Five studies were conducted to investigate factors influencing acquisition and inoculation of Candidatus Liberibacter asiaticus (Las) by Diaphorina. citri, e.g vector developmental stage, feeding periods, leaf age, symptom expression/bacterial titer in infected trees and insecticide treatments, in order to improved Huanglongbing management strategies. In study 1, we compared the ability to acquire Las by different developmental stadia and adults of D. citri on Las-infected plants over a range of acquisition access periods (AAPs). Mean acquisition efficiencies by adults and 1st, 2nd, 3rd, 4th and 5th instars were 42.9, 68.4, 60, 66.6, 84 and 65.3%, respectively. Third-instar nymphs and adults acquired Las in AAPs as short as 30 min, but efficient acquisition (>90%) required longer AAPs (1 and 4 days, respectively). Transmission efficiency reached a maximum of 9.4 and 4.8% (per individual) with a 12-h AAP by nymphs and adults, respectively. By comparing electrical penetration graphs (EPG) of nymphs and adults, we found no significant variations in probing behavior that could explain differences in transmission efficiency. In study 2, we observed that efficient acquisition depends on the availability of young leaves in Las-infected plants, apparently because phloem ingestion by D. citri adults is more frequent and last longer on younger leaves . In study 3, vector competence was assessed as the proportion of successful acquisition events by adult psyllids over a gradient in Las infection levels in citrus trees and over time after graft inoculation of this pathogen. We observed that plant infection levels (pathogen titer) increased rapidly over time, saturating at uniformly high levels (approximately 108 CN per g of plant tissue) near 200 days after inoculation ‘ the same time at which all infected trees first showed disease symptoms. Pathogen acquisition by vectors was positively associated with pathogen titer and time since inoculation, with acquisition occurring as early as the first measurement, at 60 days after inoculation. This result is problematic from a disease management perspective because it means that infected citrus trees may be sources of infectious vectors well before the disease symptoms become evident. Therefore, there is ample potential for psyllids to acquire the pathogen from trees during the asymptomatic (or latent) phase of infection, reducing the efficacy of diseased tree removal (rouging). In study 4, we conducted detailed transmission experiments coupled with EPG analysis of D. citri probing behavior in order to determine minimum time required for Las acquisition and inoculation, as well as the latent period. We observed Las transmission within an inoculation access period (IAP) as short as 45 min. EPG results showed that both acquisition and inoculation occur during the phloem phase, mainly during sustained sap ingestion (waveform E2). Nymphs and adults take an average of 63 min (16 – 241) and 75 min (16 – 241), respectively, to start E2. Thus, psyllids require ’15 min on citrus plants to reach citrus sieve elements and possibly acquire and inoculate Las. Once Las is acquired, D. citri is able to inoculate it in healthy plants after a latent period of at least 10 days at 25C. The mean latent period (LP50) is 16.5 and 18 days after acquisition by nymphs and adults, respectively. In Study 5, we evaluated the effects of insecticide mode of application (foliar vs. drench) on Las acquisition by D. citri nymphs and adults. Insecticides were not effective to avoid Las inoculation to healthy citrus trees, although they drastically affected bacterial acquisition. Therefore, insecticides should be more effective in preventing secondary spread, which requires bacterial acquisition by psyllids on infected plants, than primary spread by immigrating psyllids, which may be already infective when landing on healthy citrus trees in the orchard. The information on transmission biology of Las by D. citri generated in this project have direct implications on management of HLB and highlight the need of additional research to develop new control tools.
During this season, we are focusing our efforts on evaluating different formulations of imidacloprid. This research objective is in response to grower questions about the consistency of different imidacloprid formulations at a time when there are multiple generic products available. We are running two trials in commercial citrus groves – one trial on minneola tangelos and a second trial on grapefruits. We have completed 2 years of trials where we evaluated the uptake of clothianidin applied by soil drench and trunk spray. The results of the 2011 trials were very promising, whereas the results of the 2012 trials showed no residues in the trees. Due to the inconsistency of the results, we are conducting a third trial at Lindcove during the 2013 season. Soil drench and trunk spray applications at a single timing are currently being evaluated. In 2012, we evaluated the efficacy of thiamethoxam treatments on navel and Valencia oranges at several field sites in Redlands. At one of two sites where treatments were applied in October, thiamethoxam was only detected at 4 weeks. At a third site, where treatments were applied in mid-July, no thiamethoxam was detected up to 16 weeks after applications. At the latter site, monitoring was only begun at 8 weeks, so thiamethoxam residues may already have been established within the tree and then dissipated. Nevertheless, our data shows that thiamethoxam is not a persistent chemical in citrus trees. Our objective to conduct imidacloprid residue analysis on fruit from treated trees is now complete. Fruit from trees treated at 2 timings (Feb. and Mar.) with a 2X label rate of imidacloprid from 3 field sites in Bakersfield (Valencia oranges), Lindcove (navel oranges) and Hemet (grapefruit) have now been tested for residues. Samples were collected for up to 8 weeks after treatments at the Bakersfield and Lindcove sites and 16 weeks at the Hemet site. At the Bakersfield and Lindcove sites, residues were not detected (limit of detection of the assay was 0.01 ppm). At the Hemet site, residues were not detected in trees treated with a single application. However, in trees that were treated in 2 successive years with the 2X rate, residues of 0.01 ppm were detected, which is below the MRL established for citrus in the USA.
During this season, we are focusing our efforts on evaluating different formulations of imidacloprid. This research objective is in response to grower questions about the consistency of different imidacloprid formulations at a time when there are multiple generic products available. We are running two trials in commercial citrus groves – one trial on minneola tangelos and a second trial on grapefruits. We have completed 2 years of trials where we evaluated the uptake of clothianidin applied by soil drench and trunk spray. The results of the 2011 trials were very promising, whereas the results of the 2012 trials showed no residues in the trees. Due to the inconsistency of the results, we are conducting a third trial at Lindcove during the 2013 season. Soil drench and trunk spray applications at a single timing are currently being evaluated. In 2012, we evaluated the efficacy of thiamethoxam treatments on navel and Valencia oranges at several field sites in Redlands. At one of two sites where treatments were applied in October, thiamethoxam was only detected at 4 weeks. At a third site, where treatments were applied in mid-July, no thiamethoxam was detected up to 16 weeks after applications. At the latter site, monitoring was only begun at 8 weeks, so thiamethoxam residues may already have been established within the tree and then dissipated. Nevertheless, our data shows that thiamethoxam is not a persistent chemical in citrus trees. Our objective to conduct imidacloprid residue analysis on fruit from treated trees is now complete. Fruit from trees treated at 2 timings (Feb. and Mar.) with a 2X label rate of imidacloprid from 3 field sites in Bakersfield (Valencia oranges), Lindcove (navel oranges) and Hemet (grapefruit) have now been tested for residues. Samples were collected for up to 8 weeks after treatments at the Bakersfield and Lindcove sites and 16 weeks at the Hemet site. At the Bakersfield and Lindcove sites, residues were not detected (limit of detection of the assay was 0.01 ppm). At the Hemet site, residues were not detected in trees treated with a single application. However, in trees that were treated in 2 successive years with the 2X rate, residues of 0.01 ppm were detected, which is below the MRL established for citrus in the USA.
We are currently pursuing our first objective to evaluate the effects of nursery practices (watering, citrus variety, potting media) on imidacloprid uptake and retention. Two citrus varieties were selected for this trial – Parent Washington navel and Limoneira 8A lemon, both on Carrizo rootstock. The trees were budded in June 2012 and grown in 5′ pots at LREC. Trees were transferred to UCR Agricultural Operations on June 13 2013 and transplanted on June 17 to no. 5 pots. Two potting media were chosen for the study that differed principally in the proportions of sand (10-30%) and redwood (40-60%). The evapotranspiration rate was determined for the two soil mixes and this data was used to select three watering levels for the study. The three levels were defined as ‘replacement watering’ (100-120% ET), ‘overwatering (150-200% ET) and ‘severe overwatering’ (300-400% ET). For the trial, trees were placed in a 12 x 25 grid pattern and randomized according to variety, soil mix and watering level. When the trees had adapted to their new potting media environment, they were treated with 0.33 mls Admire Pro per cu ft potting media. Monitoring for ACP on all trees is being conducted at monthly intervals for the duration of the trial, and imidacloprid residues are being determined for all trees by ELISA at 2, 4, 8, 16 and 32 weeks post-treatment.
This is a cooperative research project between Co-PIs Joseph Morse, Jim Bethke, Frank Byrne, Beth Grafton-Cardwell, and Kris Godfrey. One objective is to coordinate with researchers working on chemical control of ACP in Florida, Texas, Arizona, and elsewhere. Towards that end, Grafton-Cardwell, Godfrey, and Morse met with Jawwad Quershi, Phil Stansly, and 2 CA PCAs in Orlando 2-5-13 at the Third International HLB conference to discuss cooperative trials focusing on organic products; the same group toured the UF Immokalee Research Center 2-6-13 to see ongoing field trials; and Byrne and Morse traveled to the UF Lake Alfred Research Center with 3 CA PCAs 2-8-13 to discuss ongoing research and collaboration with Michael Rogers and Lukasz Stelinski. The Orlando HLB conference is an excellent opportunity to network with other scientists working on ACP and HLB and much was learned and discussed. In a similar fashion, many of the PIs will attend the CHRP meeting in Denver 15-17 October 2013. We are rearing ACP in a contained greenhouse at the Chula Vista Insectary (San Diego County; about 6 miles north of the Mexican border) under permit (#2847) from CDFA. To date, Jim Bethke et al. have run 24 screening trials (11 focusing on organic products). A second location where we are working with ACP is at UC Riverside, working under permit inside the UCR Insectary Facility. Frank Byrne is conducting trials on various neonicotinoid insecticides and Morse is evaluating the baseline susceptibility of CA ACP to various pesticides (30 trials to date) in comparison with studies done in Florida. Three field trials have been run to date evaluating organic products useful in control of ACP; the first 10-4-12 evaluated the impact of 5 treatments on adult ACP; the second trial 3-5-13 evaluated 8 treatments against young nymphs and eggs; the third 8-1-13 was recently concluded in San Juan Capistrano on organic lemons in cooperation with PCA Matt Hand and compared 7 treatments. Kris Godfrey obtained a permit to rear ACP inside UC Davis’ Contained Research Facility and now has a healthy colony going. She will focus on evaluating new organic products. One study has been concluded looking at Pest Out (combination of cottonseed, clove, and garlic oils) and she has initiated studies looking at the impacts of Grandevo on oviposition and development of ACP. Beth Grafton-Cardwell has updated online ACP pest management guidelines. She serves as a focal point for communicating with ongoing ACP management programs in the SJV, Ventura, and elsewhere. Both she and Morse also serve on CDFA’s recently convened Science Advisory Panel dealing with ACP and HLB; 2 conference calls of the SAP have been conducted to date. In summary, we continue to expand our ability to conduct ACP research in California with a clear focus on management of HLB (rather than ACP).
Imidacloprid (IMD) fate and transport in Immokalee Fine Sand (IFS) soils for three citrus tree ages was studied in SW Florida Flatwoods at selected irrigation rates. Sorption and degradation studies have shown That IMD is a weakly-sorbed (log Koc, 1.1-2.4) and non-persistent chemical (half-lives 0.9-2.3 years) with high potential for leaching. We also established field experiments in three age classes of Hamlin trees with micro-sprinkler irrigation at three irrigation rates, during growing seasons between 2011 and 2013. IMD was soil-drench applied in the root zone (PA), and in a control zone not affected by roots (NPA). IMD concentrations (.g g soil-1) were higher for the NPA than PA as a function of time due to uptake by the citrus trees. IMD leached out of the root zone about 3 to 4 weeks after application in the summer, and about 6 to 8 weeks after application during spring. Nonetheless, there was effective systemic control of the CG vector Diaphorina citri Kuwayama (ACP) at about 2 weeks after application, where treated trees showed consistently lower ACP adults and immatures infestation, continuing for at least 8 weeks. A method to analyze IMD from citrus tissue (ng g-1) was developed using HPLC-MS/MS detection, and preliminary data are in close agreement with our findings in soils and insect counts on trees. Leaf tissue concentrations of IMD increase for the initial two weeks and stay constant for an additional 6 to 8 weeks. Leaf IMD concentrations are significantly greater with increased irrigation amount. Close irrigation and rainfall monitoring during IMD application is of the utmost importance to avoid leaching problems in groundwater resources of SW Florida, and to ensure systemic control of the ACP.
Progress on development of an acoustic trap to capture male Asian citrus psyllids (ACP) that are searching for females on citrus tree branches continued in the first second of 2013. In early stages of the research, we had programmed an Arduino Uno microcomputer system to detect and record psyllid-produced vibrations from a microphone attached to a citrus tree branch. In the second phase of the research, we programmed the system to play back ACP communicatory signals whenever it detects a male calling on the branch. The signal playback is done through an inexpensive piezoelectric buzzer system. During this last quarter, we have begun incorporated the microphone-buzzer microcomputer-controlled system into traps that were tested in field environments. Several technical issues were identified in the field tests that restricted the trapping efficiency and the duration of time that the traps could be used in the field. Both the traps and the microprocessor systems are being modified to address these issues. In March, we presented some of the initial findings of the research at the 3rd International Research Conference on Huanglongbing in Orlando, FL. One paper on the initial phase of the study has been published in the Annals of the Entomological Society of America, and two papers discussing a later phase of research have been submitted to the Florida Entomological Society. A fourth paper is in progress on the results of laboratory tests using the new system that plays back ACP communicatory signals whenever it detects a male calling on the branch.
This proposal aims to continue improvement to a novel psyllid trap and to use the trap to gather new information on the behavior, biology, population dynamics and biological control of ACP/Candidatus Liberibacter asiaticus. Lab and field testing was and continues to be conducted to increase trap efficiency by exploiting unique vector behaviors in response to traps and behaviorally active components. We continue to conduct field and laboratory studies toward obtaining an understanding of ACP trap response behavior by manipulating psyllid behavior around the trap or farther away so that they are moved close enough to the trap to perceive it (i.e., increase trap active distance). We have a number of positive results from our bioassays with which to attempt to increase trap efficiency but have yet to reach a level of trap capture rate which is a satisfactory representation of ACP populations in the vicinity of the trap or equal to the capture rate of the current industry standard, the AlphaScents green-yellow sticky trap . During this quarter we have cooperated with several USDA-ARS personnel to develop, test and incorporate other trap features that affect psyllid visual response but also involve other sensory modalities including sound productions during psyllid courtship. This work is ongoing and more experiments are being conducted. We have run in the last 45 days an extensive field test of our trap prototypes in Ft Pierce groves to discern what trap features are key to exploiting psyllid behavior to increase capture rate. We have also used combinations of semiochemical lures that supposedly attract psyllids provided by other scientists. So far these have not made a large difference in capture rates of any traps tested. Following another round of field tests with new traps improved based on the field data gathered this quarter, we will initiate the areawide psyllid sampling objective.
Xambr’ Paran’ Brazil Plots: The experiment is designed to quantify the effect of windbreaks, copper sprays, and insecticide sprays individual and combined effects on citrus canker management. In 2010 replicated field plots were established. I was soon evident that the screens used as windbreaks were not strong enough to provide the required effect. Thus in addition to the screens, we established natural windbreaks using Casuarina. Casuarina trees that are currently ~7 m high. There were a few new canker-affected plants in the experimental area that will be pruned every 3 weeks. Trees are well established now and have developed enough canopy to allow inoculation and to start the experiment in September/October 2013. During 2013 inspections, 25 plants were discovered with HLB symptoms. Florida Windbreak plots: The final data taken through September 2012 from the preliminary experiments last season confirmed that the number of wind gusts >11 mph increased with distance from the windbreak whether measured within the north-to-south rows or across the rows from east to west. Consequently, the highest fruit disease recorded in September in the cv. Valencia orange orchard (10 percent fruit cankered) was in the center of the 11-acre block and the lowest incidence (<2 percent fruit cankered) was in the east-west row location nearest the windbreak. The weather stations at the two locations (cv. Valencia orange, Estes Farm, Indian River Co. and cv. Rubyred grapefruit, Scott Farms, St. Lucie Co.) continue to be monitored this season with weather data being recorded and downloaded on a monthly basis. The disease data on leaves and the first sample on fruit have been taken and will be related to the wind and leaf wetness conditions from the weather data recorded in these orchards during the 2013 season. Programmable leaf wettnes controller: Further tests on the 'pin sensor' have proven problematic, as water retention control is difficult. First, to facilitate putting the silk onto the pins, the pins were lengthened off the board (approximately 1 cm). This allowed for 'cavities' between the pins, which allows too much water to be held, which lead to drying times being too long. We also determined that the board itself that the pins are mounted in will hold water, especially with the silk on top of it, which again led to longer water retention times. So, the next series of tests will include coating this board to prevent water uptake (we will attempt to use wax to coat the board to emulate the leaf surface a bit), followed by trying to keep the pins long enough to put through the silk, yet not be so long as to create cavities. Finally, a future test would be to NOT push the pins through the silk, but to keep the silk taunt across the tops of the pins, thus using the silk as not only a water absorption layer, but also a 'blocker' to prevent the water from sitting anywhere else within the sensor. Project publications: (In addition to the 7 previously reported publications relative to this project, the following was recently submitted as well) Bock, C. H., Gottwald, T. R. and Graham, J. H. 2013. A comparison of the bioassay test and culture to detect Xanthomonas citri subsp. citri . Plant Pathology 62: xx-xx.
1. Development and testing of efficient methods of statistical inference to estimate epidemiological parameters from maps of emerging epidemic.) Using citrus canker as a test system, we are developing and testing quantitative models to link weather variables to the rate of disease spread. We are also analysing models for anisotropic dispersal of the pathogen, in order to correlate the direction and extent of pathogen dispersal with wind direction/strength during extreme weather events (rainstorms). The outcome will be a set of epidemiological parameters that can be used as inputs for predictive modelling, with (at least) two different final goals: i. To produce risk maps for disease spread for large (state-wide) geographical areas and long time scales (several years), under different “what if” scenarios for favorable and unfavorable weather conditions; and ii. To produce a risk map on a smaller (county-wide) scale in response to a local storm. With knowledge of the distribution of disease prior to the storm, it should be possible to estimate the new distribution of disease using the strength and direction of wind, and other relevant weather parameters. This would help directing a post-storm sampling procedure (e.g. which areas surveyors should target for new symptoms). We have developed and tested a set of advanced spatial statistical methods for goodness of fit. The methods are based on the comparison of simulated and observed spatial patterns of infection. They have been successfully tested using model estimates for the Miami outbreak, and they were able to discriminate between good and less good dispersal models. Such methods are also a fundamental ingredient, hence a significant step forward, for the ABC (simulation-based) estimation algorithms that are currently being developed. The methodology being tested with citrus canker can be generalized to HLB. A paper on Markov chain Monte Carlo for estimation of epidemiological parameters in the Miami area has been submitted. 2. Develop a generic epidemiological model that can be used to compare control scenarios and to optimize the probability of controlling and managing high-risk pathogens of agricultural significance. We have built a model that represents the spread of HLB within an individual tree. Successful spread of the pathogen depends on transmission both within the vascular system of the tree and between leaves via psyllid vectors. The relative importance of these transmission routes is being explored, with the speed of spread of the pathogen dependent on whether transmission via psyllid vectors is frequency- or density- dependent: this depends on the population density of psyllids relative to the number of leaves. A novel feature of our modelling approach is that it explicitly incorporates the ability of Las bacteria to survive in the vascular system of the plant. Control: We are using this model for a variety of purposes. Firstly, we are investigating and comparing the efficacy of roguing, application of insecticide and the use of nutritional products and thermotherapy for disease control and mitigation. Secondly, we are using it to inform grove scale models for the spread of HLB. Grove scale models typically assume that each citrus plant is either entirely infected or not (instead of considering the proportion of the tree that is infected). This model is being used to inform the force of infection of each infected citrus tree in grove scale models. 3. Develop user friendly model ‘front ends’ that can be used by researchers and regulatory agencies. We are finalising the publication on Webidemics (http://www.webidemics.com/), the user-friendly front-end for control of citrus disease.
Soil application of systemic neonicotinoid insecticides for control of psyllid vectors of Huanglongbing disease on young citrus trees also produces season-long SAR control of citrus canker caused by Xanthomonas citri subsp. citri. The neonicotinoids imidacloprid (IMID; Admire Pro, Bayer) and thiamethoxam (THIA;Platinum, Syngenta) were compared with soil or sprinkler applications of the commercial SAR inducer acibenzolar-S-methyl (ASM, Actigard, Syngenta) and foliar sprays of copper hydroxide (CH) and/or streptomycin (STREP) to evaluate their effects on the percentage of canker-infected leaves on 2-yr-old ‘Vernia’ orange and 3-yr-old ‘Ray Ruby’ grapefruit trees in Southeast Florida. All treatments significantly reduced incidence of foliar canker compared to the untreated check. Soil drenches of ASM and season long rotations with IMID and THIA were highly effective for suppressing foliar canker on young grapefruit and orange trees under weather conditions absent of high intensity rains or tropical storms. Sprinkler application of ASM was less effective than soil drench. The level of control for SAR treatments was comparable to eleven 21-day interval sprays of CH and/or STREP. SAR induced by soil-applied insecticides provides substantial benefits for canker disease management on young citrus trees that may be augmented with ASM. Currently, use of neo-nicotinoids insecticides for control of the psyllids and leafminer is limited to trees less than 2.75 m tall in part due to the potential risk for leaching of soil applied materials into the groundwater at the higher rates required for pest control on larger size trees. An alternative approach is spray application of the chemicals on the trunk. Four soil drench or trunk applications of IMID and THIA and ASM were compared to untreated controls and 9 or 10 standard 21-day interval copper sprays for protection of foliage and fruit on 5 to 6-yr old ‘Ray Ruby’ grapefruit trees. Soil drench and trunk applications similarly reduced the incidence of canker lesions on foliage and fruit but were less effective than copper sprays. SAR inducers appeared to protect fruit by reducing incidence of foliar disease, and thereby, when integrated with 21-day interval copper sprays may improve control canker on young, fruiting trees. Currently, we are collaborating with Syngenta in an EPA-approved Experimental Use Program (EUP) in two east coast grapefruit groves to support the labeling of Actigard (ASM) integrated with copper sprays for control of canker on bearing grapefruit.
Objective 1: Assays of non-bearing trees indicate that soil drench is an effective and consistent application method for increasing copper status of young leaves well above the baseline concentration. The preventative and curative activity of the copper bactericides are being followed in pre- and early stage HLB infected trees. In a trial located at Picos Farm in USDA-Ft. Pierce, Hamlin on Swingle citrumelo trees were root drenched with the copper bactericides: EXP, Cop-R-Quik, (CQ) Magna-Bon (MB), copper phosphite (CP) and zinc phosphite (ZP). In two other locations in ridge groves, healthy pre-bearing trees were drenched with copper chelates (CQ and MB). All three locations were assayed for HLB status this past winter. Thus far, insufficient HLB positive trees have been detected in the non-treated checks to measure treatment effects. A trial in a block of 4-yr old Valencia trees with symptomatic and pre-symptomatic HLB infection as determined by PCR status, no effect of MB or CQ on HLB decline rating was detected in December 2012. Samples for PCR status of the trees in all trials will be collected and analyzed this fall. In the meantime, grower trials of MB and CQ are underway based promising visual observations of improved tree health after 18 months of soil applications of rates varying from 1-2 gal product per application and 2-3 applications per season. One location will provide the opportunity to compare fruit drop in matched 3 acre sub-blocks with and without soil applications of CQ for the last seasons.
Florida growers have reported that enhanced nutritional programs (ENPs) maintain productivity of HLB-infected trees. However, efficacy and sustainability of the nutritional approach for HLB disease management remains uncertain. Complementary studies of multiple ENPs and their individual components compared to the standard nutritional program (SNP) on nursery and field trees were initiated in 2010. Two independent nursery trials were initiated with final data collection finished and data analysis currently underway. From monitoring temporal bacterial movement and replication it was found that Candidatus Liberibacter asiaticus (Las) populations are similar for ENPs and the SNP. Minor differences in Las movement have been observed. Las invaded new flush tissue faster in ENP treated trees than SNP trees. Phosphite treatments have caused Las to favor early invasion of root tissue compared to other treatments. A reduction in visual symptom expression was observed with ENPs. However, there were no significant differences in the HLB-associated phloem plugging or reduction in leaf number, leaf size, and fibrous root mass. There was a consistent trend that the enhanced nutrition actually leads to a reduction in both root and canopy biomass. This suggests that while ENPs improve the appearance of trees by reducing the amount of blotchy mottle and nutrient deficiency associated yellowing, this change in visual appearance is masking the loss of biomass in these trees, which is more difficult to quantify with visual observations. Prior to the trial, field trees had been maintained with good horticultural practices using leaf testing to guide application of foliar nutrients to maintain optimal nutrition. In the field trial, ENPs did slow the development of HLB-induced nutrient deficiency in leaf tissue, but did not prevent the development of deficiency. However, this difference is not reflected in a difference in yield, disease incidence, or disease severity compared to trees in treatments lacking the micronutrient treatments. This supports the growing quantity of data suggesting that nutrient deficiencies are a secondary symptom of HLB and not responsible for HLB-induced yield loss. Combined with results from the greenhouse trial, this suggests that ENPs are masking visual symptoms rather than improving tree health.
Spring/summer 2013: we monitored 8 trials of leafminer attraction to pheromone blends, movement and suppression of trap catch where pheromones were deployed in collaboration with ISCA Technologies and grove managers. These trials address pheromone carrier, timing, deployment pattern and efficacy of mating disruption and leaf damage. Trial 1: Winter and spring application of pheromone (2013). Cooperator: Packers of Indian River. We continued a trial to test application of pheromone in winter and spring on suppression of CLM using a factorial design with split plots in four replicated blocks. Rubber dispensers loaded with triene (834 mg/ha) were treated in winter (Feb 6-8) and spring (Apr 24). Trap catch disruption in treated areas (winter + spring) has remained >95% at 20 wks after application. Flush shoots were evaluated a third time on 5 June. There were no differences in infestation between treated and untreated plots. Infestation severity was about 1-6 mines per flush shoot. Trials 2-3: Rubber dispensers (St. Lucie Co.). Cooperator: Golden River Fruit Co. We finished monitoring trap catch disruption in small plots (0.14 ha) treated with rubber dispensers (318 per ha) loaded with (1) a ‘natural’ 3:1 blend and (2) triene only. Trap catch disruption declined to about 32-41% at 49 wks after application. The equivalence of the two treatments suggests that both work by the same mechanism of non-competitive disruption. Near the same site, 56 ha were treated with a new rubber dispenser loaded with triene (2.5 mg) on 22-24 April, and these proved inferior for trap catch disruption compared to previous forms of the rubber dispenser. The site will be retreated in July 2013. Trial 4: Large plot rubber dispensers (St. Lucie Co.). Cooperator: Packers of Indian River. On 7 Sept an experiment was started to compare trap catch disruption in plots (0.87 ha) treated with rubber dispensers containing ‘natural’ 3:1 blend (330 dispensers/ha) versus untreated plots (0.87 ha). Plots were four times longer than they were wide. Trap catch disruption was 94% at 44 weeks (16 July). Evaluation of leafminers (11-12 June) showed no differences in incidence or severity of leafminers between treated and untreated plots. Trial 5: Large plot rubber dispensers (Charlotte Co.). Cooperator: TRB Groves. We monitoring a grower validation trial treated on 20-23 Aug (42 ha) with a deployment of rubber dispensers loaded with pheromone (natural blend). Modified rubber dispensers were deployed in April; however, this structure of rubber dispenser was found to be inferior for trap catch disruption. Therefore, another treatment of rubber dispensers is planned for July. Trial 6-7: Movement of citrus leafminer (St. Lucie Co.). Cooperator: Blue Goose. We continued a new experiment to test how far P. citrella flies to traps baited with pheromone and placed in a grassland pasture adjacent to a citrus grove. Traps were placed at 75, 150, 300, 600, and 1,200 meters along a transect west of a citrus grove that serves as a source for P. citrella. We captured moths at all these distances. We also set up potted citrus plants as sentinel plants along similar transects adjacent to the source and found that plants were infested at 1,200 m by 6 weeks after placement. Trial 8: Attraction of P. citrella and other Phyllocnistis species to ternary and binary pheromone blends (St. Lucie Co.). Several Phyllocnistis species are attracted to lures containing a binary pheromone blend optimized to catch P. citrella. We started an experiment on 19 April to determine if a ternary blend is as attractive to non-target Phyllocnistis species. Populations of non-target species are low; no differences in trap catch has been observed.
Using gas chromatograph-coupled electroantennography, we have discovered six compounds that elicit antennal responses from the Asian citrus psyllid. Two compounds elicited consistent results from both male and female antennae: neryl acetate and linalyl acetate. Less consistent results were obtained when beta-caryophyllene or geranyl acetate was puffed over the antenna of either males or females. We confirmed antennal responses to formic and acetic acids as degradation products from either citral (a racemic mixture of neral and geranial) or ‘-ocimene (a racemic mixture of cis and trans). Ocimene yielded relatively more acetic than formic acid; citral degradation resulted in more formic than acetic acid as determined by GC-MS. Female antennae responded more (n = 5) to acetic acid compared with males. No difference was detected in male and female responses to formic acid. Results suggest that formic is more stimulatory than acetic acid. These six compounds are now being tested in laboratory bioassays to determine their effect on Psyllid behavior.
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