Sustainable Management Options for Whitefly-transmitted Viruses in Cucurbit Production

Objective 1. Pre-planting integrated management

Seedlings in the greenhouse are vulnerable to whiteflies and viruses. The goal here is to evaluate protection measures on transplant seedlings in the greenhouse. Experiments for objective one were conducted at the UGA Tifton Greenhouse facility (Plant Pathology Department). Seedling protection options such as whitefly-proof screen (ProtekNet0.01 x 0.01 mesh), tray drenching with systemic insecticide cyantraniliprole (Verimark), biorational insecticide (Requiem), and plant defense modulator (salicylic acid analogue Actigard) were evaluated. Treatments were replicated in a split plot design. The presence of the insect proof screen and lack thereof were considered as main plot effects. The insecticide, biorational insecticide, and plant defense modulator were considered as sub-treatments. The main plot treatments were replicated three times in the open greenhouse. The main treatments effects were applied in alternating bench rows. The sub-plot treatments were replicated as five seedling flats for each. Whitefly infestations and virus infection percentages in each main and sub-treatment were assessed two weeks after seeding, and treatment efficacy were statistically evaluated.

Adult whiteflies on two leaves in each seedling flat were counted by turning leaves. In addition, two leaves were removed from each seedling flat and taken to the vector biology laboratory in Griffin GA where eggs and immature whiteflies were counted at 20X magnification using a dissection microscope. All flats were visually screened for virus symptoms. Plant samples were randomly sampled from each seedling flat and DNA extractions were conducted using specific primer sets and already established protocols. DNA samples were used to test for CuLCrV, Polymerase chain reaction (PCR) was performed to confirm CuLCrV infection in the seedlings using protocols already optimized in the molecular diagnostic laboratory in the Department of Plant Pathology at UGA, Tifton GA

Data analyses

Random count data of eggs, nymphs, and adults were analyzed using generalized linear mixed models by PROC GLMMIX in SAS (SAS Enterprise 4.2, Cary, NC) with suitable (Logistic) transformation at a 95% confidence level. Main plot and sub-plot treatments were considered as fixed effects and replications were considered random effects. Treatment means were separated using the LS means option in SAS at alpha =0.05. Percentages of infection data were treated as binomial responses (infected vs non-infected). Proportions of plant samples testing positive in each treatment/sub-treatment were analyzed by logistic regression using PROC GENMOD in SAS. Treatment means were separated using paired contrasts at alpha =0.05.

Objective 2

Post-planting integrated management

a. Screening for tolerance to whiteflies and/or viruses in squash cultivars.

Commercially available yellow and zucchini squash cultivars (up to 20, including new) were screened for susceptibility/tolerance to viruses. This trial was conducted both in Georgia and Florida.  Squash cultivars were transplanted as seedlings after germinating seeds in seedling flats in the greenhouse. Transplants were planted in raised beds covered with white plastic mulch. The raised beds were fumigated using Chlorpicrin at least two weeks prior to transplanting. Beds were on 6-ft centers with 1-ft plant spacing within rows. Single row plots were utilized for screening cultivars. Plots were15-ft long with 10-ft unplanted borders between plot ends. The test was conducted as a randomized complete block design with four replications. Plots were drip irrigated and fertilized weekly. All inputs fertilizers, fungicides, and insecticides were administered as per grower standards and as prescribed by UGA/UFL cooperative extension guidelines.

Insect counts, virus incidence, symptom severity counts were obtained weekly. Adult whiteflies on one leaf located in the middle of the plant on five plants in each plot were counted. In addition, one leaf was removed from each of the five-sampled plants and taken to the laboratory where eggs and immature whiteflies were counted at 20X magnification using a dissection microscope in one square inch. Immatures were divided into small nymphs (first and second instar) and large nymphs (third and fourth instar). The percentage of plants exhibiting virus symptoms such as severe stunting, leaf curling, yellowing, and stunting at least four times from planting to harvest. Severity rating of virus symptoms were assessed on a 1 to 5 scale with 1 representing mild or no symptoms and 5 representing very severe symptoms or death. Plant vigor were rated on a 1-9 scale with 1=death and 9=extremely vigorous. Leaf samples were collected from symptomatic and non-symptomatic genotypes and tested by polymerase chain reaction (PCR) to confirm virus infection. Yields were obtained by hand picking and weighing fruits from ten plants in each plot.

Random count data of eggs, nymphs, and adults were analyzed using generalized linear mixed models by PROC GLMMIX in SAS with suitable transformation at a 95% confidence level. Percentages of infection data were treated as a binomial response (infected vs non-infected). Proportions of plant samples testing positive in each treatment were analyzed by logistic regression using PROC GENMOD in SAS. Treatment means were separated using paired contrasts at alpha =0.05. The symptom severity and yield data will also be analyzed using PROC GLIMMIX assuming a Gaussian distribution.

b. Reflective mulch and companion crop and effects on whiteflies and viruses

The second trial evaluated the effects of reflective and live mulch; white mulch was treated as the standard control. The live mulch chosen in this trial was buckwheat (Fagopyrum esculentum). Three-row plots were used for this trial. The plots were at least 20 ft in length with 10 feet alleyways between adjacent plots within a row. Seedlings were transplanted in raised beds with the white and reflective mulch. For the live mulch plot, the crop was planted in white plastic, and the inter-row space on either side of the middle row was seeded with buckwheat seeds using a modified grain seed drill. Buckwheat seeds were planted at least two weeks prior to planting the squash crop. A randomized complete block design was used, and each treatment was replicated four times. Buck wheat plants were also irrigated through drip lines.

Whitefly adults and immatures counts Adult whiteflies on five leaves in each plot were counted by turning leaves. In addition, five leaves were removed from each seedling flat and taken to the Vector biology laboratory in Griffin GA (Srinivasan) where eggs and immature whiteflies were counted at 20X magnification using a dissection microscope, virus incidence, virus severity, and yields were assessed as described in trial 1. Random count data of eggs, nymphs, and adults were analyzed using generalized linear mixed models by PROC GLMMIX in SAS as described above. Percentages of infection data were treated as a binomial response (infected vs non-infected). Proportions of infected plants in each treatment were analyzed by logistic regression using PROC GENMOD in SAS as described above. The symptom severity and yield data will also be analyzed using PROC GLIMMIX assuming a Gaussian distribution.

c. Insecticides, biorationals, and row cover: effects on whiteflies and virus incidence.

Three conventional insecticides (Imidacloprid (Admire Pro) cyantraniliprole (Exirel) and Fluorpyradifurone (Sivanto), biorational insecticides including Chenopodium extract (Requiem), Crop oil, and Chromobacterium subtsugae strain PRAA4-1 (Grandevo), and row cover (Remay) were evaluated. All insecticide treatments were evaluated using randomized complete design with four replications for each treatment. All insecticides were applied as foliar sprays using a tractor mounted sprayer at the rate of 50 gallons per acre using fan tip nozzles and guard to reduce drift. Row cover was used as a treatment for the first four weeks after planting or at first female flowering whichever is earlier. No insecticide sprays were undertaken for the row cover treatment until the covers were removed. Upon removal, weekly sprays with a conventional insecticide were undertaken.

Whitefly adults and immatures counts, virus incidence, virus severity, and yields were assessed as described in trial a.