likely because the nymphs are rapidly growing an need lots of new protein synthesis and because they uptake large amounts of phloem sap. Although we have found sequences that can prevent normal expression of specific genes in psyllids, we are looking at several genes to identify sequences that work best. We recently have found that the RNAi target sequences that are expressed from near the 3′ terminus of the CTV vector appear to not be stable enough to be useful for controlling HLB spread in the field. We are recloning all of the target sequences in other positions within the CTV genome looking for an optimal compromise between efficacy and stability.
During this period we continued efforts for artificial micro RNA (amiRNA) delivery and efficacy assessment towards B. cockerelli, but also performed a series of new experiments targeting D. citri via artificial diet, dsRNA feeding. We chose 7 D. citri targets based on our analysis of which target RNAs may be susceptible to RNAi activity, and which could give detectable phenotypic effects. Our intent was to use dsRNAs to help identify mRNAs that we could later target by amiRNAs. We took two approaches. First we generated dsRNAs in vitro, and second, we purchased some in order to perform larger scale experiments. Like in previous studies by ourselves and others, we found that artificial diet feeding of dsRNAs to adult D. citri can induce mortality. All seven tested by us (AI, ATA, ATD, CD, CL, CS and TR) induced effects significantly greater than did the GFP dsRNA controls used at the same concentrations and over the same testing periods. The CS, CD and AI dsRNAs gave slightly greater effects in some experiments. Therefore, we next assessed quantitative effects of these dsRNAs on their specific mRNA targets by using RT-qPCR. All three gave significant target mRNA knockdown, even at very low concentrations used by us. Our efforts now will proceed to design amiRNAs for these targets by using bioinformatics approaches, but also NGS analysis of the D. citri degradome.
Canker disease on fruit and foliage was under evaluation in the following 2014 trials. 1) Soil-applied Actigard was evaluated for the second growing season an EPA-approved Experimental Use Program (EUP) in two east coast grapefruit groves in a collaboration with Syngenta. In general disease pressure was much higher this season and started earlier. Even though the1st applications were made at the same timing as last season, they were late. These factors, along with tree size (8 ft. tall) in one of the locations were reflected in about 20% greater disease control on fruit after four 3.2 oz/acre applications of Actigard compared to copper program for canker control. In a replicated plot trial in an adjacent block of 8 ft. trees, soil drenches of Actigard and season long rotations with Admire and Platinum were effective for suppressing canker and protecting fruit on young Ruby Red grapefruit trees. The control responses were comparable to those obtained with 21-day copper sprays. Registration of non-insecticidal Actigard will address concerns for use of neonicotinoids of larger size trees. 2) Foliar applications of soluble copper and novel bactericidal formulations were compared with standard film-forming copper formulations. Several results were obtained from 2014 trials of Ruby Red grapefruit in Vero Beach and Hamlin orange in south-central Florida: 1) Nordox 75WG, ChampION++ 30WDG and Kocide 2000 and 3000 with ‘1.0 lb of metallic copper per application significantly reduced canker incidence to ~ 20% on Ruby Red grapefruit 2) Nordox 30/30 with 0.5 lb of metallic copper and an equivalent amount of metallic Zn was less effective than Nordox 75WG with 1.0 lb of metallic copper. Nordox 30/30 with 1.0 lb metallic copper gave similar control to Nordox 75WG with 1.0 lb of metallic copper as would be expected if copper is the primary mode of action. 3) Two new Zinkicide formulations from Dr. Swadesh Santra a Nano-chemist at Univ. Central Florida with similar Zn content to Nordox 30/30 reduced canker incidence to 7.0-9.0% which is significantly lower than for all other copper standards. 4) Nordox 75 WG for 5 sprays followed by 5 sprays of Magna-Bon at the 100 ppm rate was nearly as effective as 1.33 lb of Nordox alone at 50% of the metallic copper per season. 5) Magna-Bon at 100 ppm and experimental nano-formulations of copper with or without quaternary ammonium: CS-CuSiNPs, MVCuSiNG pH 4.0, MV1-CuSiNG pH 4.0 plus Quat, or quaternary ammonium alone, Quat DDAC @ 2X or 1X gave nearly equivalent canker control to the copper standards at lower metallic rate. 6) For ‘Hamlin’ orange with average rainfall, canker incidence on fruit and drop was only 10 fruit per tree. Fruit drop from other causes (mainly HLB) was as high as the canker induced drop. 7) The effect of HLB on Hamlin tree vigor, susceptibility of foliage and fruit to infection, as fruit drop may have contributed to the lack of differences among copper formulations in incidence of fruit canker or fruit drop due to canker 3) The performance of two applications of FireWall (streptomycin) with and without the penetrant Nanocanopy was evaluated in a Ruby Red grapefruit trial. Canker incidence on fruit treated with Firewall + Nanocanopy was no different from FireWall alone. In greenhouse assay, the residual activity of FireWall was 10 weeks post foliar application. This confirms that the streptomycin a.i. is locally systemic, remains active and moves acropetally in citrus shoots. 4) Cu bioavailability and residual activity on grapefruit for copper oxide (Nordox 75 WG) and the copper-zinc formulations Nordox 30-30 was evaluated during two periods of summer season (July and August). Zinc may be enhancing the bioavailbility of Cu from the copper oxide in the 30/30 formulation compared to 75 WG copper oxide alone. These results will be presented to the product manufacturer to gain their perspective on this result. 5) Selected results will be presented at the Florida Citrus Show in Ft. Pierce on January 2015
Citrus leafminer (CLM) is a major pest of citrus, causing direct damage and increased incidence/intensity of citrus canker. Pheromone traps are valuable for monitoring CLM but need further evaluation to optimize use for management. The objectives of the proposed project include: (1) assess species composition of trap catches, (2) optimize spatial and temporal distribution of pheromone traps. Sampling from Silver Strand B9 block and the Okaloacoochee Slough State Forest continues. Species composition in trap captures continues to be evaluated by analysis for Cytochrome Oxidase 1 (CO1) sequence generation incorporating an anchored hybrid enrichment probe set from Illumina HiSeq data with the Lemmon Lab at FSU. The probe set is designed to capture a tiled, 250 bp region of the 658 bp CO1 ‘barcode region’. Blast-searches against the comprehensive BOLD sequence library have allowed a faster, bulk-sequencing approach for identification of leafminers. ‘Vince Ficarrotta continues to process samples from P. citrella and related specimens that were shipped to us in 2013. He has been extracting DNA and sequencing samples for COI barcoding. The Kawahara lab recently hired an additional assistant to help with extractions. Vincent is now fully competent in the Geneious program and other related for maximum likelihood phylogenetic analysis, and is planning to submit COI barcodes to Genbank for archiving. So far, 12% of the barcoded Phyllocnistis specimens were identified as species other than P. citrella, and increased the known species of Florida Phyllocnistis species to five. Photography of specimens prior to extraction, so that new species collected in OK Slough can be distinguished from P. citrella by external morphological characters, also continues. Photographs of the specimens will help elucidate these characters in order to provide more accurate identifications. David Plotkin continued work photographing moths through a microscope and generating COI sequence data. David presented leaf-miner work at the ESA meeting in Portland Oregon in November. Work by senior undergraduate student, Emma Roulette, continued dissecting specimens of the two unknown species of Phyllocnistis that are attracted to CLM lures. Examinations of genitalia and comparisons with type specimens in the National Museum of Natural History are being conducted to verify identities. A manuscript: “Placement Density and Longevity of Pheromone Traps for Monitoring of Citrus Leafminer, Phyllocnistis citrella (Lepidoptera: Gracillariidae)” by P. Vanaclocha, M. M. Jones, C. Monz’ and P. A. Stansly has been submitted for publication in a refereed journal. We have initiated two studies to evaluate Rb uptake and detection in CLM for mark-recapture applications. In the first, potted citrus plants have been spiked with RbCl (99% pure – Sigma-Aldrich, St. Louis, MO) at concentrations of 0, 5, 10, 50, 100, 500 and 1000 ppm by introducing 100ml solutions into the potting media. Spiked plants have demonstrated no signs of toxicity to date. For the second, two artificial diet recipes have been prepared. We are currently evaluating these recipes. Artificial diet will be spiked at concentrations of 0, 5, 10, 50, 100, 500 and 1000 ppm RbCl. Adults reared from these diets will be evaluated for Rb concentration by spectrophotometry and potential toxic effects of Rb (mortality, prolonged development time and mass). Once a calibration curve is established, releases will be made and captures compared to background Rb concentrations to establish recapture rates and spatial features of CLM populations.
The canker field trial of Zinkicide compared to industry standard management on grapefruit is ongoing with final disease assessment for the 2014-2015 crop complete. We are planning to start applications this spring with the hopes of collecting a second year of disease control data to confirm positive results observed in the first season. The HLB trial in young Valencia trees continues with regular treatment and disease assessment. The timing of first treatment and the trees flush cycle make it difficult to determine efficacy this soon after initial treatment. We are preparing to collect yield data in the next few months. We will also be closely monitoring new flush for vigor, symptoms and Las titer. HLB symptom ratings were also collected for infected trees in the canker trial because treatment began earlier. However, the data may be biased because the reduction in canker could reduce leaf drop and cause the trees to look healthier.
Current HLB management strategy involves protection of citrus plants from HLB transmitting vector, Asian Citrus Psyllid (ACP) and adaptation of a good nutrition program for preventing root loss as there is no cure available to date. The objective of this research project is to develop and study a new non-phytotoxic, environmentally friendly film-forming ACP repellent material, called hereafter Organo-Silica Composite Film (OSCF). This material form a film barrier on plant surface upon spray application. This barrier is expected to change ACP feeding behavior. Previously we reported synthesis of six new OSCF formulations (OS-SG6 through OS-SG10) using different polymer and cross-linker ingredients. Phytotoxicity testing on Vinca sp and Persian Lime showed no plant tissue damage from any of these formulations. We also looked into potential heat trapping by OSCF material. By using an infra-red (IR) temperature sensor, we monitored the temperature the OSCF coated plant surface (including leaves and stem) at different time points post application of the OSCF materials. As a control, Surround’, a recommended pest deterrent based on Kaolin Clay was included in our studies. Four of the six candidate OSCF materials showed plant temperature fluctuations comparable to that of Surround’ after application on our testing citrus plant, Persian Lime. From the four candidate OSCF formulations that did not exhibit phytotoxicity, OS-SG6 and OS-SG10 were chosen as the most promising candidates. This selection was based on formulation stability at room temperature and minimal change of plant surface temperature before and after spray application (i.e. minimal heat trapping when compared to the Surround’ control). These two OSCF formulations were delivered to UF-CREC collaborators for efficacy testing which will be conducted in a Psyllid house. Detailed materials characterization results on the OS-SG6 and OS-SG10 formulations will be reported in future reports.
Successful area-wide management of the Asian citrus psyllid (ACP) and Huanglongbing (HLB) requires effective control of the psyllid in abandoned citrus groves. Since ACP adults are highly mobile, they can disperse from abandoned to productive citrus groves and spread HLB. ACP is susceptible to Isaria fumosorosea (Ifr), a native entomopathogenic fungi. ‘Autodissemination’ is insect dispersal of a pathogen to members of its own population. Our project goals are to develop and field test an ‘autodissemination’ system for inoculating ACP with Ifr and use these infected psyllids to instigate epizootics that will decimate ACP populations in abandoned citrus groves and; thereby, reduce movement of both ACP and HLB to nearby managed groves. The Ifr formulation is being provided by Dr. Mark Jackson (USDA-ARS, Peoria, IL) and is based on a south Texas isolate (Ifr 3581). During the first year of our project, we field-tested the durability and efficacy of different cardboard and plastic autodisseminator components fabricated to our specifications by AlphaScent Inc. By September of 2014, we determined which components were most suitable for Texas conditions and ordered sufficient quantities from AlphaScent for fall and winter trials (September to December). Due to a delay in obtaining the raw plastic materials required, AlphaScent was not able to deliver the components until the second week of October 2014. During the fall and early winter (October-December) of 2014, we collaborated with Paramount Citrus to repeat field trials in Hildago County with one of their managed groves and an adjacent abandoned grove. The site was a managed Valencia Orange grove next to an abandoned Rio Red Grapefruit grove. The managed grove was treated with insecticides for ACP earlier in the fall and flood-irrigated before our field trial. The abandoned grove was not treated with insecticides or irrigated. At this site, we evaluated the impact of autodisseminators on ACP movement and infestation in plots of abandoned citrus trees and managed citrus trees. For treatment plots, we hung pairs of autodisseminators coated with Ifr-spore formulation and equipped with citrus-blend lures in trees on the edge rows of the abandoned groves. For control plots, we hung autodisseminators with only citrus-blend lures. One week before the start of our trial, we hung ACP sticky traps on the managed trees directly across from the abandoned trees to assess ACP movement from the abandoned grove to the managed grove prior to deployment of autodisseminators. To facilitate monitoring of ACP movement during our trial, we captured and released up to 4800 ACP adults marked with fluorescent powder on abandoned trees every week for three consecutive weeks. We used different fluorescent colors to distinguish psyllids released in treatment plots from psyllids released in control plots. Every seven days during the first to fourth weeks of the trials, we inspected and replaced dispensers and ACP traps. During the fifth week, we also ‘tap sampled’ managed trees and abandoned trees. Movement of ACP from abandoned trees to managed trees was greatly reduced by the deployment of autodisseminators coated with Ifr-spore formulation. Prior to autodisseminator deployment, the numbers of ACP adults trapped on managed trees in treatment plots and control plots were not significantly different. After autodisseminator deployment, weekly trap counts of ACP adults were 28-50 % less for treatment plots compared to control plots. Our findings showed that deployment of Ifr-autodisseminators at the end of the active growing season for commercial citrus in Texas can reduce the numbers of ACP adults moving from abandoned groves to managed groves. We plan to repeat this trial during the spring (March-May) of 2015 to determine if our autodisseminators can reduce movement of ACP from abandoned groves to managed groves at the start of the active growing season in Texas. Deployment of Ifr-autodisseminators at the end and / or start of the growing season should minimize intraguild interactions between Ifr and Tamarixa radiata.
Work continues with a susceptible CLM colony towards estimation of LD50s and LD80s for some commonly used insecticides. Larvae and adults from the susceptible colony are exposed for 48 h to different doses (0; 0.01; 0.03; 0.1; 0.3; 1; 3; 10; 30; 50; 100, 300, 600 and 1000 ppm) of: Actara 25WG, Agri-Mek 0.15EC, Cyazypyr, Danitol, Delegate WG, Dimethoate, Intrepid 2F and Micromite 80WGS. For CLM larvae: Actara (thiamethoxam) LD50=4.18 (CL95: 1.86-8.51) ppm, LD80=524.12 (CL95 209.70-1827) (n = 1047; .2 = 9.54; d.f = 10; Heterogeneity = 0.95); Agri-Mek (abamectin): LD50= 0.314 ppm (CL95: 0.13-0.69), LD80=16.68 ppm (CL95: 6.14-78.56) (n = 1249; .2 = 13.17; d.f = 8; Heterogeneity = 1.65); Cyazypyr (cyantraniliprole): LD50=43.36 ppm (CL95: 15.80-131.55), LD80=49,413 (CL95: 0.19E+07) ppm, (n = 1,196; .2 = 7.64; d.f = 8; Heterogeneity = 0.96); Danitol (zeta-cypermethrin): LD50=381.78 (CL95: 86.33-11,771), LD80= 37, 220 (CL95: 31.70-53761.1) ppm (n = 561; .2 = 5.29; d.f = 9; Heterogeneity = 0.59); Delegate (spinetoram): LD50=2.67 (CL95: 0.87-6.42), LD80= 93.75 (CL95: 33.89-489.58) ppm (n = 938; .2 = 10.69; d.f = 7; Heterogeneity = 1.53); Dimethoate (dimethoate) LD50=1.56 ppm (CL95: 0.15-56.31) and LD80=497.45 ppm (CL95: 22.27-0.37E+09) (n = 546; .2 = 13.07; d.f = 6; Heterogeneity = 2.18); and Micromite (diflubenzuron): LD50=74.30 (CL95: 13.63-285.15), LD80=5,723 ppm (CL95: 994-0.129E+07), (n = 816; .2 = 19.92; d.f = 9; Heterogeneity = 2.21). For adults: Micromite LD50=387.40 (CL95: 143-2817), LD80= 9392 (CL95: 0.192E+07) ppm have been estimated but further replicates are required. Contact toxicity for Intrepid LD50 and LD80 values remain inestimable at the concentrations tested (up to 10 ppm). We will continue tests of higher concentrations. Delegate (spinetoram) LD50=2.84 (CL95: 0.77-73.74), LD80= 183.52 (CL95: 16.47-0.44E+07) ppm (n = 430; .2 = 6.30; d.f = 4; Heterogeneity = 1.58). Additional tests of Actara 25WG, Agri-Mek 0.15EC, Cyazypyr, Danitol, and Dimethoate are slated for the coming weeks. Work also continues on the insect growth regulator, methoxyfenozide (Intrepid). This insecticide acts as an ecdysone agonist, interfering with molting, and exhibiting low contact toxicity as indicated above. Therefore, we have initiated larval bioassays on caged branches infested with CLM in the laboratory. Applications of doses of 0; 0.01; 0.03; 0.1; 0.3; 1; 3; 10; 30; 50; 100, 300, 600 and 1000 ppm are applied when infested leaves contain 3-4th instar larvae. Evaluations of mortality will be conducted 14d after application. We have also initiated work to develop a bioassay method using artificial diet and two diet recipes and are being tested. We are developing a method to collect large numbers of CLM eggs from the established laboratory colony. Eggs will be transferred to cups containing insecticide-spiked artificial diet to determine diagnostic doses for this compound against larval CLM. Field-collected larvae are currently being evaluated for their responses to these insecticides. Although cases of field resistance have thus far not been observed, populations of CLM are currently high in groves in SW Florida and subjected to frequent insecticide applications for control of ACP, CLM and other pests.
Plasmid vector and helper DNA construct microinjections into ACP embryos have continued, though the survival of G0 nymphs has decreased from 25% to 15%. This is due in part to decreased numbers of fertilized eggs and embryonic viability which may be the result of seasonal changes and/or ACP colony inbreeding. The survival of hatched nymphs from injected eggs continues to be challenging with less than 20% surviving the transfer to flush (with most becoming stuck to double-stick tape), and less than 20% of these G0 nymphs surviving to adulthood. From 19 G0 adults, 8 were fertile and none yielded putative fluorescent transformed progeny. Plant health and flush quality have been variable affecting ACP viability, and improvements in both remain a high priority. To improve the number of surviving nymphs after hatching, alternatives to double-stick tape (used to hold eggs in place for injection) are being tested, in addition to setting up eggs on filter paper (that can be moistened) versus glass slides. Initial tests with two types of glue (E6000 and Tacky glue) have resulted in 60-80% nymphal hatching from uninjected eggs on either slides or filter paper. Once reaffirmed, survival of nymphs from glued injected eggs will be tested with and without moistened filter paper. While we expect the transformation marker gene to be expressed in ACP, this must be confirmed, and RT-PCR tests for transient DsRed fluorescent protein gene expression regulated by the IE1 and polyubiquitin promoters have been initiated. ACP transformation tests (since 10/1/2014) ‘ 3,680 eggs injected ‘ 550 G0 nymphs hatched (~15%) ‘ 99 transferred to flush ‘ 19 G0 adults ‘ 8 fertile G0 adults ‘ 40 G1 progeny screened – none fluorescent In addition to the above, assays are continuing to confirm passage of transinfected Wolbachia cell lines from injected parents (P1) to offspring (F1 generation). The survival of injected psyllids is high, with around 80% survival. Horizontal, or transovarial, passage of injected Wolbachia appears to be possible for at least one Wolbachia strain, however the frequency of passage to offspring appears to occur at a low frequency. This may be due to limitations of the injection procedure (e.g. developmental stage of psyllid, duration of infection with foreign Wolbachia, titer of foreign Wolbachia in P1 psyllids). Alternatively, injected Wolbachia strains may not be colonizing reproductive tissues sufficiently to promote transovarial passage to F1 psyllids. Concurrently with these studies, psyllids injected and naturally infected with Wolbachia strains of interests are being evaluated in bioassays to determine their capacity for induction of cytoplasmic incompatability and pathogen interference.
The large-scale validation of citrus leafminer (CLM) disruption with the ISCA DCEPT CLM technology has continued. During this past quarter we have continued to assess efficacy data, although many of the experiments were suspended during the dormant winter season. Our role continues to be with assisting in collection of efficacy data by trapping CLM males with pheromone traps as a surrogate measure of mating disruption. We have been analyzing our data and also modeling the economic return of using this product. Specifically, we have been investigating the need for two pheromone components as the active ingredient versus only a single component, which would make the product more cost effective. Cost contour plots of our data indicated that there was an interaction between the amount of pheromone released and the number of trees treated with pheromone. However, the difference in costs between a 3:1 blend (two component-more expensive) and triene only (one component-less expensive) application were insubstantial. This economic analysis suggests that choosing a triene application at a relatively low release rate with a point source density of approximately 89% (40 treated trees out of 45 total trees per plot) should be economically more desirable to growers than the 3:1 blend at the same rate of application. Although damage by CLM was not eliminated in these trials, there appears to be significant and fairly consistent reduction in damage. Planning for 2015 field trials will begin with a meeting at the Florida citrus show in Fort Pierce in late January.
The overall objective of this project is to investigate the feeding deterrence provided by reflective colored kaolin clays on citrus plants from ACP. Previously we reported the development of colored kaolin clays using select dyes, including FDA certified colorants approved for agricultural use by the EPA (list ‘180.920), which covers the range of visible spectrum for coating clays. During this reporting period a scale up synthesis to produce colored clays was carried out for ACP deterrence assays and further characterization. Ten different colored clays were developed with reflectance in the visible region with dyes e.g. FD&C Blue 1, FD&C Red 40, Basic Blue 54, Crystal Violet, and FD&C Yellow 6, D&C Violet 2 etc. ACP greenhouse bioassays are underway to determine reflective wavelength deterrence efficiency. The characterization of the colored clays has been carried out to determine their optical and physical properties. The optical properties of the clays have been determined by total reflectance measurements using an integrating sphere (Lambda 800 UV/VIS Spectrometer). Measurements of all modified clay materials were made using unmodified kaolin clay as a reference. Reflection measurements showed that each material has an altered absorption peak and reflectance not exhibited by the unmodified clay. For example modified clays showed high reflectance in 350- 490 nm (D&C Violet 2), 350- 550nm (FD&C Blue 1), 575-750nm (FD&C Red 40), 505-750nm (FD&C Yellow 5), and 625 -750nm (Basic Violet 16), covering the visible spectrum range. Future experiments are being planned to investigate the change in the reflectance of citrus leaves with developed colored clay materials. The dispersion of the clay particles in suspension, the ability to apply as foliar spray, and eventual adhesion to the leaves is dependent upon the surface charge of the particles. Zeta Potential experiments were performed for each colored clay material to estimate the surface charge. While the unmodified kaolin clay zeta potential was found to be negative, the surface modifications and doping of clay material with the dyes, led to significant changes in the zeta potential. These results will help the selection of appropriate adjuvants necessary to provide dispersion of clay particles. Tests are also being conducted on modified kaolin formulations to increase rain-fastness.
Issue date for this grant was 13 July 2012. Objectives are: 1. Assess effects of abiotic factors (light quality, photoperiod, air flow, temperature fluctuations) on psyllid movement, 2. Evaluate physiological limits and biotic factors effecting of movement including feeding, egg load, infection status, and population density, 3. Evaluate techniques for tracking psyllid movement in the field for mark recapture studies, 4. Characterize seasonal patterns of ACP distribution and movement at different scales in the field, 5. Develop strategies to protect young trees from colonization by ACP utilizing UV reflection for repellency and insecticide treated trap crops (such as Bergera koenigii) to attract and kill. Objective 1. Bioasssays using a wind tunnel to determine ACP response to different stimuli have shown that ACP movement stopped at temperatures below 67 F, and in the absence of light. Wind speeds above 4.25 mph reduced movement as did humidity over 70%. ACP tended to move with the wind when lighting is uniform but also moved into the wind. Increased light intensity did not increase ACP movement and lighting type with varying spectrum of light intensity did not impact ACP behavior. Objective 2. Dr. Stelinski’s group recently analyzed results from three field experiments where citrus trees were sprayed with egg and milk proteins. Subsequently, sticky traps were deposited at various distances from the sprayed area. Psyllids collected were submitted to ELISA for detection of milk and egg proteins. Therefore, we knew that each psyllid positive for milk protein as indicated by ELISA had dispersed from the sprayed area. The result of these field experiments demonstrated that: 1) psyllid movement from abandoned citrus groves to adjacent managed citrus groves was greatest during the spring and summer months and decreased significantly during the colder winter months; 2) Psyllids were able to traverse geographic barriers such as roads and fallow fields; 3) Psyllids were able to disperse at least 2 km within 12 days; 4) wind direction was not correlated with the number of marked psyllids captured; 5) grove management and presence of flush were the most important factors that increased the capture of marked psyllids. The results further confirm that abandoned citrus groves in Florida serve as reservoirs for ACP which can disperse over long distances despite geographical barriers. These results are in accordance with our laboratory results obtained with a flight mill. Objective 3. Sticky cards were placed in a field of young citrus placed between trees on tomato stakes and monitored hourly for 15 different 24 hour periods. Wind speed, air temperature, and solar radiation were found to be good indicators of ACP movement while barometric pressure was not. Rainfall and time of day were marginally correlated to ACP movement. No ACP were observed to land on sticky cards at night. Objective 4. Two-sided clear and yellow sticky cards were placed in several groves to compare directional responses in an attempt to determine whether ACP adults flying in one direction may turn back and be captured on the opposite side of the yellow card. The results of the clear vs. yellow sticky cards thus far are inconclusive due to low ACP movement in the test areas. Additional attempts will be made to determine differences in ACP response to the two types of sticky cards when movement and populations increase in the spring. Objective 5. A paper reporting positive results using UV reflective mulch to repel ACP the first 2 years after planting has been published in the refereed journal Pest Management Science. Tap sampling has reveled the mulch still works to reduce the ACP populations on trees 2.5 years old. In another trial, repellency was demonstrated by the result that fewer ACP were captured on sticky traps set at 1 m in reflective mulch compared to 2 m, where as more ACP were captured more at 1m than 2m over white mulch or no mulch (grass). Manscripts are being prepared on environmental factors influencing psyllid behavior and movement and dispersal patterns of the Asian citrus psyllid.
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