Management of diamondback moth with emamectin benzoate and Bacillus thuringiensis subsp. aizawai insecticides

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1 Management of diamondback moth with emamectin benzoate and Bacillus thuringiensis subsp. aizawai insecticides Ronald F. L. Mau 1, Dennis M. Dunbar 2, Laura Gusukuma-Minuto 1 and Robin S. Shimabuku 3 1 Department of Entomology, College of Tropical Agriculture and Human Resources, University of Hawaii at Manoa, Honolulu, Hawaii Merck Research Laboratories, 7555 North Del Mar Avenue, Suite 24, Fresno, California Hawaii Cooperative Extension Service, 31 Kaahumanu Avenue, Building 214, Kahului, Hawaii Abstract Emamectin benzoate (= MK-244) was evaluated with Bacillus thuringiensis kurstaki (Btk), B.t. aizawai (Bta) and other insecticides against diamondback moth, Plutella xylostella (L.), (DBM) on cabbage in experimental field plots and on grower farms. Conventional insecticides were ineffective against DBM and resulted in poor yields. Bta insecticides provided moderate levels of DBM control and low to moderate cabbage yields. Emamectin benzoate provided superior control of DBM and other lepidopteran larvae in all on-station experiments as well as in field trials on four commercial farms. Growers obtained an average increase in marketable yield of 29 percent and an estimated net increase in gross revenues of about $66 per hectare. Commercial use of emamectin benzoate is being developed for use on vegetable crops in the U.S. under the trademark PROCLAIM 5 SG. This insecticide has positive characteristics for use in IPM programs. Its rapid degradation on leaf surfaces provides a good margin of safety for parasitoids of DBM and other pest species. Translaminar residual activity provides excellent protection against larvae of DBM and other cruciferous Lepidopterans. Key words: IPM, cabbage, biorational insecticides Introduction The major cabbage crops grown in Hawaii are green head and Chinese (Napa) cabbage. About 45 hectares of both types of cabbage are produced annually primarily for local consumption. Cabbage is cropped throughout the year in sequential plots. The majority of growers plant and harvest cabbage on weekly schedules. The major lepidopteran pests in order of importance are DBM, imported cabbage webworm (Hellula undalis), imported cabbageworm (Pieris rapae) and cabbage looper (Trichoplusia ni). DBM is the key Lepidopteran pest of cabbage in Hawaii. It occurs on all islands where crucifers are grown. DBM causes extensive losses primarily during the period April through October, but it has been a problem during other times of the year. DBM is a formidable pest due its prolific nature and ineffectiveness of registered pesticides (Tabashnik et al. 199, 1992; Mau et al. 1995a, 1995b, 1995c, 1995d). Although selection of tolerant cabbage varieties and conservation of parasitoids are viable management tactics, it has been very difficult to provide economic control of the pest without an effective insecticide. During the past five years, the cabbage industry has lost nearly $1.3 million due to DBM related causes. Economic losses occur as a result of direct feeding damage to the cabbage, physical presence of caterpillars within marketed heads, additional harvest labor costs, and increased insecticide and application expenses. A case study was conducted for the island of Maui. DBM reduced marketable cabbage yields practically every year during the past 9 years. The overall yield reduction during the period was 4 percent, and this occurred despite a 15 percent increase in harvested acres. Yields have decreased because of the lack of an effective insecticide that can be used when treatment threshold is surpassed. Bta products are somewhat effective during periods of lower DBM population density, but they do not provide adequate protection at higher population densities. Other non-chemical tactics provide supplementary protection and have not provided adequate levels of control on commercial farms. The results from laboratory and field evaluations of a second generation avermectin insecticide, emamectin benzoate (= MK-244) and two Bta products are presented in this paper. Emamectin benzoate is a novel semi-synthetic avermectin insecticide derived from the fermentation product, avermectin B 1 (abamectin). It is a second generation avermectin insecticide that is a chemically modified derivative of abamectin. Other closely related compounds were found to be extremely active on lepidopteran larvae (Dybas and Babu, 1988; Dybas et al., 1989; Lasota and Dybas, 1991). 178 Proceedings: The Management of Diamondback Moth and Other Crucifer Pests

2 Materials and Methods Bioassay tests Laboratory bioassays were performed using larvae obtained from a laboratory colony of DBM established from field collections of DBM at Kamuela, Hawaii. The larvae were reared on head cabbage leaves. Head cabbage variety C-G Hybrid was seeded in community pots. Each pot contained approximately 1 cabbage plants. The potted seedlings were at the 5 7 true leaf stage when they were treated. One liter of each insecticide solution was prepared by diluting the appropriate amount of insecticide with water. The spray adjuvant Nu-film P (Miller Chemical & Fertilizer Corporation) was added at a rate of.47 ml per l. Treatment was accomplished by inverting the community pot and completely dipping the plants in the appropriate insecticide solution. The plants were allowed to air dry before they were used. The bioassays were performed at 75 C, 76 % RH and a 24 hour photoperiod. After placement on the leaves, mortality was recorded daily. The original leaves were replaced with new leaves from the corresponding treated plants two days later. Larvae were reported as dead when there was no movement of the larva when probed. Results are reported as percent mortality. Actual mortality was transformed using arcsine transformation and the data set was subjected to analysis of variance (SAS Institute, version 6.4). Means were separated using Tukey s studentized range test. Bioassays with neonate DBM larvae The efficacy of emamectin benzoate.16ec (3.4 g a.i.per l) was evaluated against four Bacillus thuringiensis (Bt) insecticides. The Bt products were Dipel 2X (Abbott Laboratories), Mattch Bio-insecticide (Mycogen Corp.), MVP (Mycogen Corp.) and Xentari (Abbott Laboratories). Ten neonate DBM larvae were placed on each treated leaf. All treatments were replicated 1 times. Bioassays with fourth instar DBM larvae Feeding studies were conducted on cabbage treated with emamectin benzoate.16ec at 3.4 g a.i. per liters of water. Ten fourth instar larvae placed on leaves treated 24 hr earlier. Mortality observations were made 12 and 24 hours after placement. Field studies Three season-long field experiments were conducted during 1995 against DBM, imported cabbage webworm (Hellula undalis), imported cabbageworm (Pieris rapae), and cabbage looper (Trichoplusia ni). The experiment station trials were conducted during March to November at the University of Hawaii, College of Tropical Agriculture and Human Resources experimental farm at the Maui Agricultural Park. The farm is located at the 1,4 ft. elevation of Mt. Haleakala on the Hawaiian island of Maui. Each of the field trials utilized randomized complete block designs with four replications (blocks). Each treatment plot measured.39 ha. Treatment plots in each block were separated by a 1.67 m. row spacing. Blocks were separated by a 3 m. wide strip that allowed sprayer access. Total field size was.118 ha. Mean daily temperatures during the three studies was 22, 23.6, and 22.9 C, respectively. The mean daily rainfall was 8.9,.5, and 2.3 mm, respectively. Head cabbage variety Tastie (Takii Seed Co.) was used in all tests. Cabbage seedlings in the 3 5 true leaf stage were planted into each plot in 4 rows on.91 m. centers and.457 m. within row spacing. Adjacent rows were offset to allow equidistant plant spacing between rows. The field was irrigated by overhead sprinklers at 2 3 day intervals. Irrigation was withheld for at least 24 hours after each spray application. Treatments were made at 7-day intervals until one week before harvest. The non-ionic surfactant, Excel 9 (Brewer Environmental Industries, Inc.) was added at a rate of 585 ml per ha. A total of 7 applications were made during each field test. Each treatment was applied at a pressure of 3.2 kg per cm 2 and a rate of 1,169 l per ha using a PTO driven tractor mounted sprayer. The spray boom was fitted with one hollow cone TX-18 sprayer nozzles (Spraying Systems Co., Wheaton, Illinois) placed directly over each row. Harvest evaluations were performed 8 weeks after transplanting. Data collected for each plant were Lepidopteran numbers, damage ranks, and marketability. The following stratified sampling method was used to select the plants for evaluation. The remaining plants in the two center rows were numbered from The second row was numbered from the opposite end of the plot. Every fifth plant was harvested until a total of 1 heads had been harvested. The loose wrapper leaves were removed from each head before data was collected. Larval feeding damage ranks ( 5) were assigned for each head. A rank was assigned if there were no damage. The numerical rating increased as the degree of caterpillar damage increased. A 5 rating was assigned where there was extensive larval damage. A marketability assessment was made after each head was ranked for damage and dissected for pest numbers. The marketability assessments followed procedures used by commercial growers and was based on the severity of insect damage on the wrapper and inner leaves of the head. Lepidopteran pest data were determined by removing and inspecting ten randomly selected plants. A pre-treatment census for lepidopteran pests was performed the day before the first treatment. Subsequent pest evaluations were subsequently performed at two week intervals. This was done six days after prior treatment. Commercial farm tests Four commercial growers were selected for commercial validation tests under a U.S. EPA approved experimental use permit (EUP) for PROCLAIM.16 EC Insecticide. Two growers Chemical control 179

3 were located in the Kula district on the island of Maui and the remaining two growers were located in the Kamuela district on the island of Hawaii. All growers applied PROCLAIM.16 EC insecticide at a rate of 3.4 g a.i. per ha. Treatments began immediately after the cabbage was transplanted and were repeated at 7 8-day intervals. The final application was made 7-days before harvest. Data on lepidopteran larvae numbers were taken immediately prior to the first treatment, 6 days after the third, 6 days after the fifth treatment. Comparisons of the performance of PROCLAIM.16 EC treatments were made against each grower s standard pest control practice. Their standard practices varied considerably. Grower 1 used a combination of Xentari at 367 g/ha and naled 8 E at 383 ml/ha a total of 1 times at 3 7-day intervals. Grower 2 made a total of 13 treatments to the grower standard field. He applied Xentari at 91.7 g/ha and mevinphos at 574 ml/ha was applied at 7-day intervals for the first two treatments. Three days later an application on MVP at 766 ml/ha was made. This was followed by an application of Xentari at 91.7 g/ha and methomyl at g/ha four days later. Dipel 2X at g/ha and mevinphos at 574 ml/ha was applied three days later followed by Xentari at g/ha three days later. Thereafter, the following applications were consecutively made at 3 4 day intervals: MVP at 766 ml/ha, Xentari at g/ha, Dipel 2X at g/ha, MVP at 766 ml/ha, Xentari at g/ha, Dipel 2X at g/ha, and MVP at 766 ml/ha. Grower 3 made 5 applications of Xentari at g/ha at 7-day intervals. No other insecticide applications were made. Grower 4 sprayed to control DBM adults as well as larvae. He first treated with a combination of mevinphos at 355 ml/ha and Xentari at g/ha. It was followed by an application of Xentari at g/ha three days later. Subsequently, the following consecutive treatments at 7-days intervals were naled 191 ml/ha; Xentari at g/ha; Ambush at 27 ml/ha; mevinphos at 355 ml/ha; Xentari at g/ha. Harvest evaluations were made for the emamectin benzoate treatment and from an adjacent plot that were treated with the growers standard pest management protocol. Ten plants were randomly selected from each of the plots and harvested using commercial protocols. Data was collected in the same manner described above for the experiment station tests. Statistical analysis The number of immatures counted were subjected to analysis of variance (ANOVA). Mean separation was accomplished using Tukey s mean separation test (SAS for Windows version 6.8). Results Laboratory DBM Feeding Studies Bioassays with neonate DBM larvae Leaf ingestion bioassays were conducted to compare the relative effectiveness of emamectin benzoate, Btk, and Bta products against neonate DBM larvae. The emamectin benzoate (= MK-244) treatment gave very rapid kill compared with that of the Btk and Bta products (Table 1). All of the DBM larvae died within 1 day after placement on treated leaves. In comparison, none of the Btk (Dipel 2X and MVP) and Bta (Mattch and Xentari) treatments gave complete kill, and mortality in all of the Bt treatments increased more slowly than in the emamectin benzoate treatment. Bta treatments were more effective than Btk treatments. Mattch gave the best overall control among all of the Bt treatments. Bioassays with fourth instar DBM larvae Larvae fed briefly and most of them ceased feeding within 4 8 hours after placement. Fifty one percent of the larvae were paralyzed within 4 hours after placement. Seventy-one percent of the larvae were paralyzed within 8 hours of placement, and all of the larvae were paralyzed within 24 hours after placement. All larvae died within 48 hours after placement. Comparative studies showed that the total leaf area consumed by a fourth instar larva in the Table 1. Evaluation of emamectin benzoate (MK-244,.16 EC) and Bacillus thuringiensis insecticides against diamondback moth larvae Treatment rate per ha Mean percent mortality (DAT) Mattch 4.66 l 5.2b 42.3b 7.2b 83.9abc Mattch 2.33 l 3.2b 36.8b 71.8b 82.1ab Xentari 1.12 kg 9.2b 22.9bc 46.9bc 61.7bcd Xentari.56 kg 9.4b 37.2b 49.1bc 57.7bcd Dipel 2X 2.24 kg 11.b 15.9bc 28.5cd 51.9cd MVP 4.66 l.b 15.3bc 34.8cd 49.7d MK EC 72 ml 1.a 1.a 1.a 1.a Untreated check 4.7b 7.7c 9.7d 11.5e DAT = days after treatment. Numbers within the same column followed by a different letter are significantly different (P<.1, Tukey s studentized test, SAS Version 6.4). Data were subjected to arcsine transformation prior to analysis. Untransformed mean percent mortality is presented. Nu-Film ml/ l of spray solution was used as a surfactant to assist in wetting the cabbage leaves. 18 Proceedings: The Management of Diamondback Moth and Other Crucifer Pests

4 emamectin benzoate treatment was slightly larger than its head capsule (.55 cm 2 ). Larvae on untreated leaves consumed about ten times more tissue (5.7 cm 2 ). The residual activity of emamectin benzoate within treated leaves was estimated using 3rd and 4th instar DBM larvae. Placements were made on leaves excised from treated plants, 1, 2, 3, 7, and 1 days after treatment. The plants were held in an enclosed greenhouse after treatment. Emamectin benzoate residues were highly active for the entire duration of the experiment. Ninety-four to one hundred percent of the larvae died within 3 days of placement in all of the post-treatment residue groups. These results demonstrated that emamectin benzoate could conceivably provide adequate levels of DBM control in the field for 1 days. On-station experiments Both of the field experiments were conducted during periods when DBM populations were high. Both formulations of emamectin benzoate provided excellent control of DBM and kept larval densities at relatively low levels when compared with other treatments (Figures 1 and 2). Bacillus thuringiensis treatments (Xentari, Dipel 2X, and Mattch) provided only moderate levels of control when compared to the Untreated Check (UTC) and emamectin benzoate treatments. Permethrin, methomyl, and endosulfan did not provide any DBM control. In these and other field experiments, we observed increased larval densities about 2 3 weeks after transplanting. It appears that this is due to a combination spray coverage and efficiency of the treatment issues. It is difficult to effectively deliver sprays to the apical bud because of the cupped leaves. There were very distinct differences in marketable yield among the treatments. Both formulations of emamectin benzoate provided the best overall yield (88 95%) in both experiments. Yield from Bacillus thuringiensis treatments varied from poor to moderate (18 68% yield). Mattch provided moderate yields when used at the.77 l per ha application rate. There was no marketable yield from the permethrin, methomyl, endosulfan and the Untreated Check treatments. Grower evaluation of PROCLAIM.16 EC insecticide Significantly better control of DBM was obtained with fewer pesticide applications of PROCLAIM.16 EC compared with their respective standard programs. The harvest DBM densities presented in Figure 3 are indicative of DBM larval densities that were maintained through the crop season. PROCLAIM.16 EC provided rapid kill of DBM larvae and consequently very little direct damage occurred in these plots. DBM larvae were scarce in PROCLAIM.16 EC treated plots compared with that of the growers standard program. There were significant increases in marketable cabbage yields in fields treated with PROCLAIM.16 EC compared with those with each grower s standard program (Figure 4). Yields of all PROCLAIM.16 EC treated fields exceeded the comparison fields and the Cooperative Extension average yield target (613 kg/ha). In two cases, PROCLAIM.16 EC yields exceeded Extension s maximum yield target (7343 kg/ha). Yields in three of four tests using standard grower practices fell short of extension s estimate for average Mean DBM Larvae/Plant Per Cent Marketable MK EC (72 ml/ha) Xentari (182 g/ha) Dipel 2X (182 g/ha) Mattch (382 ml/ha) Permethrin 3.2E (47 ml/ha) Untd Check MK EC (72 ml/ha) Mattch (382 ml/ha) Xentari (182 g/ha) Dipel 2X (182 ml/ha) Permethrin 3.2E (47 ml/ha) ;Untd Check June 29 Dates June 13 May 3 May 15 (Pre-trtmt) Figure 1a. Evaluation of emamectin benzoate.16 EC and other insecticides against diamondback moth larvae. Comparison of larval number at different stages of crop growth. Figure 1b. Evaluation of emamectin benzoate.16 EC and other insecticides against diamondback moth larvae. Comparison of harvest yields. Chemical control 181

5 Mean DBM Larvae/Plant MK-244 (72 ml/ha) MK-244 5SG (27 g/ha) Mattch (764 ml/ha) 8 Endosulfan 2CO (764 ml/ha) Methomyl 2.4 LV (574 ml/ha) Untd Check 6 Per Cent Marketable MK EC (72 Ml/ha) MK-244 5SG (27 g/ha) Mattch (764 ml/ha) Endosulfan 2CO (764 ml/ha) Methomyl 2.4 LV (574 ml/ha) Untd Check Sept. 5 Aug. 21 Aug. 7 July 24 Dates July 15 (Pre-trtmt) Figure 2a. Evaluation of emamectin benzoate.16 EC, 5 SG and other insecticides against diamondback moth larvae. Comparison of larval numbers at different stages of crop growth. Figure 2b. Evaluation of emamectin benzoate.16 EC, 5 SG and other insecticides against diamondback moth larvae. Comparison of larval numbers at different stages of crop growth. Mean DBM Per Plant at Harvest Grower standard Proclaim.16 EC (72 ml/ha) Grower 1.1 Grower 2.14 Grower 3.2 Grower 4 Figure 3a. Evaluation of PROCLAIM.16 EC by commercial growers in Hawaii. Comparison of DBM larval control with PROCLAIM and the growers standard treatment. Marketable yield (%) Grower 1 Grower 2 Grower 3 Grower 4 Figure 3b. Evaluation of PROCLAIM.16 EC by commercial growers in Hawaii. Comparison of harvest yields from PROCLAIM and growers standard treatment plots. Grower standard Proclaim.16EC (72 ml/ha) 182 Proceedings: The Management of Diamondback Moth and Other Crucifer Pests

6 Yield per hectare (kg) 8 ;; 6 4 ;; 2 State mean CES target ave yield CES target max yield Proclaim.16 EC (72 ml/ha) Grower STD Figure 4. Comparison of grower cabbage yields with State of Hawaii and Cooperative Extension standard yields. cabbage yields. The grower that surpassed extension s average yield made high insecticide expenditures by spraying twice each week for practically the entire crop cycle. Discussion The present Bta insecticides that growers use, Xentari and Mattch Bio-insecticide, gave varying rates of control. Although they were the most effective U.S. EPA registered DBM larvicides, the products performed best at lower DBM population densities and did not come close to providing adequate levels of control during periods when population levels were highest. Taking this into account, University of Hawaii Cooperative Extension recommended their use in combination with DBM adulticides (mevinphos and naled) with the Bta larvicides. Early evening applications of either mevinphos or naled in combination with one of the Bta insecticides reduced egg deposition by killing the adults and reduced larval damage. This practice generally gave better control than with the Bta insecticides alone, but many growers did not follow the recommendation. To complicate matters, the use of mevinphos was banned in November In contrast, emamectin benzoate.16 EC and 5 SG provided superior control of DBM and other Lepidopteran larvae at low as well as at very high population levels. Both formulations were equally effective. Although the grower validation trials involved the use of only the emulsifiable concentrate formulation, we found no reason to expect that the soluble granule formulation would not perform as well. The use of PROCLAIM.16 EC under the EUP by the commercial growers resulted in an average increase in marketable yield of 29 percent and an estimated net increase in gross revenues of about $66 per ha. Yields were particularly impressive when they were compared with the State s average yield and Cooperative Extension target yields (Figure 4). Grower satisfaction of the new product was very high. They felt that it was very easy to use and that the results were predictable. As a commercial product, emamectin benzoate will fit well into the present DBM management program. It has positive characteristics for use in an IPM program. Its rapid degradation on leaf surfaces provides a good margin of safety for parasitoids of DBM and other pest species. The rapid toxicity and long residual activity within the leaf tissue should provide excellent protection against larvae of DBM and other cruciferous Lepidopterans. Although the outlook for DBM management in Hawaii is positive, there is a potential downside to registration of the product. Due to the lack of other highly effective products, we can expect that growers might rely only on this product and select for genetic resistance. We have no knowledge about the propensity for resistance development, but have taken action to reduce the use of emamectin benzoate. Hawaii Cooperative Extension entomologists devised a DBM management program that involved the use of a tolerant cabbage cultivar (Scorpio) and the application of insecticide combinations to reduce adult and larval densities. Growers were advised to make weekly Bta treatments in combination with mevinphos or naled. The applications were made immediately after sunset to have the greatest impact against adults. A resistance management protocol was mandated when the industry in the State of Hawaii was granted a emergency exemption for use of PROCLAIM 5SG on cabbage in June Until other highly effective products become available, we will attempt to manage resistance by integrating cultural controls with judicious use of PROCLAIM 5 SG and Bta products. Growers are encouraged to plant varieties such as cv. Scorpio that seem to be more tolerant to DBM than others. Insecticidal control of DBM during the first half of the season is limited to Bta or other insecticide products unless DBM larval densities exceed 1 larva per plant. PROCLAIM 5 SG can be used if this threshold is exceeded, but its use is normally limited during the second half of the crop season. To limit unnecessary use, treatments are generally governed by a.5 DBM larva per plant treatment threshold except during the last two weeks prior to harvest when marketability can be greatly reduced at even low DBM densities. References Dybas, R.A., N. J. Hilton, J.R. Babu, Ranz A. Preiser, and G. J. Dolce (1989). Novel second-generation avermectin insecticides and miticides for crop protection. In: Novel Microbial Products for Medicine and Agriculture. A. L. Demain, G.A. Somkuti, J. C. Hunter-Cevera and H.W. Rossmoore [eds]. pp Chemical control 183

7 Dybas, R.A. and J. R. Babu (1988). 4"-deoxy-4"- methylamino-4-epiavermectin B1 hydrochloride (MK243): A novel avermectin insecticide for crop protection. Brighton Crop Protection Conference. pp Lasota, J.A. and R.A. Dybas (1991). Avermectins, a novel class of compounds: Implications for use in arthropod pest control. Annu. Rev. Entomol. 36: Tabashnik, B.E., N. L. Cushing, N. Finson and M. W. Johnson (199). Field development of resistance in Bacillus thuringiensis in diamondback moth (Lepidoptera: Plutellidae). J. Econ. Entomol. 83: Tabashnik, B.E., N. Finson, J. M. Schwartz, M. A. Caprio and M. W. Johnson (1992). Diamondback moth resistance to Bacillus thuringiensis in Hawaii. pp In: N.S. Talekar [ed.], Proceedings of the second international workshop on the management of diamondback moth and other crucifer pests. Asian Vegetable Research and Development Center, Tainan, Taiwan. Mau, R.F.L., L. Gusukuma-Minuto, and W. S. Ruggero (1995a). Field evaluation of bio-rational insecticides and combinations for controlling DBM, Arthropod Management Tests, 2: Mau, R.F.L., L. Gusukuma-Minuto, W. S. Ruggero and R. Shimabuku (1995b). Field evaluation of a neem and Bt product for DBM control, Arthropod Management Tests, 2: Mau, R.F.L., L. Gusukuma-Minuto, W. S. Ruggero and R. Shimabuku (1995c). Field evaluation of bio-rational insecticides for controlling DBM, Arthropod Management Tests, 2: Mau, R.F.L., L. Gusukuma-Minuto, and R. Shimabuku (1995d). Laboratory evaluations against bioinsecticides against DBM larvae, Arthropod Management Tests, 2: Proceedings: The Management of Diamondback Moth and Other Crucifer Pests