Potential of Nuclear Polyhedrosis Virus (NPV) and Bacillus thuringiensis (Bt) for Spodoptera Control in Yellow Granex Onion [Microbial Control of Spodoptera in Onion] Investigators: L.E. Padua ', A.C. Lapus ', V.P. Gapud 2, C.V. Pile 2, B.E. Santiago 2, N.S. Talekar 3, V.F. Recta 4, and E.G. Rajotte 4 Abstract Microbial field experiments for 1999 dry season saw the emergence of Helicoverpa armigera as a new pest of onion in addition to Spodoptera litura. Since the onion plants were also damaged by H. armigera, the larval counts of both species were combined. Larval densities on onion from the Bongabon Demo Farm showed lowest counts in plots treated with NPV alone and NPV+Bt, followed by Bt plots and Lambda-cyhalothrin (Karate) plots. Although the differences among these treatments did not appear significant, they are better than the untreated control plots. The same trend was also observed for the San Jose site, except that only Bt-treated plots had significantly lower larval counts compared to untreated control plots during the last week of sampling. The degree of onion leaf damage in Bongabon is the same for the treated and untreated control plots. However, NPV-treated plots in San Jose had the lowest leaf damage and were significantly different from both the Lambda-cyhalothrin and untreated control plots. Onion yields were highest in NPV-treated plots in both Bongabon and San Jose. In Bongabon, yield differences among the treatments were not significant, but they were better than the untreated control. NPV-treated plots in San Jose, however, differed significantly to the Lambda-cyhalothrin and untreated control plots. Objectives To mass produce NPV-CRSP for field trials in combination with Bt. To develop a better formulation for NPV. To evaluate the efficacy of NPV-CRSP against S. litura. To continue field trials using NPV-CRSP and Bt as microbial control agents against S. litura. IPM Constraints The absence of alternatives to insecticides in controlling S. litura in onions has left farmers with no option but to use insecticides. The study on microbials is expected to provide other options for the farmers in managing S. litura. Research Methods The wettable powder formulation of NPV-CRSP developed in 1998 at the Insect Pathology Laboratory in U.P. Los Banos was used for the 1999 dry season field experiments replacing the crude suspension. Mass rearing of S. litura in the laboratory was continued to provide for the mass production of NPV-CRSP needed for the experiments and bioassay studies. The concentration of the microbial treatments Bt alone, NPV alone and NPV+Bt was raised to 40 grams wettable powder per liter of water from the previous 20 grams/liter. Likewise, the sampling size was increased from 10 to 20 plants per plot with 4 replications per treatment. A shift from daytime sampling to night hours was also proposed to reflect the actual Spodoptera population. A farmer's field in Palestina, San Jose and a field in the Demo farm in Bongabon, Nueva Ecija were used to compare the effects of the microbial treatments and Lambda-cyhalothrin application (farmers' practice) on cutworm larval densities and onion yields. Each treatment was set up in RCBD using 4 replicated plots measuring 4m x 5m each. s were applied four times at bi-weekly intervals. Monitoring of larval populations was also done bi-weekly. 1 University of the Philippines Los Banos 2 Philippine Rice Research Institute 3 AVRDC Penn State University 350
ASIAN REGION The degree of onion leaf damage caused by larval feeding among the treatments was measured by taking the percent of leaves damaged per plant, every two weeks starting 4 weeks after transplanting. Onion yields were also recorded after harvest. Results NPV-CRSP was mass-produced in the laboratory. Out of the six kg wettable powder formulated with 6.79 x 10 8 polyhedral inclusion bodies (Pubs) per gram, 3.8 kilograms were used for the microbial field experiments. A 2-kg WP (in addition to 1 liter crude extract) was applied as a blanket spray to minimize the effects of insect damage on the weed and plant pathology field experiments. Currently, a total of 3 kg NPV crude extract have been produced for future field experiments and for bioassay studies on S. litura, S exigua and H. armigera, which will be the focus of next year's activities. Polyhedral inclusion bodies (PIBs) of 25 NPV-infected larvae were quantified/estimated with an average count of 3.61 x 10 8 PIBs per larva for the third instar. Estimation of PIBs for the 4 th and 5 th instar larvae will be done next year. Initial bracketing of NPV-CRSP for the first (1 - day old), second (3-day old) and third (6-day old) instar larvae of S. litura is near completion in preparation for the determination of the median lethal concentration (LC 50 ). Initial findings show significant larval mortalities on the fourth and fifth day of exposure of the insect to treated diet. Bioassay of the NPV Indian isolate and semi-purified NPV-CRSP will follow. The comparative effects of Bt product, NPV-CRSP crude extract and Lambda -cyhalothrin on S. litura + Helicoverpa larval populations from farmer's field in NOGROCOMA Demonstration Farm, Bongabon, Nueva Ecija are shown in Table 1. Since the onion plants were also damaged by H. armigera, the larval counts of both species were combined. Table 1. Effect of B. thuringiensis product, NPV-CRSP wettable powder and L. cyhalothrin (Karate) on larval count (S. litura + Helicoverpa) per plant. Bongabon, Nueva Ecija, 1998-99 Dry Season. Mean No. of Larvae per Plant (n=20 plants) 4 WAT 6 WAT 8 WAT 10 WAT 12 WAT Bt.02.01 u.14 u.29 u.24 u NPV.02.08 u.10 u.25 u.22 u NPV+Bt.01.01 u.12 u.29 u.24 u L. cyhalothrin (Karate).01.04 u.14 u.29 u.26 u Untreated Control.00.29.49.90.79 Signi.0004.0008.691.054.016.013.106.019.009.310.017.000.240.013.000 Cutworm larval densities remained relatively low during the 1999 season in Bongabon despite the increased sample size from 10 to 20 plants per plot and per treatment. Increasing the concentration of the microbial treatments to 20-40 g/l likewise showed no significant advantage of microbials over insecticide applications, yet they are comparable. One problem identified was the widely spaced sampling intervals for all treatments. As observed during one of the field visits, dead larvae were recovered from the treated plots 4 days after spraying. This was also confirmed from initial bioassay studies of NPV-CRSP where high larval mortality was recovered during the 4 th to 5 th day after exposure to treated diet. Furthermore, S. litura larvae were observed to be quite mobile, hence, larval movement from one plot to another was possible owing to the close distance (one meter border) between plots. The proposed night sampling did not materialize due to security reasons. However, univariate ANOVA applied on the results from each sampling date show that the larval counts from all treated plots are significantly lower than those from the untreated control plots from weeks 6 through 12. The same results were obtained for the Palestina site, except for the last week of sampling, when only Bt-treated plots had significantly lower larval counts compared to untreated control (Table 2). 351
Table 2. Effect of B. thuringiensis product, NPV-CRSP wettable powder and L. cyhalothrin (Karate) on larval count S. litura + Helicoverpa) per plant. San Jose, Nueva Ecija, Philippines, 1999 Dry Season. Mean No. of Larvae per Plant (n=20 pits) 4 WAT 6 WAT 8 WAT 10 WAT Bt.01.06 u.15 u.08 u NPV.02.19 u.04 u.14 NPV+Bt.01.11 u.11 u.15 Karate.00.26 u.34 u.15 Untreated Control.00.81 2.80.20.0003.0006.747.368.080.018 5.627 2.658.141 " Significantly different from untreated control, using LSD test at.05 level; WAT - weeks after transplanting.008.007.366 In Bongabon, the results of leaf damage from all five sampling dates showed no significant differences due to treatment. However, univariate ANOVA applied on the results from each sampling date showed that there were differences toward the later part of the season (Table 3). In San Jose, only NPV-treated plots were significantly different from the Lambda-cyhalothrin-treated and untreated control plots 8 weeks after transplanting (Table 4). Nevertheless, leaf damage levels for both sites were not impressive enough to show any substantial impact on onion yields. The great disparity in yields between San Jose and Bongabon sites appears to be influenced by other variables, e.g., soil properties, the cultivation techniques involving rice hull burning, planting density, and incidence of soil-borne diseases, particularly pink root, and root-knot nematodes. Onion yields were highest in NPV-treated plots in both the Bongabon Demo Farm and in San Jose. Although the yield differences among treatments in Bongabon were not significant, yields were better than in the untreated control plots. Yields from NPV-treated plots in San Jose, however, were significantly higher than those in the Lambda cyhalothrin and untreated control plots (Table 5). Table 3. Extent of leaf damage from five treatments observed over five sampling periods, Bongabon, Nueva Ecija, 1998-99 Dry Season. Mean percent leaf damage (n=20 plants) 4 6 8 10 12 Bt 0.85 11.65 14.70 18.25 20.30 NPV 0.32 16.42 16.48 14.15 u 18.22 u NPV+Bt 0.32 14.58 12.98 19.20 17.75 u Karate 0.42 17.45 16.78 17.50 19.82 Untreated Control 1.20 16.85 17.05 18.95 21.45 0.60 1.25 0.75 22.10 30.77 0.60 " Significantly different from untreated control, using LSD test at.05 level. 11.95 5.67 0.14 16.72 5.76 0.07 9.24 2.09 0.02 352
ASIAN REGION Table 4. Percent onion leaf damage from five treatments observed over four sampling periods. San Jose, Nueva Ecija, 1998-99 Dry Season. Weeks After Transplanting (WAT) 4 6 8 10 Signifi Bt 1.65 1.92 u 5.50 7.15 NPV 2.45 3.62 u 2.92 u k 5.60 NPV+Bt 0.98 2.92 u 4.10 5.55 Karate 2.22 3.40 u 5.72 5.95 Untreated Control 2.48 6.75 6.18 7.30 1.64 2.46 0.63 13.15 1.86 0.004 k Significantly different from Karate (chemical control), using LSD test at.05 level. 7.21 3.13 Table 5. Effect of B. thuringiensis product, NPV-CRSP wettable powder and Lambda cyhalothrin on onion yields (t/ha). Bongabon and San Jose, Nueva Ecija, 1999 Dry Season. Signific 0.12 Bongabon Site San Jose Site Bt 21.45 89.60 u NPV 26.25 u 97.50 u k NPV+Bt 25.78 u 87.90 Karate 24.92 u 80.32 Untreated Control 19.28 77.90 36.82 6.47 0.008 k Significantly different from Karate (chemical control), using LSD test at.05 level. 244.56 52.50 0.017 2.90 3.47 0.53 Impact It has been shown that NPV and Bt can be viable alternatives to chemical insecticide use. Thus, it would greatly benefit onion farmers who are dependent on chemical insecticides for control of cutworm. Direct effects are reduced pesticide use, better health of farmers, and sustainable Spodoptera management. In addition, farm-produced NPV will cut down on cost of crop protection by farmers. As a result, the market quality of their onion produce will be greatly enhanced by their low insecticide residue levels, thereby meeting the export requirements of foreign markets. Networking Activities Poster/Paper Presentations 1. A paper describing results of IPM-CRSP studies on Spodoptera management using microbials in rice-onion systems was presented at the annual conference of the Pest Management Council of the Philippines held in May 4-6, 1999 at PhiRice, Munoz, Nueva Ecija. 2. Presented a poster entitled: "Potential of Bt+NPV for the Control of Spodoptera on Onions in Central Luzon, Philippines," during the joint annual meeting of the Entomological Society of America and Phytopathological Society of America held November 8-12, 1998 at Hilton Hotel, Las Vegas, Nevada, USA. 353
Networking E. Rajotte traveled to PhilRice in October 1998 to evaluate research results and participate in planning for Year 7 experiments. Research Information Prepared three (3) fact sheets on the Pests of Onion and their Management for adoption by onion farmers. Publications and Presentations L.E. Padua, V.P. Gapud, E.C. Martin, C.V. Pile, B.A. Santiago, N.S. Talekar, G.F. Recta, E.G. Rajotte and A.C. Lapus. 1999. Use of Nuclear Polyhedrosis Virus (NPV) and Bacillus thuringiensis (Bt) for Spodoptera control in Yellow Granex Onions. Philippine Entomologist. (In Review). Presented IPM-CRSP Research/Technologies in collaboration with NOGROCOMA members on July 30, 1999 at PhilRice, Maligaya, Munoz, Nueva Ecija and participated in by NOGROCOMA members and Barangay Captains of Bongabon, Nueva Ecija. Training Output Took part in the training entitled, "Training for Agriculture Technicians and Farmer-Leaders on Rice-Vegetables with Emphasis on IPM" on August 2-6, 1999 at PhilRice, Maligaya, Munoz, Nueva Ecija. Project Highlights Six kilograms of NPV-CRSP wettable powder have been produced and were used for microbial field experiments and as blanket spray for the weed and plant pathology experiments. An additional three (3) kilos of NPV are being kept in cold storage for future use. Completed the final draft of three (3) fact sheets on Insect Pests of Onion (Common cutworm, S. litura; Onion armyworm, S. exigua and Onion leaf miner, Liriomyza sp.) and their Management. Quantified polyhedral inclusion bodies (PIBs) of twentyfive (25) NPV-infected 3' d instar larvae of S. litura with an average count of 3.61 x 10 8 PIBs per larva. H. armigera emerged as a new pest of onion in both Bongabon and Palestina sites for the 1999 dry season. The combined larval densities of S. litura and H. armigera did not differ among the treatments but were significantly different from the untreated control plots in both Bongabon and San Jose sites. Onion yields were highest in NPV-treated plots, for both Bongabon and San Jose, but not significantly different among the treatments in Bongabon. NPV-treated plots in San Jose differed significantly from Lambda-cyhalothrin and untreated control plots. Differences in cutworm larval densities, level of onion leaf damage, and onion yields between San Jose and Bongabon are attributed to planting density, soil type and the presence of other lepidopterous larvae, e.g. H. armigera. NPV alone and NPV+Bt are potential alternatives to insecticides for managing Spodoptera litura and H. armigera in onions.