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Optimization of pheromone dispenser density for managing the rice striped stem borer, Chilo suppressalis (Walker), by mating disruption
Crop Protection 28 (2009) 567–572
Contents lists available at ScienceDirect
Crop Protection journal homepage: www.elsevier.com/locate/cropro
Optimization of pheromone dispenser density for managing the rice striped stem borer, Chilo suppressalis (Walker), by mating disruption C. Alfaro*, V. Navarro-Llopis, J. Primo Centro de Ecologı´a Quı´mica Agrı´cola, Universidad Polite´cnica de Valencia, Camino de Vera s/n, 46022 Valencia, Spain
a r t i c l e i n f o
a b s t r a c t
Article history: Received 28 July 2008 Received in revised form 8 January 2009 Accepted 18 February 2009
The rice striped stem borer, Chilo suppressalis (Walker) (Lepidoptera: Pyralidae) is one of the most important rice pests worldwide. Rice is frequently grown in an intensive production system in areas adjacent to environmentally sensitive areas. Therefore, the use of insecticides is problematic and new techniques, including mating disruption, are being introduced. Due to the high cost of pheromones, it is essential to optimize the density of the pheromone dispensers. The main purpose of this research was to determine the minimal dosage and optimal dispenser distribution for effective mating disruption of C. suppressalis. To this end, we conducted a wide-area trial to test several dispenser densities, 31, 25 and 16 dispensers/ha during 3 years. Results were compared with a standard mating disruption treatment (51 dispensers/ha) and a standard aerial chemical treatment with tebufenozide. Treatment efficacy was determined by pheromone trap catches and crop damage assessment. The release rate of the pheromone dispensers was also quantified. The results of these trials suggest that such treatments provide effective pest control even with reduced pheromone dispenser densities. Longer-lasting dispensers with lower residual load at the end of trials will greatly improve the efficacy of mating disruption for C. suppressalis. Moreover, an important consideration is that these newly tested dispenser densities imply a significant reduction in the cost of the treatment. These results are essential in order to expand the use of mating disruption in rice crop protection. Ó 2009 Elsevier Ltd. All rights reserved.
Keywords: Pheromone dosage Lepidoptera Chilo suppressalis Mating disruption
1. Introduction Rice stem borers cause damage to the crop at the larval stages. Larvae bore into the plant stems and feed on plant nutrients causing, in many cases, severe crop loss (Beevor et al., 1990). Chilo and Scirpophaga spp. are difficult to control with foliar contact insecticides because these are only effective during a short period, the period between hatching and penetrating the plant stem (Nesbitt et al., 1975; Beevor et al., 1990; Howse et al., 1998). The rice stem borer, Chilo suppressalis (Walker), is one of the most important rice pests worldwide. It is particularly an economically important rice pest in some Asian countries, such as Japan, Korea and China. C. suppressalis also causes serious damage on rice in the Middle East and Spain (Casagrande, 1993). The sex pheromone of female C. suppressalis was initially identified in 1975 as a binary mixture of (Z)-11-hexadecenal (Z11– 16:Ald) and (Z)-13 octadecenal (Z13–18:Ald). The pheromone was present in female ovipositor extract (Nesbitt et al., 1975; Ohta et al., 1975, 1976). In 1983, another active compound of the sex
* Corresponding author. Tel.: þ34 96 387 9058; fax: þ34 96 387 9059. E-mail address: [email protected] (C. Alfaro). 0261-2194/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.cropro.2009.02.006
pheromone from C. suppressalis, (Z)-9-hexadecenal (Z9–16:Ald), was discovered (Tatsuki et al., 1983). Studies have showed that Z9– 16:Ald could significantly enhance the attractiveness to males when it was mixed with the binary mixture in the natural ratio. Therefore, it was concluded that the female sex pheromone of C. suppressalis was the three-component blend of Z11–16:Ald, Z13– 18:Ald and Z9–16:Ald in an approximate ratio of 48:6:5 (Tatsuki, 1990). The three-component blend with aldehyde-stabilizing agents has been the primary compounds for commercial development of controlled-release formulations (Howse, 1998). Several field trials with this pheromone formulation have been conducted for monitoring proposes or using as a mating disruption agent for managing the rice stem borer (Mochida et al., 1984; Tatsuki, 1990; Kondo and Tanaka, 1991; Casagrande, 1993; Kondo et al., 1993). Gaston and colleagues published in 1967 (Gaston et al., 1967) the first experimental confirmation that pre-mating communication between the sexes can be disrupted by permeating the atmosphere with an insect pheromone. This general phenomenon was demonstrated using Trichloplusia ni, cabbage looper, as the test organism. Since that time, the feasibility of interfering with mate finding by the broadcast release of synthetic pheromone has been demonstrated with innumerable moth species (Carde and Minks, 1995).
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C. Alfaro et al. / Crop Protection 28 (2009) 567–572
In Valencia, Spain, rice is grown in an environmentally protected area where application of chemical insecticides for managing rice pests could cause several environmental problems. Pest control using pheromones offers a good alternative to the chemical control methods. Past studies have shown that applying pheromone dispensers could completely disrupt the mating activity of C. suppressalis in the open field conditions. Mating disruption trials for controlling C. suppressalis was initiated in 1986 in Spain at a density of 2500 PVC dispensers/ha, each containing 8 mg of active ingredient. Mating disruption rates at this density were close to 100%. Several successive trials were conducted to optimize the application densities and methods. Based on the information generated from these studies, a density of 100 dispensers/ha, each containing 400 mg of active ingredient/dispenser was recommended for managing C. suppressalis (Casagrande, 1993; Howse et al., 1998). The Department of Agriculture of the government of Spain has also tested various dispenser densities and found that the dispenser densities could be further reduced without decreasing the disruption rates. In Spain, application of pheromone dispensers for mating disruption has become the primary method in management of C. suppressalis on rice. In 2003, the mating disruption technique was applied to 68.2% of the rice-growing area in Spain. In the pheromone control region, 52% of the area used a density of 69 dispensers/ha and 48% used 51 dispensers/ha. On the remaining area, chemical insecticides, tebufenozide, was applied for controlling the rice stem borer. The major limitation of the use of pheromone dispensers for controlling the rice stem borer is the relatively high costs compared to the chemical control. This study is design to test if the application costs of the mating disruption technique could be further reduced by optimizing the pheromone dosage and dispenser density. Our objective was to determine the minimum dosage and optimal dispenser distribution for effective mating disruption of C. suppressalis. We also studied the release profile of SELIBATE CSÔ dispensers to check if their pheromone release rate is enough to control the three generations of C. suppressalis that typically occur in Spain.
corresponding to the follow layout of the dispensers, 18 m 18 m, 20 m 20 m, 25 m 25 m, respectively. There were two control plots. One of them extended 1600 ha and was given the standard mating disruption treatment (SMDT), i.e. 51 dispensers/ha (14 m 14 m). The other plot (1900 ha) was given chemical treatment. Both control plots are around 5 km away from the trial area. The insecticide used was tebufenozide (24% w/w), 0.6 L/ha, applied in two aerial applications. The 95 ha trial field had never been treated with mating disruption pheromone. The SMDT plot had been treated with the mating disruption technique for nearly 10 years. The chemical treated area had been treated with insecticides since the 1960s. In the areas given mating disruption treatment, the dispensers were placed in field at the beginning of June. Table 1 shows a summary of the experimental design. 2.2. Evaluation of treatment efficacy Treatment efficacy was determined by (i) pheromone trap catches and (ii) crop damage assessment. The absence of trap catches during mating disruption treatment can be a good indication that the technique is effective, but crop damage assessment provides the ultimate proof (Howse, 1998).
2.1. Field trial location and layout
2.2.1. Pheromone trap catches Bucket funnel pheromone traps (Aragonesas Agro, SA, Madrid, Spain) were used for trapping C. suppressalis moths. The traps were baited with 7.5 mg of the pheromone lure ARALUREÔ Chilo (Aragonesas Agro, SA, Madrid, Spain) and DDVP Strip 20% dichlorvos (w/w) (Agrisense-BCS Ltd, Pontypridd, UK). In both control plots, standard mating disruption treatment and chemical treatment, pheromone traps were placed at the density of one trap per 10 ha at the beginning of the first flight of C. suppressalis adults, usually in the end of May. At the same time, nine pheromone traps were placed inside of each of the nine test plots. Pheromone trap catches were recorded weekly. The pheromone lures and DDVP strip were replaced every 6 weeks. Trap catches in treatments and control plots must be compared to verify the performance of the treatment. The absence of trap catches during mating disruption treatments in comparison with catches obtained in plots treated with insecticides is a good indication of the technique effectiveness.
The efficacy of commercial SELIBATE CSTM dispensers of C. suppressalis pheromone (Aragonesas Agro, SA, Madrid, Spain) was evaluated at different densities. Tests were conducted during the rice growing season from 2003 to 2005 on a 95 ha paddy rice field. This field bounded by citrus crop to the north, south and west, and by rice crop to the east. In order to reduce the impact of C. suppressalis moved from the neighboring eastern rice fields, a 10 ha buffer zone was treated with 100 dispensers/ha. The 95 ha field was divided into nine plots. In these plots, three different densities (31, 25 and 16 dispensers per ha) were tested with three replicates for each density. The dispenser densities
2.2.2. Crop damage assessment Crop damage assessment gives the ultimate proof to evaluate the efficacy of mating disruption technique. Crop damage was estimated by counting the number of infected plants per m2 (NIP/ m2) of rice field. Ten percent of each tested field surface and one percent of each control field surface were inspected. In each inspected plot, plants of 30 randomly selected 1 m2 areas were examined. The tillers of plants with symptoms were observed to verify that the damage was produced by rice stem borer. Plants were inspected at the end of the first, second and third moth flight generations.
2. Materials and methods
Table 1 Summary of treatments used during field experiments on C. suppressalis between 2003–2005. Test no.
Year
Area treated (ha)
Distance between dispensers (m)
Number dispensers/ha
Total pheromone dosage (g/ha)
Mean release (mg/day/ha)
CT SMDT Treatment 1 Treatment 2 Treatment 3
2003–2005 2003–2005 2003–2005 2003–2005 2004–2005
1900 1600 26.93 27.42 40.07
14 18 20 25
51 31 25 16
20.4 12.4 10 6.4
115.26 63.14 51.15 36.16
CT, chemical treatment; SMDT, standard mating disruption treatment.
C. Alfaro et al. / Crop Protection 28 (2009) 567–572
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Fig. 1. Moths per trap and day (MTD) of treatments 1, 2 and controls in 2003.
2.3. Pheromone release rates Release rates of pheromone dispensers were determined under field conditions. This was done by putting out 30 SELIBATE CSTM pheromone dispensers in the field and removing 5 of them at 0, 15, 30, 50, 60, 90 day intervals. Dispensers were stored at 18 C until they were analyzed. The residual pheromone amount of the dispensers was extracted and quantified by gas chromatography with flame ionization detector (GC/FID). The method used for extraction was pressurized solvent extraction using the One PSEÔ apparatus (Applied Separations, Bethlehem, PA, USA). Extraction conditions were 100 bar, 60 C, nine cycle extractions using dichloromethane as solvent. Each cycle lasted 5 min. The resulting extracts were concentrated under reduced pressure and analyzed by GC/FID. Residual pheromone analysis was made by GC/FID from dispenser extracts on the basis of Z11–16:Ald, the major C. suppressalis pheromone component. Gas chromatography and mass spectrometry (GC/MS) were used to confirm that the three components of C. suppressalis pheromone were released at the same rate by the dispenser. Z11–16:Ald was quantified by GC/FID
using hexadecane as internal standard. A ClarusÒ 500 gas chromatograph from PerkinElmer (Wellesley, MA, USA) was used for the analysis. All injections were made onto a ZebronÔ ZB-5MS (30 m 0.25 mm 0.25 mm) column (Phenomenex, Torrance, CA, USA) held at 100 C for 2 min and then programmed at 15 C/min to 170 C, held at 170 C for 5 min, and then at 20 C/min to 240 C and held at 240 C for 1 min. The carrier gas was helium at 1.2 ml/min with split flow value of 25 ml/min. 2.4. Statistical analysis Three pre-defined periods were established for statistical analysis corresponding with the three C. suppressalis flight generations. Under our weather conditions, the first generation of C. suppressalis take place throughout June, the second throughout July and the third from first of August until first of September. Trap catch data were transformed to sqrt(x), while crop damage data were transformed to log(x þ 1) scale to normalize the data. The transformed data were analyzed using one-way analysis of variance (ANOVA) followed by the LSD test to determine treatment differences.
Fig. 2. Moths per trap and day (MTD) of treatments 1, 2, 3 and controls in 2004.
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Fig. 3. Moths per trap and day (MTD) of treatments 1, 2, 3 and controls in 2005.
3. Results 3.1. Trap catches During the first year with mating disruption treatment, significant numbers of C. suppressalis moths were catches from pheromone treated and the chemical treated areas (Fig. 1). The trap catches were significantly reduced during the second and third years from the pheromone treated areas (Figs. 2 and 3), suggesting mating disruption treatments are more effective during the second and third years of the treatment. The cumulative trap catches during 2003 from SMDT field were least, followed by, in decreasing number, Treatment 1, Treatment 2 and chemical treatment (Table 2). The total moth catches in Treatment 1 with a dispenser density of 31 d/ha was 7.7, which was not significantly (P > 0.05) different compared to that from SMDT but it was significantly (P < 0.05) less than that from the chemical treated plots. During the second and third years, a significant
number (>15 moths) of C. suppressalis moths were captured by pheromone traps from the chemical treated area. The total moth catches from SMDT plots and the three pheromone treatments (Treatments 1, 2, and 3) during the second and third years were low (<2 moths) and did not differ significantly among these four treatments. The results were very consistent across insect generations in the two years. 3.2. Crop damage Crop damage assessment is the most evident way for showing the efficiency of mating disruption (Karg and Sauer, 1995; Howse, 1998). In the first year, there were generally no significant differences in the damage levels among treatments (including chemical control treatment) (Table 3). The only significant difference was the damaged observed from chemical treatment and Treatment 2, the lowest dispenser density (25 dispensers/ha). However, the difference was small, <0.6 plants/m2 and both damage rates were well
Fig. 4. Residual pheromones of the SELIBATE CSÔ dispenser.
9.62 2.76 b 1.20 0.58 a 1.10 1.10 a 0.66 0.66 a 0.66 0.33 a
3.3. Pheromone release rates Pheromone was released throughout the field trial, but the release rate decreased after 50 days. However, field trials indicated that the reduced emission rates of the pheromone were still high enough to produce good mating disruption in the field. The residual amount of pheromone in the dispenser at the end of the season was approximately 30% of the initial dosage, suggesting unnecessary amount of pheromone was presented in the dispensers (Fig. 4). The residual amount of unreleased pheromone could be minimized in order to reduce the cost of the mating disruption technique for managing the stem borer pest. 4. Discussion
Treatments followed by the same letter are not significantly different, P > 0.05 (ANOVA).
Total 3rd 2nd
1.87 0.47 b 0.35 0.16 a 0.66 0.66 ab 0.33 0.33 a 0.33 0.33 a 18 3.38 b 1.16 0.28 a 0.00 0.00 a 1.00 0.57 a 1.33 0.88 a
4.00 1.55 b 0.35 0.15 a 0.00 0.00 a 0.00 0.00 a 0.00 0.00 a
1st Total
1.06 0.63 b 0.27 0.13 a 0.00 0.00 a 0.00 0.00 a 0.33 0.33 a 0.60 0.34 a 0.16 0.12 a 0.00 0.00 a 0.00 0.00 a 0.33 0.33 a
571
below the treatment threshold prescribed by the Spanish Ministry of Agriculture for 3 infected plants per m2. In 2004, no first-generation damage was observed in any of the areas evaluated. This corresponded with the low number of moth catches by the pheromone traps during the first generation (Fig. 2). There were also no significant differences in the damage from the second generation stem borer (Table 3). During the third generation, the most damage was recorded in the plots treated with tebufenozide, which was significantly greater than those observed from the pheromone treated areas (Table 3). Results from the 2005 field trial showed that there were generally no significant differences in plant damage among the five treatments during the first and second generations, while plots of the chemical control and SMDT appeared to have a less number of plants damaged by the stem borer. However, plant damage caused by the third generation may not be economically important, because the grain is already formed and grain yield will be not significantly affected if the damage is not too several (Poitout and Bues, 1978).
15.8 3.01 b 0.72 0.25 a 0.00 0.00 a 1.00 0.57 a 0.66 0.33 a
2nd 1st Total 3rd
31.27 7.39 c 0.88 0.38 a 5.00 3.60 ab 11.66 1.20 bc 10.63 4.17 b 0.00 0.00 a 0.33 0.33 a 2.33 1.20 ab 1.09 0.41 ab 0.28 0.16 a 2.33 1.20 bc 4.33 2.33 c CT SMDT (51 d/ha) Treatment 1 (31 d/ha) Treatment 2 (25 d/ha) Treatment 3 (16 d/ha)
2nd
43.00 10.09 c 1.16 0.43 a 7.66 4.66 ab 18.33 4.09 bc
1st
3rd
Generations 2005 Generations 2004 Generations 2003 Treatments
Table 2 Average and standard error of cumulative trap catches in tests and controls per generation and total in each year.
15.50 3.44 b 1.9 0.61a 1.66 0.88 a 1.00 0.57 a 1.00 0.57 a
C. Alfaro et al. / Crop Protection 28 (2009) 567–572
Chemical rice crop protection with tebufenozide relies on killing insects through contact at its most vulnerable stage of development. For most lepidopteran pests such as rice stem borers, this means the first instar larvae, which bore into the stems immediately after hatching and thus offer a very short period of control opportunity (Howse, 1998). With the mating disruption technology, the timing of the application is still important but not so critical as for the chemical control. The application of pheromone for managing stem borers also provides farmers with protection for their crops that is season-long, userand environment-friendly. In addition, chemical treatments require a safety period of approximately 20 days free from application before harvesting. This prevents treating the third generation of the moth, or even the second generation when the fly period is delayed, which occurred in the second year of our field trials. Because rice is frequently grown in the vicinity of environmentally protected areas, it is essential to dispose of an effective yet environment-friendly technique to control C. suppressalis. However, the environmental cost of chemical treatments is usually not included in calculation of treatment costs. Consequently cost has been seen the main obstacle in adoption of new environmentally friendly techniques. On the other hand, application of mating disruption for managing rice stem borer is expensive for farmers because pheromone market is still developing and dispenser technology is not optimized. Results of this study showed that treatments with reduced pheromone dispenser densities also provided efficient pest control, suggesting application costs using mating disruption could be significantly reduced. For example, in Spain the price of the standard
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Table 3 Average and standard error of crop damages (NIP/m2) in tests and controls per generation in each year. Treatments
CT SMDT (51 d/ha) Treatment 1 (31 d/ha) Treatment 2 (25 d/ha) Treatment 3 (16 d/ha)
Generations 2003
Generations 2004
Generations 2005
1st
2nd
3rd
2nd
3rd
1st
2nd
3rd
0.22 0.05 a 0.13 0.06 ab 0.00 0.00 b 0.13 0.13 ab
0.09 0.03 a 0.04 0.02 a 0.03 0.00 a 0.04 0.03 a
0.26 0.06 a 0.40 0.07 ab 0.45 0.06 ab 0.79 0.21 b
0.06 0.02 a 0.03 0.03 a 0.00 0.00 a 0.13 0.13 a 0.00 0.00 a
1.54 0.23 a 0.41 0.17 b 0.00 0.00 c 0.16 0.16 bc 0.17 0.10 bc
0.08 0.08 a 0.00 0.00 a 0.24þ0.24 a 0.38 0.22 a 0.33 0.18 a
0.05 0.03 ab 0.06 0.006 a 0.14 0.14 ab 0.33 0.18 b 0.03 0.02 ab
0.21 0.08 a 0.16 0.06 ab 2.15 1.36 c 1.15 0.82 bc 0.89 0.24 c
Treatments followed by the same letter are not significantly different, P > 0.05 (ANOVA).
mating disruption treatment (51 dispensers/ha) is 66 V/ha. If the number of dispensers is reduced to 31, 25 or 16 dispensers per ha, the cost of the treatment will be 43, 36 and 24 V/ha respectively (Ministry of Agriculture, personal communication). Treatment against C. suppressalis with these new densities will be cheaper than the two chemical treatments needed per year, whose cost was approximately 47 V/ha. This reduction in the cost of the treatment will be important for the implement of the mating disruption technique in the whole rice crop area in Valencia. The results of this research prove clearly that a mating disruption treatment using 31, 25 and 16 dispensers/ha is capable of providing protection against C. suppressalis for almost three years. Although it is possible that after several years of treatment with a low density of dispensers the pest populations would gradually increase. Under this circumstance, the financial saving from the first a few years should be enough to justify switching to use the mating disruption technology from chemical control method. Our results also showed that the rice stem borer population reduced progressively after mating disruption was applied suggested that several successive years of mating disruption treatment are needed to achieve optimal control (Coscolla, 1997). The study of dispenser release rates showed decreased emission during the end of each season. This might explain the loss of efficacy in the third stem borer generation even though the dispensers still contained 30–40% of the pheromone load. An improved dispenser design would make it possible to achieve even better results in moth control with the same quantity of pheromone, and thus contribute to reducing the cost of the treatment even more.
Acknowledgements This project was funded by ‘‘Fundacio´n Jose´ y Ana Royo’’. We would like to thank Fernando Alfaro Lassala for technical support and CAPA-GVA for providing trial fields.
References Beevor, P.S., David, H., Jones, O.T., 1990. Female sex pheromones of Chilo spp. (Lepidoptera: Pyralidae) and their development in pest control applications. Insect Sci. Appl. 11, 787–794. Carde, R.T., Minks, A.K., 1995. Control of moth pests by mating disruptiondsuccesses and constraints. Annu. Rev. Entomol 40, 559–585. Casagrande, E., 1993. The commercial implementation of mating disruption for the control of the rice stem borer, Chilo suppressalis, in rice in Spain. IOBC WPRS Bulletin, 82–89. Coscolla, R., 1997. La polilla del racimo de la vid (Lobesia botrana Den. and Schiff). Generalitat Valenciana, Valencia. Gaston, L.K., Shorey, H.H., Saario, C.A., 1967. Insect population control by use of sex pheromones to inhibit orientation between sexes. Nature 213, 1155. Howse, P.E., 1998. Pheromones and behaviour. In: Howse, P.E., Stevens, I., Jones, O.T. (Eds.), Insect Pheromones and their Use in Pest Management. Chapman & Hall, London 1–130. Howse, P.E., Stevens, I., Jones, O.T., 1998. Insect pests of rice. In: Howse, P.E., Stevens, I., Jones, O.T. (Eds.), Insect Pheromones and their Use in Pest Management. Chapman & Hall, London, pp. 325–330. Karg, G., Sauer, A.E., 1995. Spatial-distribution of pheromone in vineyards treated for mating disruption of the grape vine moth Lobesia botrana measured with electroantennograms. J. Chem. Ecol 21, 1299–1314. Kondo, A., Tanaka, F., 1991. Pheromone trap catches of the rice stem borer moth, Chilo suppressalis (Walker) (Lepidoptera, Pyralidae) and related trap variables in the field. Appl. Entomol. Zool 26, 167–172. Kondo, A., Tanaka, F., Sugie, H., Hokyo, N., 1993. Analysis of some biological factors affecting differential pheromone trap efficiency between generations in the rice stem borer moth, Chilo suppressalis (Walker) (Lepidoptera, Pyralidae). Appl. Entomol. Zool 28, 503–511. Mochida, O., Arida, G.S., Tatsuki, S., Fukami, J., 1984. A field-test on a 3rd component of the female sex-pheromone of the rice stem borer moth, Chilo suppressalis, in the Philippines. Entomol. Exp. Appl. 36, 295–296. Nesbitt, B.F., Beevor, P.S., Hall, D.R., Lester, R., Dyck, V.A., 1975. Identification of female sex pheromones of moth, Chilo suppressalis. J. Insect Physiol 21, 1883–1886. Ohta, K., Tatsuki, S., Uchiumi, K., Kurihara, M., Fukami, J., 1975. Sex-pheromone of rice stem borerdpurification and chemical properties. Agric. Biol. Chem. 39, 2437–2438. Ohta, K., Tatsuki, S., Uchiumi, K., Kurihara, M., Fukami, J., 1976. Structures of sexpheromones of rice stem borer. Agric. Biol. Chem. 40, 1897–1899. Poitout, S., Bues, R., 1978. Life-cycle of rice stem borer Chilo suppressalis (Walker) in rhone delta (Camargue)duse of sex trap. Ann. Zool. Ecol. Anim 10, 245–265. Tatsuki, S., Kurihara, M., Usui, K., Ohguchi, Y., Uchiumi, K., Fukami, J., Arai, K., Yabuki, S., Tanaka, F., 1983. Sex-pheromone of the rice stem borer Chilo suppressalis (Walker) (Lepidoptera, Pyralidae), the 3rd component. Z-9-Hexadecenal. Appl. Entomol. Zool 18, 443–446. Tatsuki, S., 1990. Status of application of sex pheromone of rice stem borer moth, Chilo supressalis in Japan. Insect Sci. Appl. 11, 807–812.
Contents lists available at ScienceDirect
Crop Protection journal homepage: www.elsevier.com/locate/cropro
Optimization of pheromone dispenser density for managing the rice striped stem borer, Chilo suppressalis (Walker), by mating disruption C. Alfaro*, V. Navarro-Llopis, J. Primo Centro de Ecologı´a Quı´mica Agrı´cola, Universidad Polite´cnica de Valencia, Camino de Vera s/n, 46022 Valencia, Spain
a r t i c l e i n f o
a b s t r a c t
Article history: Received 28 July 2008 Received in revised form 8 January 2009 Accepted 18 February 2009
The rice striped stem borer, Chilo suppressalis (Walker) (Lepidoptera: Pyralidae) is one of the most important rice pests worldwide. Rice is frequently grown in an intensive production system in areas adjacent to environmentally sensitive areas. Therefore, the use of insecticides is problematic and new techniques, including mating disruption, are being introduced. Due to the high cost of pheromones, it is essential to optimize the density of the pheromone dispensers. The main purpose of this research was to determine the minimal dosage and optimal dispenser distribution for effective mating disruption of C. suppressalis. To this end, we conducted a wide-area trial to test several dispenser densities, 31, 25 and 16 dispensers/ha during 3 years. Results were compared with a standard mating disruption treatment (51 dispensers/ha) and a standard aerial chemical treatment with tebufenozide. Treatment efficacy was determined by pheromone trap catches and crop damage assessment. The release rate of the pheromone dispensers was also quantified. The results of these trials suggest that such treatments provide effective pest control even with reduced pheromone dispenser densities. Longer-lasting dispensers with lower residual load at the end of trials will greatly improve the efficacy of mating disruption for C. suppressalis. Moreover, an important consideration is that these newly tested dispenser densities imply a significant reduction in the cost of the treatment. These results are essential in order to expand the use of mating disruption in rice crop protection. Ó 2009 Elsevier Ltd. All rights reserved.
Keywords: Pheromone dosage Lepidoptera Chilo suppressalis Mating disruption
1. Introduction Rice stem borers cause damage to the crop at the larval stages. Larvae bore into the plant stems and feed on plant nutrients causing, in many cases, severe crop loss (Beevor et al., 1990). Chilo and Scirpophaga spp. are difficult to control with foliar contact insecticides because these are only effective during a short period, the period between hatching and penetrating the plant stem (Nesbitt et al., 1975; Beevor et al., 1990; Howse et al., 1998). The rice stem borer, Chilo suppressalis (Walker), is one of the most important rice pests worldwide. It is particularly an economically important rice pest in some Asian countries, such as Japan, Korea and China. C. suppressalis also causes serious damage on rice in the Middle East and Spain (Casagrande, 1993). The sex pheromone of female C. suppressalis was initially identified in 1975 as a binary mixture of (Z)-11-hexadecenal (Z11– 16:Ald) and (Z)-13 octadecenal (Z13–18:Ald). The pheromone was present in female ovipositor extract (Nesbitt et al., 1975; Ohta et al., 1975, 1976). In 1983, another active compound of the sex
* Corresponding author. Tel.: þ34 96 387 9058; fax: þ34 96 387 9059. E-mail address: [email protected] (C. Alfaro). 0261-2194/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.cropro.2009.02.006
pheromone from C. suppressalis, (Z)-9-hexadecenal (Z9–16:Ald), was discovered (Tatsuki et al., 1983). Studies have showed that Z9– 16:Ald could significantly enhance the attractiveness to males when it was mixed with the binary mixture in the natural ratio. Therefore, it was concluded that the female sex pheromone of C. suppressalis was the three-component blend of Z11–16:Ald, Z13– 18:Ald and Z9–16:Ald in an approximate ratio of 48:6:5 (Tatsuki, 1990). The three-component blend with aldehyde-stabilizing agents has been the primary compounds for commercial development of controlled-release formulations (Howse, 1998). Several field trials with this pheromone formulation have been conducted for monitoring proposes or using as a mating disruption agent for managing the rice stem borer (Mochida et al., 1984; Tatsuki, 1990; Kondo and Tanaka, 1991; Casagrande, 1993; Kondo et al., 1993). Gaston and colleagues published in 1967 (Gaston et al., 1967) the first experimental confirmation that pre-mating communication between the sexes can be disrupted by permeating the atmosphere with an insect pheromone. This general phenomenon was demonstrated using Trichloplusia ni, cabbage looper, as the test organism. Since that time, the feasibility of interfering with mate finding by the broadcast release of synthetic pheromone has been demonstrated with innumerable moth species (Carde and Minks, 1995).
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In Valencia, Spain, rice is grown in an environmentally protected area where application of chemical insecticides for managing rice pests could cause several environmental problems. Pest control using pheromones offers a good alternative to the chemical control methods. Past studies have shown that applying pheromone dispensers could completely disrupt the mating activity of C. suppressalis in the open field conditions. Mating disruption trials for controlling C. suppressalis was initiated in 1986 in Spain at a density of 2500 PVC dispensers/ha, each containing 8 mg of active ingredient. Mating disruption rates at this density were close to 100%. Several successive trials were conducted to optimize the application densities and methods. Based on the information generated from these studies, a density of 100 dispensers/ha, each containing 400 mg of active ingredient/dispenser was recommended for managing C. suppressalis (Casagrande, 1993; Howse et al., 1998). The Department of Agriculture of the government of Spain has also tested various dispenser densities and found that the dispenser densities could be further reduced without decreasing the disruption rates. In Spain, application of pheromone dispensers for mating disruption has become the primary method in management of C. suppressalis on rice. In 2003, the mating disruption technique was applied to 68.2% of the rice-growing area in Spain. In the pheromone control region, 52% of the area used a density of 69 dispensers/ha and 48% used 51 dispensers/ha. On the remaining area, chemical insecticides, tebufenozide, was applied for controlling the rice stem borer. The major limitation of the use of pheromone dispensers for controlling the rice stem borer is the relatively high costs compared to the chemical control. This study is design to test if the application costs of the mating disruption technique could be further reduced by optimizing the pheromone dosage and dispenser density. Our objective was to determine the minimum dosage and optimal dispenser distribution for effective mating disruption of C. suppressalis. We also studied the release profile of SELIBATE CSÔ dispensers to check if their pheromone release rate is enough to control the three generations of C. suppressalis that typically occur in Spain.
corresponding to the follow layout of the dispensers, 18 m 18 m, 20 m 20 m, 25 m 25 m, respectively. There were two control plots. One of them extended 1600 ha and was given the standard mating disruption treatment (SMDT), i.e. 51 dispensers/ha (14 m 14 m). The other plot (1900 ha) was given chemical treatment. Both control plots are around 5 km away from the trial area. The insecticide used was tebufenozide (24% w/w), 0.6 L/ha, applied in two aerial applications. The 95 ha trial field had never been treated with mating disruption pheromone. The SMDT plot had been treated with the mating disruption technique for nearly 10 years. The chemical treated area had been treated with insecticides since the 1960s. In the areas given mating disruption treatment, the dispensers were placed in field at the beginning of June. Table 1 shows a summary of the experimental design. 2.2. Evaluation of treatment efficacy Treatment efficacy was determined by (i) pheromone trap catches and (ii) crop damage assessment. The absence of trap catches during mating disruption treatment can be a good indication that the technique is effective, but crop damage assessment provides the ultimate proof (Howse, 1998).
2.1. Field trial location and layout
2.2.1. Pheromone trap catches Bucket funnel pheromone traps (Aragonesas Agro, SA, Madrid, Spain) were used for trapping C. suppressalis moths. The traps were baited with 7.5 mg of the pheromone lure ARALUREÔ Chilo (Aragonesas Agro, SA, Madrid, Spain) and DDVP Strip 20% dichlorvos (w/w) (Agrisense-BCS Ltd, Pontypridd, UK). In both control plots, standard mating disruption treatment and chemical treatment, pheromone traps were placed at the density of one trap per 10 ha at the beginning of the first flight of C. suppressalis adults, usually in the end of May. At the same time, nine pheromone traps were placed inside of each of the nine test plots. Pheromone trap catches were recorded weekly. The pheromone lures and DDVP strip were replaced every 6 weeks. Trap catches in treatments and control plots must be compared to verify the performance of the treatment. The absence of trap catches during mating disruption treatments in comparison with catches obtained in plots treated with insecticides is a good indication of the technique effectiveness.
The efficacy of commercial SELIBATE CSTM dispensers of C. suppressalis pheromone (Aragonesas Agro, SA, Madrid, Spain) was evaluated at different densities. Tests were conducted during the rice growing season from 2003 to 2005 on a 95 ha paddy rice field. This field bounded by citrus crop to the north, south and west, and by rice crop to the east. In order to reduce the impact of C. suppressalis moved from the neighboring eastern rice fields, a 10 ha buffer zone was treated with 100 dispensers/ha. The 95 ha field was divided into nine plots. In these plots, three different densities (31, 25 and 16 dispensers per ha) were tested with three replicates for each density. The dispenser densities
2.2.2. Crop damage assessment Crop damage assessment gives the ultimate proof to evaluate the efficacy of mating disruption technique. Crop damage was estimated by counting the number of infected plants per m2 (NIP/ m2) of rice field. Ten percent of each tested field surface and one percent of each control field surface were inspected. In each inspected plot, plants of 30 randomly selected 1 m2 areas were examined. The tillers of plants with symptoms were observed to verify that the damage was produced by rice stem borer. Plants were inspected at the end of the first, second and third moth flight generations.
2. Materials and methods
Table 1 Summary of treatments used during field experiments on C. suppressalis between 2003–2005. Test no.
Year
Area treated (ha)
Distance between dispensers (m)
Number dispensers/ha
Total pheromone dosage (g/ha)
Mean release (mg/day/ha)
CT SMDT Treatment 1 Treatment 2 Treatment 3
2003–2005 2003–2005 2003–2005 2003–2005 2004–2005
1900 1600 26.93 27.42 40.07
14 18 20 25
51 31 25 16
20.4 12.4 10 6.4
115.26 63.14 51.15 36.16
CT, chemical treatment; SMDT, standard mating disruption treatment.
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Fig. 1. Moths per trap and day (MTD) of treatments 1, 2 and controls in 2003.
2.3. Pheromone release rates Release rates of pheromone dispensers were determined under field conditions. This was done by putting out 30 SELIBATE CSTM pheromone dispensers in the field and removing 5 of them at 0, 15, 30, 50, 60, 90 day intervals. Dispensers were stored at 18 C until they were analyzed. The residual pheromone amount of the dispensers was extracted and quantified by gas chromatography with flame ionization detector (GC/FID). The method used for extraction was pressurized solvent extraction using the One PSEÔ apparatus (Applied Separations, Bethlehem, PA, USA). Extraction conditions were 100 bar, 60 C, nine cycle extractions using dichloromethane as solvent. Each cycle lasted 5 min. The resulting extracts were concentrated under reduced pressure and analyzed by GC/FID. Residual pheromone analysis was made by GC/FID from dispenser extracts on the basis of Z11–16:Ald, the major C. suppressalis pheromone component. Gas chromatography and mass spectrometry (GC/MS) were used to confirm that the three components of C. suppressalis pheromone were released at the same rate by the dispenser. Z11–16:Ald was quantified by GC/FID
using hexadecane as internal standard. A ClarusÒ 500 gas chromatograph from PerkinElmer (Wellesley, MA, USA) was used for the analysis. All injections were made onto a ZebronÔ ZB-5MS (30 m 0.25 mm 0.25 mm) column (Phenomenex, Torrance, CA, USA) held at 100 C for 2 min and then programmed at 15 C/min to 170 C, held at 170 C for 5 min, and then at 20 C/min to 240 C and held at 240 C for 1 min. The carrier gas was helium at 1.2 ml/min with split flow value of 25 ml/min. 2.4. Statistical analysis Three pre-defined periods were established for statistical analysis corresponding with the three C. suppressalis flight generations. Under our weather conditions, the first generation of C. suppressalis take place throughout June, the second throughout July and the third from first of August until first of September. Trap catch data were transformed to sqrt(x), while crop damage data were transformed to log(x þ 1) scale to normalize the data. The transformed data were analyzed using one-way analysis of variance (ANOVA) followed by the LSD test to determine treatment differences.
Fig. 2. Moths per trap and day (MTD) of treatments 1, 2, 3 and controls in 2004.
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Fig. 3. Moths per trap and day (MTD) of treatments 1, 2, 3 and controls in 2005.
3. Results 3.1. Trap catches During the first year with mating disruption treatment, significant numbers of C. suppressalis moths were catches from pheromone treated and the chemical treated areas (Fig. 1). The trap catches were significantly reduced during the second and third years from the pheromone treated areas (Figs. 2 and 3), suggesting mating disruption treatments are more effective during the second and third years of the treatment. The cumulative trap catches during 2003 from SMDT field were least, followed by, in decreasing number, Treatment 1, Treatment 2 and chemical treatment (Table 2). The total moth catches in Treatment 1 with a dispenser density of 31 d/ha was 7.7, which was not significantly (P > 0.05) different compared to that from SMDT but it was significantly (P < 0.05) less than that from the chemical treated plots. During the second and third years, a significant
number (>15 moths) of C. suppressalis moths were captured by pheromone traps from the chemical treated area. The total moth catches from SMDT plots and the three pheromone treatments (Treatments 1, 2, and 3) during the second and third years were low (<2 moths) and did not differ significantly among these four treatments. The results were very consistent across insect generations in the two years. 3.2. Crop damage Crop damage assessment is the most evident way for showing the efficiency of mating disruption (Karg and Sauer, 1995; Howse, 1998). In the first year, there were generally no significant differences in the damage levels among treatments (including chemical control treatment) (Table 3). The only significant difference was the damaged observed from chemical treatment and Treatment 2, the lowest dispenser density (25 dispensers/ha). However, the difference was small, <0.6 plants/m2 and both damage rates were well
Fig. 4. Residual pheromones of the SELIBATE CSÔ dispenser.
9.62 2.76 b 1.20 0.58 a 1.10 1.10 a 0.66 0.66 a 0.66 0.33 a
3.3. Pheromone release rates Pheromone was released throughout the field trial, but the release rate decreased after 50 days. However, field trials indicated that the reduced emission rates of the pheromone were still high enough to produce good mating disruption in the field. The residual amount of pheromone in the dispenser at the end of the season was approximately 30% of the initial dosage, suggesting unnecessary amount of pheromone was presented in the dispensers (Fig. 4). The residual amount of unreleased pheromone could be minimized in order to reduce the cost of the mating disruption technique for managing the stem borer pest. 4. Discussion
Treatments followed by the same letter are not significantly different, P > 0.05 (ANOVA).
Total 3rd 2nd
1.87 0.47 b 0.35 0.16 a 0.66 0.66 ab 0.33 0.33 a 0.33 0.33 a 18 3.38 b 1.16 0.28 a 0.00 0.00 a 1.00 0.57 a 1.33 0.88 a
4.00 1.55 b 0.35 0.15 a 0.00 0.00 a 0.00 0.00 a 0.00 0.00 a
1st Total
1.06 0.63 b 0.27 0.13 a 0.00 0.00 a 0.00 0.00 a 0.33 0.33 a 0.60 0.34 a 0.16 0.12 a 0.00 0.00 a 0.00 0.00 a 0.33 0.33 a
571
below the treatment threshold prescribed by the Spanish Ministry of Agriculture for 3 infected plants per m2. In 2004, no first-generation damage was observed in any of the areas evaluated. This corresponded with the low number of moth catches by the pheromone traps during the first generation (Fig. 2). There were also no significant differences in the damage from the second generation stem borer (Table 3). During the third generation, the most damage was recorded in the plots treated with tebufenozide, which was significantly greater than those observed from the pheromone treated areas (Table 3). Results from the 2005 field trial showed that there were generally no significant differences in plant damage among the five treatments during the first and second generations, while plots of the chemical control and SMDT appeared to have a less number of plants damaged by the stem borer. However, plant damage caused by the third generation may not be economically important, because the grain is already formed and grain yield will be not significantly affected if the damage is not too several (Poitout and Bues, 1978).
15.8 3.01 b 0.72 0.25 a 0.00 0.00 a 1.00 0.57 a 0.66 0.33 a
2nd 1st Total 3rd
31.27 7.39 c 0.88 0.38 a 5.00 3.60 ab 11.66 1.20 bc 10.63 4.17 b 0.00 0.00 a 0.33 0.33 a 2.33 1.20 ab 1.09 0.41 ab 0.28 0.16 a 2.33 1.20 bc 4.33 2.33 c CT SMDT (51 d/ha) Treatment 1 (31 d/ha) Treatment 2 (25 d/ha) Treatment 3 (16 d/ha)
2nd
43.00 10.09 c 1.16 0.43 a 7.66 4.66 ab 18.33 4.09 bc
1st
3rd
Generations 2005 Generations 2004 Generations 2003 Treatments
Table 2 Average and standard error of cumulative trap catches in tests and controls per generation and total in each year.
15.50 3.44 b 1.9 0.61a 1.66 0.88 a 1.00 0.57 a 1.00 0.57 a
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Chemical rice crop protection with tebufenozide relies on killing insects through contact at its most vulnerable stage of development. For most lepidopteran pests such as rice stem borers, this means the first instar larvae, which bore into the stems immediately after hatching and thus offer a very short period of control opportunity (Howse, 1998). With the mating disruption technology, the timing of the application is still important but not so critical as for the chemical control. The application of pheromone for managing stem borers also provides farmers with protection for their crops that is season-long, userand environment-friendly. In addition, chemical treatments require a safety period of approximately 20 days free from application before harvesting. This prevents treating the third generation of the moth, or even the second generation when the fly period is delayed, which occurred in the second year of our field trials. Because rice is frequently grown in the vicinity of environmentally protected areas, it is essential to dispose of an effective yet environment-friendly technique to control C. suppressalis. However, the environmental cost of chemical treatments is usually not included in calculation of treatment costs. Consequently cost has been seen the main obstacle in adoption of new environmentally friendly techniques. On the other hand, application of mating disruption for managing rice stem borer is expensive for farmers because pheromone market is still developing and dispenser technology is not optimized. Results of this study showed that treatments with reduced pheromone dispenser densities also provided efficient pest control, suggesting application costs using mating disruption could be significantly reduced. For example, in Spain the price of the standard
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Table 3 Average and standard error of crop damages (NIP/m2) in tests and controls per generation in each year. Treatments
CT SMDT (51 d/ha) Treatment 1 (31 d/ha) Treatment 2 (25 d/ha) Treatment 3 (16 d/ha)
Generations 2003
Generations 2004
Generations 2005
1st
2nd
3rd
2nd
3rd
1st
2nd
3rd
0.22 0.05 a 0.13 0.06 ab 0.00 0.00 b 0.13 0.13 ab
0.09 0.03 a 0.04 0.02 a 0.03 0.00 a 0.04 0.03 a
0.26 0.06 a 0.40 0.07 ab 0.45 0.06 ab 0.79 0.21 b
0.06 0.02 a 0.03 0.03 a 0.00 0.00 a 0.13 0.13 a 0.00 0.00 a
1.54 0.23 a 0.41 0.17 b 0.00 0.00 c 0.16 0.16 bc 0.17 0.10 bc
0.08 0.08 a 0.00 0.00 a 0.24þ0.24 a 0.38 0.22 a 0.33 0.18 a
0.05 0.03 ab 0.06 0.006 a 0.14 0.14 ab 0.33 0.18 b 0.03 0.02 ab
0.21 0.08 a 0.16 0.06 ab 2.15 1.36 c 1.15 0.82 bc 0.89 0.24 c
Treatments followed by the same letter are not significantly different, P > 0.05 (ANOVA).
mating disruption treatment (51 dispensers/ha) is 66 V/ha. If the number of dispensers is reduced to 31, 25 or 16 dispensers per ha, the cost of the treatment will be 43, 36 and 24 V/ha respectively (Ministry of Agriculture, personal communication). Treatment against C. suppressalis with these new densities will be cheaper than the two chemical treatments needed per year, whose cost was approximately 47 V/ha. This reduction in the cost of the treatment will be important for the implement of the mating disruption technique in the whole rice crop area in Valencia. The results of this research prove clearly that a mating disruption treatment using 31, 25 and 16 dispensers/ha is capable of providing protection against C. suppressalis for almost three years. Although it is possible that after several years of treatment with a low density of dispensers the pest populations would gradually increase. Under this circumstance, the financial saving from the first a few years should be enough to justify switching to use the mating disruption technology from chemical control method. Our results also showed that the rice stem borer population reduced progressively after mating disruption was applied suggested that several successive years of mating disruption treatment are needed to achieve optimal control (Coscolla, 1997). The study of dispenser release rates showed decreased emission during the end of each season. This might explain the loss of efficacy in the third stem borer generation even though the dispensers still contained 30–40% of the pheromone load. An improved dispenser design would make it possible to achieve even better results in moth control with the same quantity of pheromone, and thus contribute to reducing the cost of the treatment even more.
Acknowledgements This project was funded by ‘‘Fundacio´n Jose´ y Ana Royo’’. We would like to thank Fernando Alfaro Lassala for technical support and CAPA-GVA for providing trial fields.
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