We have previously shown that dsRNA identitical to two separate Asian citrus psyllid genes (Cathepsin F and Vacuolar ATPase) are toxic when fed to the psyllid in an artificial diet. As a result of this finding, we have worked in collaboration with Dr. Bill Dawson and Dr. Siddarame Gowda to express these dsRNAs in citrus using their infectious Citrus Tristeza Virus (CTV) vector. Such plants have been developed and are now being evaluated for the presence of psyllid gene dsRNA in the phloem tissues. These plants are also being propagated to challenge with psyllids to analyze psyllid preference and performance with respect to mortality and efficacy when either provided with these plants as the sole host plants or provided with a choice of these dsRNA expressing plants and non-dsRNA expressing plants.
This is the second annual report for a three year project on protecting newly planted citrus trees from Asian citrus psyllid (ACP) and huanglongbing (HLB). Key information from the project is summarized here. In an area under high ACP and HLB pressure, regular treatments of insecticides may reduce ACP levels but outbreaks still occurred even under an intensive insecticide program (monthly applications of traditional insecticides). Repetitive (every three weeks) applications of petroleum oil failed to prevent periodic ACP outbreaks. There was little evidence that interplanting orange jasmine with citrus reduced ACP infestations in citrus, regardless of whether the jasmine was treated with pesticides or not – however, problems establishing and maintaining jasmine plants was difficult, thus definite conclusions regarding the value of jasmine as a trap plant could not be made. Regardless of the specific insecticide program evaluated and degree of ACP control achieved, none of the programs entirely circumvented HLB infections. In experiments where infected trees were removed as they were identified, less than 10% trees tested positive for HLB by 12 months after planting. In a test where infected trees were not removed, the incidence of HLB in young citrus generally increased faster with up to 25% trees testing positive within a year. In the experiments where infected trees were removed as they were found and not replaced, 40 to 50% trees had been removed by 33 months after planting and some plots began to look barren. Had the infected trees been replaced with resets, it was likely that resets would have developed the disease at a rate equal to the older young trees and it is doubtful that enough trees would have ever began producing enough fruit to offset planting expenditures. In tests where trees were not removed, at 14 months after planting percentages of trees testing positive for HLB (Ct<32) reached 35 to 50% but then declined to as low as around 15% at 19 months after planting. In experiments where infected trees were not removed, some trees (both healthy and infected) began producing fruit during the second year after planting. During June at 22 months after planting, although infected trees were shorter, they were equivalent to healthy trees with respect to trunk diameter and number of fruit; whether the fruit of infected trees will remain on trees until harvest remains to be determined. In one planting, an average of 5% trees died by 17 months after planting but only about 1% of dead trees were known to be HLB-infected. Among other dead trees, some were killed by freezes and some by Phytophthora. Although HLB may weaken a tree and predispose it to other problems such as Phytophthora, at 22 months after planting there was the same percentage of trees infected by Phytophthora among trees infected by HLB (10.8%) and trees not infected by HLB (10.3%). During the third year of this project, we will continue subjecting young citrus to the various insecticide programs and monitoring the young plantings for HLB. Growth and yield data will be obtained for healthy and HLB-infected trees. Juice quality will be assessed for healthy and diseased trees. The epidemiology of HLB in citrus under different insecticide programs will be investigated. Collectively, the results of these experiments will provide growers insight into the efficacy of different ACP control programs for protecting young citrus from HLB and, if infected trees are not removed, what they may expect from a new young citrus planting with respect to tree growth, survival and fruit production.
A series of research greenhouse research experiments have been completed that show that a number of commercial formulations of plant growth regulators (PGRs) (GA biosynthesis inhibitors) reduce Asian citrus psyllid fitness. Specifically, we found that two of the PGRs tested (prohexadione calcium and mefluidide) reduced ACP fecundity and survivorship while two others, uniconazole and paclobutrazol, reduced fecundity or survivorship, respectively, when psyllids were caged on treated trees 10 days after PGR application. These effects on psyllid biology are independent of vegetative growth, i.e. they occur at PGR rates below which vegetative growth is reduced, and are apparently not a result of PGR toxicity to the psyllids. These results have been summarized in a manuscript currently in review with the Journal of the American Society for Horticultural Science that we anticipate will be published in late 2011. Follow up work has been ongoing to understand the plant biochemical changes occurring in response to PGR treatment that may be correlated with changes in psyllid biology. We have found that plants treated with prohexadione calcium exhibit significant changes in a number of amino acids between 9 and 15 days after PGR application. At least three of these amino acids are considered to be essential dietary nutrients for plant-feeding insects. Analyses of other dietary compounds (carbohydrates, vitamin A and E) are ongoing. We anticipate that these results will be submitted for publicatio in late 2011. One of our proposed objectives is to determine the effect of PGRs on acquisition of Candidatus Liberibacter asiaticus (Las) by the Asian Citrus Psyllid (ACP). Previously, we reported that a population of citrus trees was graft-inoculated with Las-infected budwood for use in experiments. We recently tested for the presence of Las in the grafted plants using quantitative polymerase chain reaction (qPCR) assays and determined that approximately 40% of grafted plants were infected with Las. Subsequently, we have conducted experiments to evaluate the effect the PGR prohexadione calcium on Las acquisition. Healthy and Las-infected plants were treated with prohexadione (sprayed until run-off) and allowed to dry. Twenty adult psyllids of mixed gender were released onto treated plants and enclosed in nylon mesh sleeve cages. As a control, twenty ACP were also released onto healthy and Las-infected plants treated with water + adjuvant. Each treatment was replicated 6 times on two separate dates. Plants and insects were maintained in a temperature-controlled environmental chamber for 28 or 14 days prior to collecting insects. Following each acquisition experiment, qPCR analysis of psyllids revealed that 54% and 51% of psyllids acquired Las from treated plants after 28 and 14 days, respectively, compared with 50% and 40% of psyllids feeding on untreated plants. Although these data suggest that prohexadione does not decrease acquisition, these results are preliminary and may indicate that the residual activity of prohexadione against psyllids does not extend to two weeks. These experiments, which evaluated acquisition after two weeks or one month of feeding, will be replicated two more times. In addition, subsequent experiments will be conducted to determine the rate of acquisition after 24 hours, one week, and during nymph development.
Under Objective 1 (define rates and formulations of copper sprays for more effective control) one trial with Red grapefruit was conducted in Ft. Pierce, FL and one trial with Hamlin orange in Hardee County, each with 14 treatments of formulations, rates and combinations with other chemical treatments of interest. This season, trials had low to moderate disease epidemics as a result of a periodically wet spring, moderately wet summer and an absence of tropical storm events. Evaluations of fruit infection in these trials were made in November 2010 (grapefruit) and January 2011 (Hamlin). Copper formulations containing copper hydroxide or basic copper sulfate (metallic rates of 0.67 to 1.12 kg/ha), varied from low (Hamlin) to moderately effective (grapefruit) for canker control of fruit disease incidence. Copper pentahydrate, at a lower metallic copper rate/ha per application, provided equivalent control to film-forming copper formulations in grapefruit but less control in Hamlin. Greater canker susceptibility of fruit occurred in later season and was likely because of prolonged opening of stomates in cooler weather and enhanced bacterial entry, coincident with more numerous windblown rain events. Although Hamlin fruit disease incidence was higher, copper protection against early season fruit infection was moderately effective for prevention of premature fruit drop. Under Objective 2 (establish the period of fruit susceptibility, residual activity and phytotoxicity of copper). In 2010, we compared the copper fruit residues from different copper-containing products up to 28 days after application to grapefruit. The copper product used and the number of days after application significantly affected the amount of copper residue. These effects were consistent whether the data were analyzed as copper/fruit or copper/surface area. The rate of copper residue decrease was different for the various products tested. Copper products forming films, Kocide 3000 (copper hydroxide), Cuprofix Ultra 40 Disperss (copper sulfate), and Badge X2 (copper hydroxide and oxycloride) decreased at a similar rate, whereas the soluble formulation Magna-Bon CS2005 (copper sulfate pentahydrate) decreased at a higher rate and left about 75% less fruit residue. This was expected as Magna-Bon is applied at approximately 15% of the metallic contained in film-forming coppers. Since Magna-Bon performed as well as film-forming copper products for control of canker on grapefruit the last two seasons, we have hypothesized that the Magna-Bon copper could be locally systemic in fruit rind tissues. This coming season, we will attempt to assay the activity of copper in the rind against Xcc to determine whether the copper is located internally as well as externally. Under Objective 3 ( evaluate the use of streptomycin [Firewall]) As in the past three seasons, applications of Firewall in July and August, were effective for canker control on grapefruit either in combination with a reduced rate of copper or when substituted for copper in the spray program. Based on Firewall’s efficacy and ability to lower the risk of copper phytotoxicity, an application has been submitted by FFVA and FDACS to gain EPA Section 18 emergency registration for use of Firewall against canker on fresh grapefruit. Under Objective 4 (To define risk for development of bacterial resistance to copper and streptomycin in FL citrus groves) a number of factors favorable for the development of copper resistance in Xcc were identified. Findings are being prepared for publication. Under Objective 5 (rapid transfer improved canker management technology to the Florida citrus industry), 2011 canker management recommendations have been submitted for publication in the Florida Citrus Pest Management Guide and to Citrus Industry Magazine. Oral presentations have been delivered at the Florida Citrus Show and at a multi-county extension meeting. Results of the Hamlin trials will be presented at the 2011 FSHS annual meeting.
Objective 1 is to conduct a field evaluation of nutritional sprays for control of HLB and HLB symptom expression and yield. The field study was set up May 2010 in Southern Grove, Hendry Co., FL. Six treatments were located in 4 plots of 150 trees per treatment (interior 10 trees in each block were identified for PCR, leaf nutrition sampling, tree health and yield evaluation). Trees were 8 years old at the initiation of the trial. Treatments were 1) non-treated check; 2) Nutri-Phite sprayed 4 times bimonthly; 3) N-Sure sprayed bimonthly; 4) Agra Sol Mn/Zn/Fe plus Nutri-Phite plus triazone urea sprayed bimonthly; 5) Keyplex 1400 DP plus Nutriphite plus triazone urea sprayed bimonthly; 6) Wettable powder nutrients (Diamond R #2) plus Nutri-Phite P+K sprayed bimonthly. The materials were applied to both sides of the tree in 125 gallons per acre with an airblast sprayer driven at 2 mph to obtain thorough coverage. Five disease ratings have been taken so far with the latest in June 2012 and a slight decline in tree health has been observed, but no significant treatment effects have been detected. The 2011 harvest increased 27% yield in all treatments compared with the 2010 harvest, but no significant treatment effects were observed. Objective 2 is to determine the mechanism of HLB symptom suppression by foliar nutritional application, Rep 1 using Hamlin sweet orange trees inoculated with HLB and treated bimonthly with the nutritional sprays treatments 1, 2, 3, and 5 from objective 1 has finished and results are in previous reports. Rep 2 monthly samplings of leaf and root tissue are continuing. Root samples are split for qPCR Las quantification and starch analysis for a quantitative measurement of phloem function throughout the plant. Microscopy will be continued, however the high variability of phloem plugging and collapse even within the same midrib from a symptomatic leaf makes interpretation of results difficult. 72 of the 75 inoculated trees are now positive with disease symptoms developing. No treatment effects have been observed, except that the phosphite alone treatment results in higher than expected infection of roots within 4 months of inoculation which continues to occur at 6 months post inoculation. Now that symptoms have developed and consistently positive tissue has been identified, samples have been collected for microscopy of leaves and roots. The samples have been fixed and sectioning of samples for microscopy is underway. Some trees have begun to show significant signs of decline, but it is too early to tell if decline is more common in a specific treatment. No significant differences in quantity of flush between treatments or Las titer has been identified other than the initial infection of roots mentioned above.
Exp.1 ‘ Test plant from the first and second trials are being observed for the presence of new symptomatic trees and leaf sampling of suspected plants are being PCR tested. We are waiting for laboratory results. Exp. 2- Emerging ACP adults from nymphs reared on inoculum sources plants were used for inoculation in Mar/10. Until now no symptomatic test plant was observed. Exp. 3 ‘ New inoculation of Las in three shoots of each younger trees of each of three sweet orange varieties on screenhouses was done in July/11. The two older class of plants are being prunned to be inoculated in November/11. Some symptomatic trees that appeared before the inoculation were maintained in the screenhouse and the severity progress was assessed during last year project. For Hamlin/Rangpur 99, the annual severity increased from 24.3 to 27.5% (average of 7 trees); for Valencia/Rangpur 99, from 24.5 to 38.5% (average of 5 trees); and for Hamlin/Rangpur 04, from 35.0 to 57.5% (average of 2 trees). As an alternative for this experiment, 30 trees from each five citrus blocks at different scion/rootstock combinations (Hamlin/Swingle 07, Valencia/Volkamer 07, Valencia/Swingle 07, Valencia/Swingle 99, Valencia/Swingle 95), showing very low symptoms severity, were selected in Jun/10 in farms without removal of symptomatic trees but with strong ACP control program. After 400 days, the disease severity increased in average from 6.2 to 9.7% for Val/Sw 95, from 4.6 to 10.3% for Val/Sw 99, from 7.9 to 14.5% for Val/Sw 07, from 8.6 to 23.5% for Val/Volk 07, and from 10.8 to 9.3% for Ham/Sw 07. For all plots the disease severity increased except for Ham/Sw 07. In that plot many selected trees did not show visual symptoms at last assessment (the presence of Las in those asymptomatic trees will be checked). Also it was observed that during drought season, because of water stress and defoliation, the disease severity assement is more complicated and difficult, usually resulting in lower severity. All assessed tree of Val/Sw 95, Ham/Sw 07 and Val/Volk 07 were harvested in July/11 and the average yield were compared with average yield of healthy trees from the same plot. Yield reduction (%) was higher for diseased trees of younger plots Val/Volk 07 (31.6%) and Ham/Sw 07 (35.2%) than older plot Val/Sw 95 (16.5%). In the plot Val/Sw 99, for a mistake the healthy trees were pruned and the selected trees were not. The plot Val/Sw 07 was not harvested yet.
Leprosis infected samples were recently sent to quarantine facilities at the USDA, APHIS, PPQ, CPHST, Beltsville, MD in order to check on the shipment and the viability of the samples for the mite transmission work to begin as soon as the funding is in place. The materials were successfully shipped from Mr. Guillermo Leon of COPIACA, our cooperator in Colombia. Both protein and nucleic acid extractions were done and PCR verified the presence of the virus in the samples. Dr. Jorge Pena, University of Florida, Homestead, FL has begun preparing colonies of endemic Brevipalpus mites for transmission experiments. Mr.Leon in Colombia already has Brevipalpus colonies and will begin transmission experiments there as soon as funding is in place for him to be able to pretest samples for virus. Drs. Schneider and Damsteegt have prepared space for the transmission work at the USDA, ARS, FDWSRU in Ft. Detrick and as soon as we are able to purchase the isolation chamber for the transmission work we will begin the first transmission experiments.
Under Objective 1 (define rates and formulations of copper sprays for more effective control) and Objective 2 (establish the period of fruit susceptibility, residual activity and phytotoxicity of copper) a three year trial with Hamlin orange in Hardee County, with treatments of formulations, rates and number of sprays was presented at the 2011 FSHS annual meeting and submitted for publication in the proceedings. The results are summarized as follows: The objective was to evaluate copper formulations for control of fruit infection and drop in 6- to 8 year-old ‘Hamlin’ trees. Copper sprays were applied at 21-day intervals after fruit reached 0.5 to 1.0 cm dia. The period of susceptibility of fruit to canker infection and fruit drop was established by increasing the number of applications through the fruit growth period. Separate treatments ended at each 21-day interval so that there were four to seven applications per season. In 2008, early season infection occurred during rains before copper sprays commenced in late April. Subsequently, five sprays of copper formulations at rates exceeding 0.5 kg metallic copper/ha significantly reduced incidence of lesions on fruit. Fruit disease and cumulative fruit drop were co-ranked among copper treatments (r = 0.83). Although a tropical storm in early August promoted disease on fruit late in the season, most of the premature drop was due to infection occurring before July. In 2008 and 2009, sprays after the period of early fruit susceptibility did not further reduce canker incidence or fruit drop. In 2009, copper sprays were initiated before significant spring rainfall and the incidence of fruit disease and fruit drop were lower and less well-correlated among copper treatments (r = 0.57) compared to 2008. In 2010, disease on fruit and premature drop were not significantly different from the untreated checks. Overall, there was little difference in efficacy among copper formulations, although there was reduced control using copper sulfate pentahydrate at lower rates of metallic copper. In each season, copper treatments controlled fruit drop by ~50% compared to the untreated check, however as ‘Hamlin’ trees grew from 6 to 8 years of age, canker incidence dropped due to the development of hedgerows which reduced windblown rain penetration into the grove. Hence, fewer copper sprays will be necessary after canopy closure promotes an internal windbreak effect. Under Objective 4 (To define risk for development of bacterial resistance to copper and streptomycin in FL citrus groves) the results were submitted and accepted for publication in European Journal of Plant Pathology. The results are briefly summarized as follows: Streptomycin was tested as an alternative to copper bactericides, which are routinely used for the control of citrus canker where the disease is endemic. A major concern is that excessive use of copper as a bactericide may lead to development of copper-resistant strains of Xcc. In this study, a semi-selective medium was developed to recover copper or streptomycin-resistant strains of Xcc from citrus leaves. The newly developed semi-selective medium was used to monitor the effect of a 21-day-interval copper or streptomycin spray program on Xcc for three consecutive seasons and on citrus epiphytic bacterial populations for two seasons in a commercial grapefruit grove. Although, no copper- or streptomycin-resistant strains of Xcc were isolated after three seasons, a significant increase over time was observed in the frequency of citrus epiphytic bacteria resistant to these chemicals. Under Objective 5 (rapid transfer improved canker management technology to the Florida citrus industry), 2011 canker management recommendations were published in the Florida Citrus Pest Management Guide and Citrus Industry Magazine. Oral presentations have been delivered at the Florida Citrus Show and at a multi-county extension meeting.
A trial was initiated in July for evaluation of Magna-Bon (copper sulfate pentahydrate) by trunk injection/infiltration on expression of HLB. The trial is located in Southern Grove in Clewiston, FL in a block of 8 year-old Valencia orange trees. There are 7 treatments with 20 trees per treatment and each treatment has 6 blocks of four trees in a completely randomized block design. Treatments: 1) Untreated Check; 2) One infiltration @ 500 ppm plus Magna-Bon 47; 3) One injection/year of Magna-Bon @ 1,000 ppm; 4) Two injections/year of Magna-Bon @ 1,000 ppm; 5) One infiltration @ 250 ppm plus Magna-Bon 47; 6) Two infiltrations @ 250 ppm plus Magna-Bon 47; 7) One infiltration Magna-Bon 47. Measurements are assay of flush leaves by qPCR at the Southern Gardens Diagnostic Lab(SHDL), sampling of flush leaves for Copper. Tree health assessment twice per season (April and October). First treament was made July 2011.
Objective 1 is to conduct a field evaluation of nutritional sprays for control of HLB and HLB symptom expression and yield. The field study was set up May 2010 in Southern Grove, Hendry Co., FL. Six treatments were located in 4 plots of 150 trees per treatment (interior 10 trees in each block were identified for PCR, leaf nutrition sampling, tree health and yield evaluation). Treatments were 1) non-treated check; 2) Nutri-Phite sprayed 4 times bimonthly; 3) N-Sure sprayed bimonthly; 4) Agra Sol Mn/Zn/Fe plus Nutri-Phite plus triazone urea sprayed bimonthly; 5) Keyplex 1400 DP plus Nutriphite plus triazone urea sprayed bimonthly; 6) Wettable powder nutrients (Diamond R #2) plus Nutri-Phite P+K sprayed bimonthly. The materials were applied to both sides of the tree in 125 gallons per acre with an airblast sprayer driven at 2 mph to obtain thorough coverage. Three disease ratings have been taken so far and a slight decline in tree health has been observed, but no significant treatment effects have been observed. There were no significant treatment differences in yield at the first harvest, after the initiation of treatments the previous April. Objective 2 is to determine the mechanism of HLB symptom suppression by foliar nutritional application, Hamlin sweet orange trees have been inoculated and are being treated bimonthly with the nutritional sprays treatments 1, 2, 3, and 5 from objective 1. Infection rate and progress are being monitored by qPCR monthly. By 6 months post inoculation (MPI), most inoculated trees tested Las positive by qPCR within the expected graft inoculation success rate. No treatment differences were apparent. At 6 months post inoculation Las detection was lost in multiple trees in treatments 2 and 5. Root samples were taken for DNA extraction and 7/7 and 6/7 trees were positive for Las in root tissue for treatment 2 and 5 respectively, while only 4/7 were positive in treatments 1 and 3. After pruning trees at 6 MPI for canopy management, sampling at 7 MPI showed a slight reduction in titer in the new flush of all treatments except treatment 1, where no Las was detected until 8 MPI. This suggests that treatments 2,3, and 5 may potentiate movement of Las to new flush where psyllids are most likely to feed and acquire Las. Sampling and fixing of plant material for microscopy of phloem and leaf blade tissue began at two months and continues monthly. Fixed samples were observed at 6 and 8 MPI and sectioning of 9 MPI samples is underway. At 6 MPI reduced phloem plugging and necrosis was observed in treatments 3 and 5, however these treatments had some symptomatic leaves without detectable Las. These leaves had abnormal starch buildup preferentially in phloem tissue instead of mesophyll cells. At 8 MPI no difference in phloem plugging or starch accumulation was observed among treatments. At 6 MPI disease symptom and tree health ratings were routinely taken. Initially no treatment differences were observed. By 8 MPI at least one tree in each treatment had begun to decline and treatment 2 showed the most severe symptoms. At 9 MPI all HLB inoculated trees in treatment 2 had significant decline, while there was no treatment effect in uninoculated trees. In the other three treatments symptom development was variable, but a strong decline of tree health was observed in HLB inoculated trees of all treatments. At 10 MPI most HLB inoculated trees were dead, so this replication was terminated and treatment of a second replication of the experiment was initiated. An additional full nutrient treatment without phosphite was added to test the nutrient treatment without phosphite.
Objective 1 is to conduct a field evaluation of nutritional sprays for control of HLB and HLB symptom expression and yield. The field study was set up May 2010 in Southern Grove, Hendry Co., FL. Six treatments were located in 4 plots of 150 trees per treatment (interior 10 trees in each block were identified for PCR, leaf nutrition sampling, tree health and yield evaluation). Treatments were 1) non-treated check; 2) Nutri-Phite sprayed 4 times bimonthly; 3) N-Sure sprayed bimonthly; 4) Agra Sol Mn/Zn/Fe plus Nutri-Phite plus triazone urea sprayed bimonthly; 5) Keyplex 1400 DP plus Nutriphite plus triazone urea sprayed bimonthly; 6) Wettable powder nutrients (Diamond R #2) plus Nutri-Phite P+K sprayed bimonthly. The materials were applied to both sides of the tree in 125 gallons per acre with an airblast sprayer driven at 2 mph to obtain thorough coverage. Three disease ratings have been taken so far and a slight decline in tree health has been observed, but no significant treatment effects have been observed. There were no significant treatment differences in yield at the first harvest, after the initiation of treatments the previous April. Objective 2 is to determine the mechanism of HLB symptom suppression by foliar nutritional application, Hamlin sweet orange trees have been inoculated and are being treated bimonthly with the nutritional sprays treatments 1, 2, 3, and 5 from objective 1. Infection rate and progress are being monitored by qPCR monthly. By 6 months post inoculation (MPI), most inoculated trees tested Las positive by qPCR within the expected graft inoculation success rate. No treatment differences were apparent. At 6 months post inoculation Las detection was lost in multiple trees in treatments 2 and 5. Root samples were taken for DNA extraction and 7/7 and 6/7 trees were positive for Las in root tissue for treatment 2 and 5 respectively, while only 4/7 were positive in treatments 1 and 3. After pruning trees at 6 MPI for canopy management, sampling at 7 MPI showed a slight reduction in titer in the new flush of all treatments except treatment 1, where no Las was detected until 8 MPI. This suggests that treatments 2,3, and 5 may potentiate movement of Las to new flush where psyllids are most likely to feed and acquire Las. Sampling and fixing of plant material for microscopy of phloem and leaf blade tissue began at two months and continues monthly. Fixed samples were observed at 6 and 8 MPI and sectioning of 9 MPI samples is underway. At 6 MPI reduced phloem plugging and necrosis was observed in treatments 3 and 5, however these treatments had some symptomatic leaves without detectable Las. These leaves had abnormal starch buildup preferentially in phloem tissue instead of mesophyll cells. At 8 MPI no difference in phloem plugging or starch accumulation was observed among treatments. At 6 MPI disease symptom and tree health ratings were routinely taken. Initially no treatment differences were observed. By 8 MPI at least one tree in each treatment had begun to decline and treatment 2 showed the most severe symptoms. At 9 MPI all HLB inoculated trees in treatment 2 had significant decline, while there was no treatment effect in uninoculated trees. In the other three treatments symptom development was variable, but a strong decline of tree health was observed in HLB inoculated trees of all treatments. At 10 MPI most HLB inoculated trees were dead, so this replication was terminated and treatment of a second replication of the experiment was initiated. An additional full nutrient treatment without phosphite was added to test the nutrient treatment without phosphite.
The objective is to evaluate soil-applied neo-nicotinoids and other SAR inducers on HLB disease progress in newly planted citrus trees subjected to psyllid-mediated infection or graft-inoculation. One yr-old Hamlin trees were planted in May 2009 and treated as follows: 1) non-treated check (UTC), 2) foliar insecticide to control psyllids, 3) soil-applied imidacloprid/thiamethoxam (IMID/THIA) to induce SAR, 4) soil-applied IMID/THIA plus foliar insecticides, 5) graft-inoculated UTC, 6) graft-inoculated with IMID/THIA. There were 50 trees per treatment (5 blocks of 10 trees). In 2009, the effect of SAR inducers on HLB infection progress was inconclusive perhaps attributable to the interaction of IMID/THIA with psyllid control which may have an uncontrolled effect on psyllid transmission. In 2010, the SAR inducer acibenzolar-S-methyl (ASM, Actigard 50WP) which does not control psyllids was substituted in treatments 3, 4 and 6. At 24 months after treatments began, 105 trees were PCR+ (35%) in the trial. Higher number of PCR+ trees occurred in the UTC (20), the UTC with graft inoculation (22), and the IMID/THIA/ASM with graft-inoculation (28). A lower number of PCR+ trees occurred in the treatments with SAR inducers (11), foliar insecticides (12), and foliar insecticide plus SAR inducers (12). Two years after treatments were initiated, the effect of SAR on HLB disease progress has been minimal, which indicates a lack of promise for SAR inducers in HLB management. This information has been communicated to citrus growers in a recent Citrus Industry magazine article entitled “The ABCs of SAR” (May issue).
Under Objective 1 and 2 the following was submitted and published in Plant Disease: Soil applications of inducers of systemic acquired resistance (SAR), imidacloprid, thiamethoxam or acibenzolar-S-methyl, at various rates and application frequencies were evaluated for control of citrus canker caused by Xanthomonas citri subsp. citri in a field trial of 3- and 4-year-old ‘Ray Ruby’ grapefruit trees in southeastern Florida. Reduction of foliar incidence of canker produced by one, two or four soil applications of imidacloprid, thiamethoxam, and acibenzolar-S-methyl was compared with 11 foliar sprays of copper hydroxide and streptomycin applied at 21-day intervals. In 2008 and 2009 crop seasons, canker incidence on each set of vegetative flushes was assessed as the percentage of the total leaves with lesions. By the end of the 2008 season, despite above average rainfall and a tropical storm event, all treatments significantly reduced foliar incidence of citrus canker on the combined Spring-Summer-Fall flushes. Sprays of copper hydroxide and streptomycin were effective for reducing canker incidence on shoot flushes produced throughout the season compared to the untreated control, whereas soil applied SAR inducers reduced foliar disease depending on rate, frequency and timing of application. Except for the treatment of four applications of acibenzolar-S-methyl at 0.2 g a.i. per tree or two applications of imidacloprid, SAR inducers were ineffective for reducing foliar disease on the flushes that were present during the tropical storm. In 2009, all treatments significantly reduced the incidence of foliar canker on the combined Spring-Summer-Fall flushes but not all treatments of Spring-Summer flushes with SAR inducers were effective compared to the untreated control. Hence, depending on rate, frequency and timing of application, soil-applied SAR inducers reduced incidence of canker on foliar flushes of young grapefruit trees under epidemic conditions. Under Objective 2 Integration of soil applied SARs with foliar applications of copper sprays for control of canker, we are testing the efficacy of the inducers for control of citrus canker on young fruiting Red grapefruit and Hamlin orange trees. We previously demonstrated that trunk application of imidacloprid is effective for canker control on fruit as 21 day copper sprays. To keep the chemicals out of the soil, we are spraying the SAR inducers on the trunk. These trials involve testing of non-registered acibenzolar-S-methyl and the insecticides imidacloprid, thiamethoxam at elevated rates. Hence, fruit from the treated trees must be dropped and destroyed (called ‘crop destruct’ of nonregistered chemical treatments). Under Objective 3, In 2011 the complementary use of ASM, THIA and IMID soil applications to increase and/or extend canker control in 3-yr-old grapefruit and 2-yr-old Vernia orange trees. The highest incidence of disease trees and/or leaves is in the unreated check in each trial compared with a very low incidence of canker in the integrated SAR treatments. A field trial with soil applied neonicotinoids in Parana, Brazil was evaluated. IMID (Confidor) as a soil drench and IMID (Winner) applied to trunk gave comparable in disease control activity on 2-yr old Valencia orange trees, as well as, the other neonicotinoids tested, THIA and Clothianidin. Clothianidin is now registered for use on non-bearing citrus in Florida, hence all of neonicotinoids registered for non-bearing citrus in Florida have been shown to have SAR-inducing activity against canker as well.
This project includes experiments being conducted by USDA-ARS and University of Florida on protecting newly planted citrus trees from Asian citrus psyllid and huanglongbing. This progress report reviews research conducted by University of Florida in Southwest Florida. Research reviewed pertains to the efficacy of insecticides for psyllid control. One study compares three rates of cyazapyr (150, 225, and 350 g ai/ha) against Admire Pro (280 g ai/ha) and Platinum 75g (140 g ai/ha) applied on 19,20 Jul to newly planted Hamlin trees. Evaluations were conducted 10,23 ‘ Sep. and no psyllids were found. However, all products have been 100% effective for leafminer control. Psyllid nymphs were observed 19-Apr-2011 on new flush and evaluations were made by removing 10 shoots per plot and counting nymphs under a stereo microscope. A more uniform sample was later obtained by pruning back and caging a single branch on 11 Apr on two trees in each plot. On 27-Apr , 10 adult psyllids from a lab reared colony at SWFREC were placed in each of the cages which was then sealed with a 6 inch twist tie to keep out predators. On 11-May, one shoot from each cage and two additional shoots randomly selected from the same trees in each plot were removed and nymphs were counted with a stereo microscope. Counted shoots were sent to a laboratory for HPLC residue analysis. On 19 April, fewer nymphs compared to the control were seen from all treatments except Admire Pro, with no differences among rates of cyazpyr or thiamethoxam. On 11 May, only the two highest rates of cyazapyr resulted in fewer nymphs compared to the control. All other treatment trees were 100 % infested with psyllids compared to 40 and 80% for the high and medium rate of cyazapyr. Thus, we were able to detect activity against psyllids from these two treatments 10 months after application to young citrus trees. Another study is designed to continue for 3 years with rotations of cyazpyr at the low and medium rate with Platinum and Admire Pro at the same rates as above every three months. The objective is to assess the ability of these treatments to delay or prevent the onset of HLB in an area of high incidence. Initial treatments were made 29-Jul on newly planted Hamlin trees and evaluations conducted 2,22-Sep with no psyllids being observed although all treatments have been 100% effective in controlling leafminer. Applications were repeated on 24-Jan and 25-April and evaluations on 13-Apr and 18- May by removing 10 shoots per plot and counting nymphs under a stereo microscope. Untreated shoots were highly infested, but no nymphs were observed on shoots from treated trees except on the last sample date when 2.4 nymphs per shoot were seen with the rotation that began with Admire Pro and employed the low rate of cyazapyr, although this was not statistically different from the other 3 insecticide treatments. Leaf samples are being analyzed for residues of all insecticides. A review of the efficacy of different insecticide treatments for suppressing psyllids and consequently HLB will be presented in this project’s next annual report.
The intent of this study is to examine the effect of windbreaks, copper sprays to reduce infection, and leafminer treatments to determine there individual and combined effects on control of citrus canker in Brazilian commercial citrus and the applicability of this strategy to the US commercial citrus industry. A preliminary study was published in Crop Protection 27:807-813, that indicated that copper and insecticide applications significantly reduced canker infection but windbreaks did not have any effect. As described previously, a new series of plots with much more extensive windbreaks were established via a USDA/ARS specific cooperative agreement with the University of Sao Paulo, and the Brazilian cooperator at an IAPAR farm, in Xambr’, Parana state, using, 2-yr-old P’ra on Rangpur lime. Windbreaks were completed and plants were be established in Mid April 2010, but severe winds damaged the windbreaks during two storm events. These windbreaks have been reinforced and rebuilt. This delayed the experiment which is now scheduled to begin in March 2012. The following treatments will then be applied: 1) no sprays (control), 2) Cu++ sprays to reduce citrus canker incidence, and 3) insecticide sprays to inhibit infestations of Asian leafminer (secondary effects). Main effects are windbreak versus no windbreaks. Citrus canker incidence will be estimated on multiple branches on each tree treated as the number of leaves per branch infected. We anticipate running these plots for 2-3 more years to collect all necessary data. The development of the Programmable leaf wetness controller (PLWC) software was written, debugged, is complete, and the control program is working well. New leaf wetness sensors were designed and constructed and calibrated. An electronic glitch was determined in the leaf wetness sensors and new control circuitry was designed and constructed to overcome the glitch. New hydrophobic materials used as the sensor bridge allowing the detection of leaf moisture are being evaluated as well. The newly designed probes with various sensor bridge materials are currently being tested Publications: 167. Bock, C. H., Graham, J. H., Gottwald, T. R., Cook, A. Z., and Parker, P. E. 2010. Wind speed effects on the quantity of Xanthomonas citri subsp. citri dispersed downwind from canopies of grapefruit tree infected with citrus canker. Plant Di Bock C.H., Graham, J.H., Gottwald, T.R., Cook, A.Z., and Parker, P.E. 2010. Wind speed and wind-associated leaf injury affect severity of citrus canker on Swingle citrumelo. Eur J. Plant Path 128:21-38 Bock, CH, Parker, PE, Cook, AZ, Graham, JH and Gottwald, TR. 2001. Infection and decontamination of citrus canker and inoculated the surfaces. Crop Protection 30:259-264. Hall, D.G., Gottwald, T.R. and C.H. Bock. 2010. Exacerbation of citrus canker by citrus leafminer Phyllocnistis citrella in Florida. Florida Entomologist. Florida Entomologist 93:558-566. Bock, C.H., Gottwald, T.R. and Parker, P.E. 2011. Distribution of canker lesions on the surface of diseased grapefruit. Plant Pathology (Accepted).
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