Work done this quarter:(1b) Adjuvant screeningPreliminary trials have been conducted with adjuvants alone to determine their lethality to lebbeck mealybug. A total of 9 adjuvants, including and 4 from Helena Agri Enterprises were mixed with DI water at label rates and sprayed until dripping on Volk lemon leaves with mealybugs attached. 8 of the adjuvants resulted in significantly greater mealybug mortality over a 7-day period compared to a DI water control. (1c) Evaluate promising materials in open grove setting: We began a field trial at a 10-acre commercial citrus planting that was heavily infested in 2020. First insecticide applications occurred as a pre-bloom prophylactic spray of a systemic material, with a second spray planned in the end of April.(1d) Ant Management · We are testing a variety of methods to remove red imported fire ants (Solenopsis invicta) from citrus groves. Removing the ants can allow natural enemies to prey on and parasitize lebbeck mealybug without interference, and provide more effective biological control. Our research goals are:1. Reduce abundance of fire ants associated with lebbeck mealybug (Nipaecoccus viridis) in central FL citrus groves. · 2. Determine long-term efficacy of different treatment types at reducing fire ant abundance · 3. Assess time and cost of different treatment types · 4. Determine effects of ant treatment on predators abundance within trees and within N. viridis clusters. We are collaborating with the King Lab from the University of Central Florida, and are conducting the experiment in the grove of a local citrus grower. Four experimental treatments will be tested and compared to an untreated control: Chlorpyrifos drench, Clinch ant bait (Abamectin), Extinguish ant bait (S-methoprene), and spot treating ant colonies with hot water. Fire ant abundance will be assessed by counting the number of surviving colonies, and by determining presence/absence of foragers with pecan sandie baits. Natural enemy abundance will be assessed by dissecting mealybug clusters for predator larvae. Sampling for each of these methods will take place every 2 weeks for a total of 3 months post-treatments. Currently, all treatments have been applied and we are conducting follow-up surveys to determine their efficacy. (1e) Evaluate management options for IPCsWe recently completed the study to evaluate several commercially available entomopathogenic fungi (EPF) products as a potential tool to control lebbeck mealybug infestation on young citrus trees within individual protective covers (IPCs). Findings of the study indicate that EPF can cause death of mealybugs and EPF conidia are viable on citrus trees within IPCs up to 42 days after treatment. Thus, EPF can offer control of mealybug for up to 1.5 months after treatment. In February, we began infesting trees in our research planting to run a similar comparison of topical insecticides for clean-up, with pretreatment drenches planned prior to the next set of trees to be infested. Additionally, a controlled greenhouse comparison of drench materials including aldicarb is planned once trees develop sufficient leaf mass to enable testing of residual post application of all chemistries used in addition to impact on mealybugs. We ran a methods trial and found that malathion lasted 2-3 times longer under IPCs compared to open field settings, suggesting that materials sprayed in IPCs that are known to have rapid UV breakdown may persist much longer and remain effective longer under IPCs. (2a) Predator assessmentsPredatory insects have been reared from locally infested plant materials the past two years, with the majority of predators emerging being parasitic flies, which consume a variety of piercing-sucking pest insects, mealybug destroyers, and one species of lacewing. A primer has been developed and evaluated to enable the detection of mealybugs in the guts of predators. Post-feeding retention time in a known predator was assessed and we feel confident that this tool will now enable screening of field caught arthropods that may play a role in the suppression of lebbeck mealybug. The commercially available predators Cryptolaemus montrouzieri, Orius insidiosus, Adalia bipuctata, and Hippodamia convergens have been screened using no-choice assays to determine if they will feed on larval lebbeck mealybugs, and on mealybug ovisacs. Additionally, several wild-caught predators have also been screened, including Harmonia axyridis and larva of the genus Ceraeochrysa (colloquially called trash bugs). Both adult and larval C. montrouzieri readily feed on lebbeck mealybug larvae and ovisacs, as do larval Ceraeochrysa. Adult Orius insidiosus, early instar Adalia bipunctata, and adult and larval Harmonia axyridis do not feed on lebbeck mealybug larvae or ovisacs. Adult Hippodamia convergens and Adalia bipunctata do feed on some lebbeck mealybug larvae, but appear to attack and kill the larvae more than fully consuming them. Neither feed on ovisacs. From our results, only Cryptolaemus montrouzieri shows promise as a commercially available biological control agent for lebbeck mealybug, although Ceraeochrysa larvae may act as natural biological control in the grove. (2d) Develop tools to minimize spread· Killing lebbeck mealybug with isopropyl alcoholWe tested different concentrations of isopropyl alcohol to determine how lethal they are to 1st instar lebbeck mealybugs. 50%, 70%, and 90% solutions of isopropanol were sprayed onto mealybug crawlers placed on cloth swatches, and compared to a DI water control. Both 1 spray at each concentration, and 2 sprays at each concentration were tested. Mortality was assessed after 5, 10, and 15 minutes. The test was repeated, this time assessing mortality at 30 min, 1hr, and 2hrs. For all concentrations of isopropanol, 1 spray resulted in significantly greater mealybug mortality compared to the control. However, several mealybug crawlers remained alive and active after a single spray for all concentrations. 2 sprays of each concentration resulted in almost 100% mortality or incapacitation at all time points. · Using steam to kill adult mealybugs and ovisacsAdult mealybugs and ovisacs may be accidentally transferred from infested groves on tools and equipment. Steam treatments may be an effective method of sanitizing equipment and killing both adults and ovisacs. Using a steam cabinet on-station, we steam treated adults and ovisacs at 100 and 120 degrees Fahrenheit for 5, 10, and 15 minutes to determine mortality. Previous preliminary experiments showed steam treatments at 130 degrees Fahrenheit for 15 and 30 minutes resulted in 100% mortality. Mortality was assessed at 0, 3, and 5 days after treatment, and ovisacs were held for an additional 2 weeks to see if crawlers emerged. At 100 degrees F for all times points, adult mealybug and ovisac mortality was not significantly different than the control. At 120 degrees F for 5 minutes, mortality also functionally 0%. However, mortality rose to 100% at 120 degrees F for 10 and 15 minutes. Currently, we are running trials on adults and ovisacs treated at 130 degrees F for 5 and 10 minutes, and preliminary results show 100% mortality. Work planned for next quarter(1b) Adjuvant screening will continue to determine optimal adjuvants to work in synergism with insecticide sprays.(1c) We will continue treating and scouting the grove we have started a management comparison trial at.(1d) The ant management project will continue throughout the upcoming quarter with the addition of monitoring for predatory insect establishment.(1e) IPC management management trials will continue, looking towards conventional materials for management and spray penetration by tractor mounted sprayers.(2a) Working with FDACS, we have permission to deploy sentinel infested materials to screen more broadly for predators that may be present in the system which we missed with haphazard rearing from infested grove sites.(2c) Feeding mechanisms will continue to be worked out using EPG with the future plan to determine if we can interfere with their feeding via chemical intervention leading to reduced offspring production.(2d) We will continue evaluating sanitation options and are planning to look at vehicle sanitation in the next quarter.