The goal of this project is to identify insecticidal or antimicrobial peptides with efficacy against phloem-feeding citrus pests, including the Asian citrus psyllid (ACP). Previously, we reported that citrus plants expressing Peptide B showed promise in terms of deterring feeding by ACP, but that only four plants had been evaluated at the time of reporting. During this reporting cycle, we have completed additional replicates that corroborate our previous findings. Additionally, the final peptide to be tested, Peptide D, was evaluated in detached leaf assays and it was found to be the most efficacious out of the four peptides at significantly deterring feeding by adult ACP. Our results also showed that by 48 hours, the feeding behavior of ACP had returned to normal levels when insects fed on Peptide A, while Peptides B and D continued to deter feeding through 72 hours. Future experiments will examine protective effects beyond the 72 hour time point with the most promising peptides. In a preliminary study, we observed that when Asian citrus psyllids harboring Candidatus Liberibacter asiaticus (Las) at rates between 20-80% were held on plants expressing Peptides A, B or C, there was a reduction in the percentage of Las-positive insects after 15 and 30 days post-exposure to the plants, as well as, a reduction in the percentage of infected offspring. Plants necessary to set up a fully replicated experiment to investigate the potential phenomenon of peptide-conferred protection against Las have been grafted with CTV-peptide constructs, and once they have been screened and test positive for CTV expression, assays will be started to investigate these findings. Meanwhile, we have developed quantitative PCR primers for three of the four peptides under investigation in order to correlated psyllid responses to peptide expression in planta. This information will provide important information as to the potential for consistent management of Las transmission by ACP with the most promising peptides.
Biorational, non-neurotoxic insecticides play an important role in integrated pest management given that they typically target a narrower range of species as compared to conventional, broad-spectrum, neurotoxic insecticides. Also, non-neurotoxic insecticides often cause lower levels of toxicity to beneficial arthropods, including parasitic wasps and ladybird beetles, than conventional insecticides. During this report cycle, we have begun evaluation of three new compounds against Asian citrus psyllid (ACP) that fall under the non-neurotoxic category: novaluron, concanavalin A (jack bean lectin), and L-canavanine. Initial experiments with the chitin biosynthesis inhibitor, novaluron, indicate that this insecticide may hold potential as a control agent against ACP by inducing sub-lethal effects on the insect, including reduced egg hatch and reduced adult emergence. Based on these initial studies, Rimon’, the commercial blend of novaluron, is being evaluated at three field rates recommended by Chemtura for testing: 10, 15 and 20 fl oz. These rates are based on efficacy against other Hemipteran species in other fruit crops and on our initial experiments. This insecticide will be evaluated to determine the effect on egg hatch, adult emergence, and fecundity at these three rates. The other compounds under current evaluation, concanavalin A and L-canavanine, are being evaluated against adult ACP to determine feeding deterrence and effects on ACP mortality. Effects on feeding deterrence and mortality have been observed for other related insect species. Initial experiments suggest that L-canavanine, a non-proteinogenic amino acid, may deter feeding of ACP on citrus. However, concanavalin A had no effect on feeding or mortality of ACP in our initial experiment. However, the initial screens for both of these compounds were conducted over a brief time period (48H) and will be extended in subsequent experiments, where mortality effects may be more apparent for both compounds. Efforts to refine an artificial feeding assay to better evaluate these particular compounds are underway given that the current results caused higher than expected mortality in control treatments at 48 hours and longer.
The objective of this investigation is to determine how enhanced nutrition of citrus plants may affect Asian citrus psyllid (ACP) biology, including host acceptance behaviors and population dynamics. We have have been conducting this investigation with complementary field and laboratory experiments. Regarding field experiments, we are still following the Keyplex’ program in the field. The most recent spray of nutritional supplements occurred on February 28th. The next application (post bloom application) will be performed mid-April. Since March, we have monitored the number of psyllids on each tree within our treatment plots weekly with the tap sampling method. We plan to continue this survey during the summer season with regular nymph counts and leaf nutrient analyses. We are also performing a qPCR analysis of all trees in the experiment, 6 months after the initial one, to investigate whether the infection rate has changed, depending on whether or not plots were treated with supplemental nutrients. Currently, our data indicate that infected trees are more attractive to ACP than non-infected trees. This has been already reported from our previous laboratory experiments, but we are observing this effect in the field for the first time. The effects of the supplemental nutrient regime on psyllid population densities are currently not yet statistically significant, but we have observed a trend of more psyllids found in plots treated with nutritional supplements as compared with untreated plots. However, in the laboratory we consistently observe that psyllids are more attracted to HLB-infected plants treated with a nutritional supplement as compared with non-supplemented controls. Interestingly, we have also observed that HLB-infected citrus plants were proportionally more infested with ACP adults in choice settling assays as Ct values (measured by quantitative real time PCR showing presumably higher bacterial concentration) were proportionally lower. We plan to perform a calibration curve for our qPCR method in order to determine the amount of bacterial DNA within these citrus plant treatment samples to confirm whether degree of bacterial infection impacts psyllid preference for choosing and feeding on plants. Our next objectives are to: (1) investigate the acquisition rate of the HLB-causing pathogen by psyllids from plants treated with supplemental nutrients as compared with controls, and (2) collect volatile chemicals from plants before and after the application of a nutrient spray cycle, in order to determine how this treatment affects the volatile emission pattens from plants. Our hypothesis is that changes in volatile emissions may be affecting behavior of ACP.
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. 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. This permit clearly notes experimental protocols and procedures so that the work is done as safely as possible to minimize the chance of ACP escape. At this site, Jim Bethke has initiated ACP tests on various organic and traditional pesticides of interest to California growers using 4 types of tests: adult topical adult drench adult residual and nymph topical tests. Data have been gathered to date on 14 traditional pesticides and 12 organic products, in many cases with multiple trials involving each material. 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 (18 trials to date) in comparison with studies done in Florida. Two 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. Kris Godfrey obtained a permit to rear ACP inside UC Davis’ Contained Research Facility and received 4 shipments of ACP (under CDFA permit) from Bethke’s colony to start the Davis colony (so that Chula Vista and Davis results might be similar). She has initiated testing of organic products and has several other lines of experimentation planned based on requests from the CRB. Beth Grafton-Cardwell has updated online ACP pest management guidelines and has given 8 presentations to various groups dealing with ACP since 10-1-12 (i.e. over the past 6 months). She has served as a focal point for communicating with ongoing ACP management programs in the SJV, Ventura, and elsewhere. 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).
Our project goal is to develop a system for inoculating Asian citrus psyllid (ACP) with a native pathogenic fungus, Isaria fumosorosea (Ifr), and use these infected psyllids as ‘autodisseminators’ to effectively spread the pathogen among ACP populations in residential citrus. Research goals for 2012-2013: (1) Conduct field trials to determine if Ifr-inoculated ACP adults can infect nymphs in residential citrus, (2) Determine seasonal and environmental effects on infection levels, (3) Evaluate pathogen dispersal, (4) Assess pathogen effects on arthropod biological control agents. Main Findings: “Infection of ACP in dooryard citrus”. A field trial was conducted during April to May of 2012 with Mexican lime trees at the ‘Victoria Palms’ RV park in Donna, TX. Ten adult ACP were dusted with Ifr inoculum and released into mesh bags placed around shoots with nymph clusters. Each test tree contained one to four clusters; a total of 15 clusters on 10 trees were tested. A separate set of nymph clusters (n = 15) measured ambient levels of Ifr infection. In the control clusters, we found few dead individuals and no Ifr-infected psyllids. In clusters exposed to dusted adults, a mean of 39% of the psyllids were infected. For a follow-up trial at ‘Victoria Palms’ during July to August of 2012, 100 ACP adults were dusted and released on the north-west and south-east canopies of Mexican lime trees (n = 4) and grapefruit trees (n = 8). Identical numbers of ‘control’ trees were used to measure ambient levels of Ifr infection. After 3 weeks, we inspected two nymph clusters from both sides of each tree and found no Ifr-infected individuals on control trees, no significant effect of canopy aspect on infection levels, a mean of 16% infected individuals on lime trees and 6% on grapefruit trees. These trials demonstrated that dusted psyllids could infect nymphs in open clusters on residential trees under summer weather conditions in Texas. “Susceptibility of immature Tamarixia radiata to Ifr spores”. During November 2012 to February 2013, we evaluated susceptibility of Tamarixia radiata, a parasitoid wasp used for biological control of ACP, to Ifr spores before and after ACP mummification. To evaluate susceptibility of T. radiata prior to ACP mummification, we exposed ACP nymphs to adult female wasps for 24 h and waited 96 h before confirming parasitism of individual nymphs. Each parasitized nymph was transferred to a citrus leaf disk and then dusted with Ifr inoculum (n = 20). A separate set of parasitized nymphs was transferred to leaf disks but not exposed to Ifr (n = 20). Wasp eclosion was 14 times greater among control nymphs than Ifr-dusted nymphs. To evaluate susceptibility of T. radiata after ACP mummification, we transferred parasitized ACP nymphs to leaf disks but waited until mummification (approximately 168 h after parasitism) before dusting half of the mummies with Ifr inoculum (n = 17) and leaving the other mummies as controls (n = 17). Wasp eclosion was 48% among mummies dusted with Ifr and 65% among controls. These trials demonstrated that susceptibility of immature T. radiata to Ifr spores is greatly reduced after ACP mummification. “Susceptibility of immature Tamarixia radiata to Ifr hyphae”. During March 2013, we evaluated the susceptibility of T. radiata to Ifr hyphae after ACP mummification. We allowed parasitized ACP nymphs to mummify on leaf disks before exposing half of the mummies to Ifr hyphae (n = 23) and leaving the other mummies as controls (n = 23). Wasp eclosion was 17% among mummies exposed to Ifr hyphae and 78% among controls. These trials demonstrated that immature T. radiata are susceptible to Ifr hyphae after ACP mummification.
This project began 11/1/2012 and our efforts are now ongoing. We have colonies of both psyllids (Bactericerca cockerelli and Diaphorina citri) to be used in our work and are using both for our studies here. We have created a series of transgenic plants to compare general tissue (35S promoter) and specific phloem (AtSus2) expression of anti-psyllid (B. cockerelli initially) sequences for their potential to induce RNAi effects on psyllids. We have also begun constructing artificial mirco RNA interfering RNAs as a strategy to deliver specific anti-psyllid RNAs to psyllid targets. Three 21-nt-sequences of the Bactericerca cockerelli ATPase (BC-ATPase) gene were used to construct artificial microRNAs (amiRNAs). MIR319a has been used as a backbone of amiRNAs and therefore is chosen for our amiRNA constructions. The sense and complementary strands of three 21-nt-sequences were generated. One of the three 21-nt-sequences was also created using primer extension to have a amiRNA with shorter stem-loop. Therefore, the total of 7 amiRNAs targeting BC-ATPase were made and cloned into pBluescript SK, which are pAMIRA1, pAMIRA1c, pAMIRA2, pAMIRA2c, pAMIRA3, pAMIRA3c, and pAMIRA2PE. These constructs, plus appropriate controls will be used 1, for in vitro transcription and the RNA transcripts will be used for psyllid-feeding experiments; 2, to construct into a which can generate the amiRNAs in plants, and 3, to clone into a binary vector for transient expression by agro-infiltration. We will begin evaluations for effects very soon and will utilize positive results to move towards targeting D. citri. We are also attempting to identify the actual forms of RNAi-inducing RNAs that are present in plants. To determine if the double-stranded RNA induced siRNAs can move systemically in plants through the phloem for psyllids to acquire, the full-length BC-ATPase sequence was inserted into an agroinfection-compatible TMV expression vector. The BC-ATPase TMV will be infiltrated into Nicotiana benthamiana plants and the newly emergent leaves will be used to detect for siRNAs of BC-ATPase by Northern blot analysis. We will also perform bioassays on these leaves to determine whether or not RNAi effects can be induced in recipient psyllids.
We have made further progress on nucleotide sequence and bioinformatics analyses of Diaphorina citri samples from several worldwide locations in order to identify candidate D. citri-infecting viruses. We are continuing to sequence and characterize the D. citri reovirus that we have already identified from Taiwan samples. We have re-constructed a new small RNA library for the Taiwan D. citri RNA samples and we hope that we can generate additional contigs this time in order to identify the full genome sequence of the reovirus, but also we hope that deeper sequence analysis may identify additional viruses as the RNA quality of these samples was excellent. This virus is confirmed also to be in psyllids from China, but not from Brazil. It is likely present in Florida D. citri samples due to previous reports, but we have failed to confirm these at this time. We have also now generated small RNA libraries for Diaphorina citri RNA samples from China and Brazil. Both of these RNAs showed good quality with RIN (RNA integrity number) ‘ 8. After validating cDNA constructs, we submitted these two libraries to BGI (Beijing Genome Institute) sequence facility in Hong Kong for deep sequencing in Illumina`s Solexa platform with HiSeq system. We are waiting for these sequences to start data analysis. We also have received five new RNA samples from Florida and samples from Pakistan. After analyzing RNA quality, three of the Florida samples were selected for small RNA library construction and deep sequencing. Right now, we are generating these libraries and they will be sent for sequencing soon. We have now established a virus-free D. citri colony within the UC Davis Contained Research Facility. We will use these psyllids for biological assays as D. citri viruses are discovered and prove to be workable.
The overall goal of this 3-year research project is to efficiently deliver antimicrobial molecules into citrus phloem against the HLB bacterium. This quarterly (from Jan 2013 to April 2013) research continued to optimize the W/O (water in oil) and O/W (oil in water) nanoemulsion formulations by evaluating the ideal adjuvant mixtures of solvents, oils, lipophilic and hydrophilic surfactants using pseudo-ternary phase diagrams and thermodynamic stability methods. Several W/O and O/W nanoemulsion formulations were screened for further preparation of optimum W/O or O/W solutions using pseudo-ternary phase diagrams with oil as the oil phase, water as the aqueous phase, and combinations of lipophilic surfactant, hydrophilic surfactant and water-miscible solvent as mixed phase. We have developed a method for screening penetrants on citrus and used this to evaluate and quantitate several penetrants on different cuticle types. This will used to select the best penetrants for further application. First round tests on LIberibacter control have been conducted on seven compounds from the Citrus Research and Education Center, IFAS, UF and one compound, epsilon-L-polyline, from China. The second round tests will be done in the next quarter. The future work will be focused on the following: 1) evaluating penetration ability of chemical compound and penetrants through the citrus cuticle; 2) Evaluating drug loading capacity using the optimized nanoemulsion formulations; 3) Optimizing the final formulations by combination of screened penetrants and high drug loading capacity formulations.
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 quarter of 2013. By the end of the previous quarter, we had programmed an Arduino Uno microcomputer system to detect and record psyllid-produced vibrations from a microphone attached to a citrus tree branch. We now have 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. In the last two weeks, we have begun the process of incorporating the microphone-buzzer microcomputer-controlled system into a trap for testing in field environments. Earlier in the quarter, we also presented some of the initial findings at the 3rd International Research Conference on Huanglongbing in Orlando, FL. One paper on this research is in press, and two others are in progress.
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. A bioassay was developed using a wind tunnel to determine ACP response to different wind induced stimuli. ACP are marked with fluorescent powders for easy observation and released into the wind tunnel for different evaluations. Objective 2. Stelinski’s lab initiated an experiment to study the effect of adult density (crowding) on ACP dispersal behavior. For two days, psyllids are allowed to settle on Valencia plant citrus plants in the laboratory. Densities of psyllid released varied from 1 ACP per cm of leaf flush to 10 ACP per cm (1:1 sex ratio). After two days, four new Valencia plants were introduced into each cage. Each day, over the course of one week, psyllids that dispersed from the original plant to the newly introduced plants were removed and sexed. An increase in the number of dispersing ACP was observed with increased psyllid density. Females were also observed to be more sensitive to ACP density than males, and consequently dispersed sooner than males at lower crowding levels. Currently, they are conducting experiments that vary the sex ratio of the ACP released in the crowding experiments. Based on previous results showing that females repel one another at high densities of crowding, it is hypothesize that for a given psyllid density, increasing the proportion a females should increase female tendency to disperse. Experiments were also initiated to investigate how infection of psyllids with Candidatus Liberibacter asiaticus (Las) affects dispersal behavior. Using a flight mill technique, it was seen that a small portion of psyllids initiate long flights, while most appear to only fly for short periods. Why do some psyllids initiate these potentially long-range flights, while others do not? Sex, mating status, level of satiation, presence of endosymbionts, and infection with the Las pathogen are all being investigated. Understanding why a small portion of psyllids undertake long range flight may provide a means of targeting long-range movers. Objective 3. Release of up to 2,000 adult ACP marked with fluorescent dye has resulted in too little recovery to draw significant conclusions so plans underway for another mark recapture trial using more adults. Objective 4. Yellow sticky traps set at three canopy heights are giving mixed results with more ACP captures in the upper canopy at in the older Charleston grove and a more even distribution at the younger and more uniform Bob Paul grove. However, traps on the east side of the trees capture more than the west side at both locations. At both locations and also the English grove, ACP captures are greatest adjacent to a hammock or wind break. Also at Bob Paul, traps set up to assess ACP movement between insecticide sprayed and unsprayed blocks with low and high populations of ACP respectively indicate no significant directional effects. Objective 5. A paper reporting positive results using UV reflective mulch to repel ACP the first 2 years after planting was accepted for the refereed journal Pest Management Science. Subsequent results indicate that the 5 ft strip of metalized mulch is not effective on trees more than 5 ft tall. In addition the mulch has shown the ability to sigificantly slow rate of HLB spread even under high pressure. Trials to test different ways of utilizing UV reflectance such as wrapping canopies with strips of metalized mulch have commenced.
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. A bioassay was developed using a wind tunnel to determine ACP response to different wind induced stimuli. ACP are marked with fluorescent powders for easy observation and released into the wind tunnel for different evaluations. In recent tests, ACP have not been responding in the wind tunnel (as previously) and photoperiod is being investigated as a possible cause. Objective 2. In the previous report, we described experiments indicating that female psyllids avoid plants that were previously infested with a high density of conspecific females, and that they are also repelled by conspecific female odors. We determined if ACP damage to plants also affects psyllid movement. We used potted citrus plants that were sectioned into two halves with Teflon guillotines. Sectioning the plants into two halves allowed us to infest only one portion of the plant and simultaneously measure psyllid movement to the non-infested portion. After 24 hr of psyllid feeding on one section of the plant, other psyllids were much more attracted to the non-infested portion of damaged plants as compared with non-damaged controls. These results indicate that plant response does not contribute to psyllid-psyllid repellency, but in contrast, there is a systemic plant response to attract other psyllids upon damage. GC-MS analysis showed that this attractiveness is likely due to increased production of methyl salicylate from damaged plants. Our current hypothesis is that specific volatiles from citrus are important for selection of trees that are already infested with psyllids (indicating a favorable host), and female odor is used by other females to avoid flush that have been over-utilized. Both signals impact psyllid movement in opposite ways. Objective 3. Preparations are underway to release more marked psyllids this year after too few were recovered last spring in the first attempt. Objective 4. Captures of ACP on yellow sticky traps set at three canopy heights are giving mixed results, depending on tree height. More ACP were captured in the upper canopy in an older mature grove. However, a more even distribution of psyllids throughout the tree canopy was observed in a younger grove. However, more ACP are captured on the east side of the trees than the west side at both locations. At all locations sampled, ACP captures are greatest adjacent to a hammock or wind break. Also, traps set up to assess ACP movement between insecticide sprayed and unsprayed blocks with low and high populations of ACP, respectively, indicate no significant differences in directional movement. Seasonal fluctuations are likely and further analysis of the data is pending. Objective 5. Results indicate that the 5 ft strip of metalized mulch is effective in repelling ACP until it is shaded out by the canopy when trees exceed 5 ft in height. However, the mulch treatment reduced incidence of HLB infection by 75% for almost 2 years. A paper reporting positive results using UV reflective mulch to repel ACP for 2 years after planting was submitted to and is now under revision for the refereed journal, Pest Management Science. Also, an article was submitted to Citrus Industry for publication in the Apr 2013 issue. Trials have begun to evaluate metalized mulch placed around 1 year old resets or adjacent to mature trees. Also metalized ribbon hung in the tree canopy is being evaluated.
March 31, 2013 Update: Activities of the CHMA program assistant (Brandon Page) Jan 1 – March 31 2013: During this reporting period, the CHMA assistant provided support to the CHMA effort by attending various local CHMA meetings as well as statewide meetings. Specifically, Mr. Page attended 14 meetings where he either gave an oral presentation, setup CHMA displays such as posters, or participated in group discussions. Venues included Citrus show in Ft. Pierce, ConservII management meeting, IRCHLB in Orlando, South Lake / West Orange CHMA meeting, Gulf CHMA meeting (2x), Florida Southern College, Futch’s Citrus Production school, Mid-Florida Citrus Foundation Field Day, Hardee CHMA meeting, IRCHLB grower day at CREC, Ft. Meade ALturas CHMA meeting, North and Central Polk Co CHMAs meeting, and the Volusia CHMA meeting. In addition to attending meetings, daily updates were made to the CHMA website based on communication with CHMA grower leaders. Work also continued on the mapping program that went live online at the end of December 2012. A considerable amount of time has also been spent working out the bugs in the program and working with the programmers to enhance the program with new features not yet available online.
March 2013 update: Field trials conducted to determine the true longevity of pesticide applications were completed this spring. The goal of the project was to use caged psyllids on treated trees to assess how long pesticide residues remain active on the foliage of trees killing adult psyllid fly-ins. This is in contrast to the manner we normally evaluate pesticide products by spraying out products and evaluating how long psyllid populations remain below the untreated control. Such an evaluation really is only a measure of how long it takes psyllids to reinfest from surrounding areas and thus can be influenced by the surrounding psyllid population levels. In this study, we examined three pesticide active ingredients, dimethoate, Danitol (fenpropathrin) and Admire (imidacloprid applied foliarly). These products were applied at label rates during the months of January, April, September and November. The products were applied during these months to represent weather extremes at different times of the year when growers are making their pesticide applications. On each application date, 24 hrs after sprays were applied, adult psyllids were caged on the treated trees. After 7 days had passed, the cages were removed to assess psyllid mortality and new cages containing alive psyllids were then cages back on the same tree in order to continue the evaluations of duration of pesticidal efficacy. The results showed that 1 day after application, all pesticide residues on leaves provided near 100% control of adult psyllids with one exception. In september, high rainfall occurred approximately 12 hrs after the treatments were applied resulting in less than 60% control 24 hrs after the applications were made. This shows that despite the fact that the products had time to dry after applied, rains that are typical during the florida rainy season can negatively influence pesticide effectiveness. By 7 days after application, percent mortality across all treatments was less than 60% in most instances with even greater reductions observed following periods of heavy rain. Time of year did not have an impact on pesticide efficacy, dispelling the notion that pyrethroids work better in the winter and organophosphates provide better control in the summer. These results suggest that it is more important to choose a product for psyllid control based on rotation from previous MOA used rather than trying to use only pyrethroids in the winter and OPs in the summer. Additionally, these results show that growers are not likely to obtain as long a duration of psyllid control as they previously thought possible. This is likely best overcome through coordination of sprays through the CHMA program.
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 look good and actually improve, despite the nearly 100% HLB incidence. Foliation of canopies is dense, and leaf color going into late Fall season was a healthy green. Tree sizes (heights and total canopy volumes) were measured and analyzed on 1 November 2013, as follows: FOLIAR FERT Tree Ht(in) STD-Ca STD+Ca Prescrip SOIL FERT Liq/Dry 89.6 87.0 93.8 Liq 89.4 82.5 97.0 Liq/Dry-Ca 90.0 86.6 86.3 FOLIAR FERT Canopy vol(ft3) STD-Ca STD+Ca Prescrip SOIL FERT Liq/Dry 287 239 306 Liq 268 207 324 Liq/Dry-Ca 286 242 237 Both the tree height and canopy volume means showed non-significant (NS) differences between treatments, which agrees with the visual observations made in the grove. Leaf samples were collected in December 10, 2013 and the results of nutrient analysis of the leaf tissue were NS except for B and S. All the different fertilizer strategies chosen for testing in this block appear to be working equally well. The slightly adverse effects of the +Ca foliar sprays on canopy size and fruit yield in 2012/13 appear to be corrected in 2013/14.
Entomopathogenic nematodes: Amending soils to increase biological control of insect pests Processed the autumn soil samples from Schumann ACPS trial in Auburndale and new trial in Bartow in which sulfur is a treatment to reduce pH from near neutral to near 5.0 and determine effect on Paenibacillus sp. infestation of Steinernema diaprepresi and on the population density of the nematode. Briefly, both trials revealed remarkable similarity in quantitative relationships (regression equation) between population density of the bacterium and the nematode. Relationship is significantly positive, but infestation rate (bacteria spores per nematode) decreases with population density of the nematode. Across 4 treatments and 40 plots, numbers of S. diaprepesi are inversely correlated with soil pH (P<0.001), supporting the relationship measured in the Auburndale trial. However, the numbers of nematode in plots treated with sulfur do not yet differ from those in other treatments. If numbers do change over time it will be the first known example of a specific agronomic practice that can be exploited to conserve services provided by entomopathogenic nematodes. Wrote 'New citriculture system suppresses entomopathogenic nematodes and biological control of subterranean herbivores' Campos-Herrera, R., Pathak, E., El-Borai, F., Schumann, A., Abd-Elgawad, M.M., Duncan, L.W. for publication in Biological Control. Plant parasitic Nematodes: Characterizing a new nematode pest and the prevalence of resistance breaking populations of the citrus nematode. Continued sampling and processing citrus soil from Indian River, St. Lucie counties. Identified sites for chemical trials to begin in February 2013. Used newly developed PCR primers and Taqman probes to confirm identities of 4 populations of putative X. citricolum and X. laevistriatum. All positive for latter species, none for former. This raises questions about validity of these species. Sampled 4 sites identified in original publication as sources for these populations. Dagger nematodes from these sites are being characterized for morphology and ITS and D2/D3 region DNA sequences. These samples should be adequate to validate or invalidate the current taxonomy in this group of citrus parasites.
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