1. Please state project objectives and what work was done this quarter to address them:
Objective 1: Evaluate currently available registered insecticides in Florida citrus against DRW.
Sub-objective 2: Evaluation of B. thuringiensis tenebrionis (Btt) activity against DRW.
Effect of combined EPN-Btt on the mortality of Diaprepes root weevils
The effects of entomopathogenic nematode (EPN)-Btt pesticidal protein combinations on Diaprepes root weevil mortality was determined. Neonatal and 5-week-old larvae were exposed to Steinernema riobrave (100 IJs/ cm2), Btt CX-2330 (3000 ppm or g AI/ml DI water) and a combination of these agents against an untreated control.
Five-week-old larvae were transferred to 589 cm3 containers filled with a soil-based mix (three parts peat moss, two parts coco peat, one part perlite, and one part gravel-sand-soil mixture). Each cup contained 28 g of moist soil with or without Btt. For treatments including Btt, the soil was moistened with Btt suspension at the appropriate concentration and was thoroughly mixed. Each treatment was replicated nine and six times with 20 neonatal and 5 5-week-old larvae per replicate, respectively. The nematodes were added 27 days after Btt application (Li et al., 2021; Koppenhofer & Kaya, 1997). Experiments were conducted in a growth chamber at 27°C with a photoperiod of 14:10 (L:D) h. Mortality and weights of surviving larvae was assessed six weeks after inoculation.
One-way ANOVA was used to test for significant differences among the treatments in greenhouse experiment. Multiple comparisons between treatments were tested by LSD test (P < 0.05). Results as of December 2023 The mortality rate of larvae differed between treatments 42 days after inoculation of neonatal larvae with treatments (F = 14.75; d.f. = 3, 32; P < 0.000). There was no difference detected between the untreated control and Btt treatments, whereas both treatments differed from treatments with EPN or Btt+EPN (Fig. 1). The combination of EPN and Btt in the rhizosphere increased the dry weight of roots and shoots in a synergistic manner (F = 9.065; d.f. = 3, 32; P < 0.000; F = 0.727; d.f. = 3, 32; P < 0.544) (Figs. 2). The average weights (fitness) of surviving larvae were affected by Btt (Figure 4) and this effect may reduce the pest's immune system in the face of EPN. The mortality rate of 12-week-old larvae increased only in the presence of EPN (F = 12.322; d.f. = 3, 20; P < 0.000). Mortality of 5 wk-old larvae exposed to EPN was unaffected by Btt (Fig. 5). Similar to the previous experiment, the dry weight of roots and shoots increased in the presence of Btt and EPN; however, without an interaction (F = 1.226; d.f. = 3, 20; P < 0.009; F = 12.322; d.f. = 3, 20; P < 0.326) (Fig. 6). Conclusion Our results suggest that combining entomopathogenic nematodes (EPN) with Btt pesticidal proteins application increases protection of citrus roots compared with using either technique alone. EPNs have been an effective technique for managing Diaprepes; however, variation in efficacy is historically known. Combining EPNs with Bt may be one avenue for improving both consistency and efficacy of tree protection from Diaprepes. 2. Please state what work is anticipated for next quarter: Obj 1: We will continue laboratory evaluations of Btt formulations +/-EPNs Obj 2: We will continue our weekly trapping for activity and plan to scout new sites for the mark-recapture work to be repeated in a location, likely on the east coast, with higher Diaprepes populations 3. Please state budget status (underspend or overspend, and why): We should be close to on track for budget use 4. Please show all potential commercialization products resulting from this research, and the status of each: none at this time