Efficacy of Insect Growth Regulators as Grain Protectants against Two Stored-Product Pests in Wheat and Maize

Transcription

1 942 Journal of Food Protection, Vol. 75, No. 5, 2012, Pages doi: / x.jfp Copyright G, International Association for Food Protection Efficacy of Insect Growth Regulators as Grain Protectants against Two Stored-Product Pests in Wheat and Maize NICKOLAS G. KAVALLIERATOS, 1 * CHRISTOS G. ATHANASSIOU, 2 BASILEIOS J. VAYIAS, 3 AND ŽELJKO TOMANOVIĆ 4 1 Laboratory of Agricultural Entomology, Department of Entomology and Agricultural Zoology, Benaki Phytopathological Institute, 8 Stefanou Delta str., 14561, Kifissia, Attica, Greece; 2 Laboratory of Entomology and Agricultural Zoology, Department of Agriculture, Crop Production and Rural Environment, University of Thessaly, Phytokou str., Nea Ionia, 38446, Magnissia, Greece; 3 Ministry of Rural Development and Food, General Directorate of Plant Produce, Directorate of Plant Produce Protection, 150 Syggrou Avenue, 17671, Kallithea, Greece; and 4 Institute of Zoology, Faculty of Biology, University of Belgrade, Studentski trg 16, 11000, Belgrade, Serbia MS : Received 27 August 2011/Accepted 2 December 2011 ABSTRACT Insect growth regulators (IGRs) (two juvenile hormone analogues [fenoxycarb and pyriproxifen], four chitin synthesis inhibitors [diflubenzuron, flufenoxuron, lufenuron, and triflumuron], one ecdysteroid agonist [methoxyfenozide], and one combination of chitin synthesis inhibitors and juvenile hormone analogues [lufenuron plus fenoxycarb]) were tested in the laboratory against adults of Prostephanus truncatus in maize and against adults of Rhyzopertha dominica in wheat. The tested IGRs were applied in maize at three doses (1, 5, and 10 ppm) and assessed at three temperature levels (20, 25, and 30uC) in the case of P. truncatus, while in the case of R. dominica the above doses were assessed only at 25uC in wheat. In addition to progeny production, mortality of the treated adults after 14 days of exposure in the IGR-treated commodities was assessed. All IGRs were very effective (.88.5% suppression of progeny) against the tested species at doses of $5 ppm, while diflubenzuron at 25uC in the case of P. truncatus or lufenuron and pyriproxyfen in the case of R. dominica completely suppressed (100%) progeny production when they were applied at 1 ppm. At all tested doses, the highest values of R. dominica parental mortality were observed in wheat treated with lufenuron plus fenoxycarb. Temperature at the levels examined in the present study did not appear to affect the overall performance in a great extent of the tested IGRs in terms of adult mortality or suppression of progeny production against P. truncatus in treated maize. The tested IGRs may be considered viable grain protectants and therefore as potential components in stored-product integrated pest management. Insect growth regulators (IGRs) consist of a group of insecticides that disrupt the normal development of insects and consequently affect development, reproduction, and behavior (40, 42, 44). They are classified as juvenile hormone analogues (JHAs), ecdysteroid agonists, and chitin synthesis inhibitors (CSIs) (42, 44). Due to their mode of action, IGRs act against the immature stages of insects by inhibiting metamorphosis and consequently insect proliferation on the infested commodity (42). These insecticides specifically act against insects and have low mammalian toxicity (39, 40, 42, 44). JHAs are responsible for the prolongation of the larval stage of insects (36). They act on the metamorphic change of the treated larvae, producing deformed pupae (40, 42), and they also block the completion of embryonic development (44). Furthermore, they affect the reproduction of treated adults by influencing vitellogenesis, development of follicles, protein synthesis in ovaries (29), sexual communication (52), or mating performance (49). JHAs may (8) or may not (54) reduce the fecundity of adults when they are applied on their larvae or pupae. * Author for correspondence. Tel: z ; Fax: z ; nick_kaval@hotmail.com. The use of ecdysteroid agonists generates the premature synthesis of the insect s cuticle especially close to the area of the head, causing feeding inhibition irrespectively of the age or instar of the insect (19, 48, 55). These compounds show chemosterilant activity of females (20), act both through stomach and contact (19), and can also penetrate the insect s cuticle (48). CSIs prevent the synthesis of new cuticle, and thus, the insects are not capable of proceeding into moulting, or they form abnormal cuticle (23, 40). They also affect the hormonal status of the treated insects and thus several physiological functions (23, 40, 46, 56). The above characteristics have resulted in the classification of IGRs as low-risk insecticides and raised interest for their evaluation as potential integrated pest management (IPM) components (19, 30). So far, only two forms of the IGR methoprene (methoprene containing the R and S isomers and s-methoprene containing the S isomer alone) are used as grain protectants in the United States, with label doses from 1 to 5 ppm (3, 5, 6, 54). In Australia s-methoprene is registered at the label dose of 0.6 ppm for a 9-month grain protection (11, 13). The IGRs fenoxycarb, pyriproxifen, diflubenzuron, flufenoxuron, lufenuron, triflumuron, and methoxyfenozide

2 J. Food Prot., Vol. 75, No. 5 EFFICACY OF INSECT GROWTH REGULATORS AGAINST STORED-PRODUCT PESTS 943 are commercially produced for use against insect pests in various agroecosystems (31, 33, 34, 37, 45, 50). Several studies have shown the potentiality of these compounds against stored-grain insect pests. Thind and Edwards (53) found in laboratory experiments that 1 ppm of fenoxycarb mixed with the food source caused complete inhibition of adult emergence of insecticide-susceptible or -resistant strains of Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae), Cryptolestes ferrufineus (Stephens) (Coleoptera: Laemophloeidae), and Oryzaephilus surinamensis (L.) (Coleoptera: Silvanidae) after 10, 8, and 7 weeks of exposure, respectively. Later, Edwards et al. (16) showed that 4.2 ppm of fenoxycarb applied on wheat in bins controlled O. surinamensis, Rhyzopertha dominica (F.) (Coleoptera: Bostrychidae), and T. castaneum for a period of 2 years. Büchi (7) evaluated fenoxycarb, mixed with food at 4, 8, and 16 ppm, against Liposcelis bostrychophila Badonnel (Psocoptera: Liposcelididae) and showed that it reduced considerably the adult production after 4, 6, and 8 weeks of exposure. However, the maximum reduction of the number of nymphs was 70.0% after 8 weeks of exposure. Furthermore, fenoxycarb caused reduction in adult fertility and hatching of the eggs of L. bostrychophila. Kostyukovsky et al. (30) reported that 0.1 ppm of pyriproxyfen applied in wheat flour inhibited completely the F 1 adults of strains of T. castaneum susceptible or resistant to pirimiphos-methyl. At 2.5 ppm, pyriproxifen caused 97% reduction of F 1 adults of a susceptible strain of Sitophilus oryzae (L.) (Coleoptera: Curculionidae) in treated wheat. Athanassiou et al. (4) reported significant reduction on the numbers of adults and nymphs of L. bostrychophila, Liposcelis decolor (Pearman) (Psocoptera: Liposcelididae), and Liposcelis paeta Pearman (Psocoptera: Liposcelididae) in comparison to the untreated controls 35 days after exposure on concrete arenas with the label rate of 2.3 mg of active ingredient per m 2. Desmarchelier and Allen (15) showed that there was no progeny of Sitophilus granarius (L.) (Coleoptera: Curculionidae) and S. oryzae adults after their exposure for a second 2- week period in wheat treated with diflubenzuron at concentrations ranging from 0.6 to 9.0 ppm. Ammar (2) evaluated flufenoxuron against S. oryzae in treated wheat and found that after 8 months posttreatment the F 1 progeny reduction was 92.9 or 96.3%, whereasthef 2 was95.9or100% at 5 or 25 ppm, respectively. Mahanthi (35), testing lufenuron and diflubenzuron as stored-maize protectants, showed that they inhibited completely the adult emergence of S. oryzae and controlled completely Corcyra cephalonica Stainton (Lepidoptera: Pyralidae). Eisa and Ammar (17) by testing the residual performance of triflumuron, fenoxycarb, and flufenoxuron in wheat against S. oryzae found that 50 or 100 ppm of these compounds provided complete control for a period of 2 years. Triflumuron and flufenoxuron controlled completely S. oryzae in treated wheat at 1 and 10 ppm for a period of 9 months, whereas fenoxycarb gave the same results at 10 ppm for a 12-month posttreatment period. Oberlander et al. (43) reported that the weight of larvae of Plodia interpunctella (Hübner) (Lepidoptera: Pyralidae) exposed in cereal food source treated with 10 and 25 ppm of methoxyfenozide increased 50% compared with the increase of 400% of the controls over 3 days. Furthermore, the mortality of the larvae in the treated diet increased dramatically for doses from 5 to 25 ppm compared with the controls. Prostephanus truncatus (Horn) (Coleoptera: Bostrychidae) and R. dominica are considered as major pests of stored maize and cassava or stored grains, respectively (21, 24). They are classified as primary pests, a fact that means that they are capable of infesting unbroken grain kernels. Although there are reports on the use of IGRs as grain protectants, very little information is available on the simultaneous evaluation of a wide range of IGRs (i.e., JHAs, CSIs, and ecdysteroid agonists) on both parental adult mortality and progeny production of P. truncatus and R. dominica. The objective of the present study was to test in the laboratory the insecticidal efficacy of IGRs, i.e., two JHAs (fenoxycarb and pyriproxifen), four CSIs (diflubenzuron, flufenoxuron, lufenuron, and triflumuron), one ecdysteroid agonist (methoxyfenozide), and one combination of CSI and JHA (lufenuron plus fenoxycarb) as grain protectants against P. truncatus in maize in different doses and at different temperature levels. The performance of the above IGRs was also evaluated against R. dominica in wheat. MATERIALS AND METHODS Insects. The insects tested were P. truncatus and R. dominica. P. truncatus was reared on whole maize, and R. dominica was reared on whole wheat. All species that were used in the experiments were obtained from the laboratory cultures of Benaki Phytopathological Institute at 25uC with 70% relative humidity and continuous darkness. For the bioassays, only adults were used,,4 weeks old. Formulations. The IGR formulations that were screened in the present study were Admiral EC containing 10% pyriproxyfen as the active ingredient (AI) (provided by Hellafarm, Amaroussion, Greece), Cascade DC containing 10% flufenoxuron (AI) (provided by BASF, Aghia Paraskevi, Greece), Runner SC containing 24% methoxyfenozide (AI) (provided by Bayer, Amaroussion, Greece), Alsystin SC containing 48% triflumuron (AI) (provided by Bayer), Match EC containing 5% lufenuron (AI) (provided by Syngenta, Anthousa, Greece), Helgar WG containing 25% fenoxycarb (AI) (provided by Hellafarm), Lufox EC containing 7.5% fenoxycarb and 3% lufenuron (AI) (provided by Syngenta), and Dimilin WP containing 25% diflubenzuron (AI) (provided by Dupont, Halandri, Greece). Commodities. Untreated, clean maize (Zea mays L. var. Dias) and hard wheat (Triticum durum Desf. var. Mexa) were used in the tests. The moisture contents of the tested commodities were 12.1 and 11.2% for maize and wheat, respectively, as determined by a moisture meter (Dickey-John Multigrain CAC II, Dickey-John Co., Auburn, IL) upon the beginning of the tests. Bioassay series 1. In the first series of bioassays, the tested IGRs were separately applied as solutions in 1-kg lots of maize at three doses: 1, 5, and 10 ppm. Spraying was carried out on a tray, where 1 kg of maize, spread into a thin layer, was treated with 3 ml of an aqueous solution that contained the appropriate volume of each IGR formulation corresponding to each dose. Spraying was carried out with an AG-4 airbrush (Mecafer, Valence, France). Furthermore, an additional 1-kg lot of maize was sprayed with 3 ml of distilled water and served as control. After spraying, the maize lots were placed into 5-liter glass jars and were manually shaken for 10 min to achieve equal distribution of the insecticide in the

3 944 KAVALLIERATOS ET AL. J. Food Prot., Vol. 75, No. 5 entire grain mass (25). Next, three samples, each of them weighing 30 g, were obtained from each treated (or untreated) maize lot and placed inside glass vials (7.5-cm diameter, 12.5-cm height). The closure of the vials had a 1.5-cm-diameter hole in the middle covered by gauze for sufficient aeration inside the vial. Thirty adults of P. truncatus were added in each vial, and the vials were inserted in incubators set at 25uC. Following this, all vials were placed in incubators set at the desired level of temperature each time. The tested levels of temperature were 20, 25, and 30uC, while relative humidity was maintained at 65% during the entire experimental period of this bioassay series. Parental mortality was assessed after a period of 14 days of exposure, while all parental individuals (dead and alive) were removed from the vials and the vials were again placed in the incubators for an additional period of 60 days under the same conditions as described above. Next, the vials were opened again and the adult progeny was counted. The entire procedure was repeated three times for each species by preparing new maize lots each time. Parental mortality rates in control groups were 5.5, 4.2, and 4.9% at 20, 25, and 30uC, respectively. Therefore, parental mortality data were transformed according to Abbott s formula (1). Parental mortality data and progeny production data were subjected to the same transformations as those used in bioassay series 1. The transformed mortality data were submitted to a three-way analysis of variance, with IGR, temperature, and dose as main effects. Similarly, the transformed progeny production counts were submitted to a three-way analysis of variance, with IGR, temperature, and dose as main effects. All analyses were done using the JMP software (47). Means were separated by the Tukey-Kramer honestly significant difference (HSD) test with P values set at 0.05(51). A preliminary Dunnett s test was used to examine the existence of significant differences in progeny production between control groups and IGRtreated maize for each of the tested temperature levels. Since Dunnett s test showed that control progeny was always statistically higher than the progeny that emerged in IGR-treated maize, control groups were not incorporated to the analyses of progeny production data. Bioassay series 2. In the second series of bioassays, the IGRs were applied in hard wheat and evaluated against R. dominica at 25uC following the same procedure as described for bioassay series 1. The parental mortality and progeny production counts were recorded as described above. Parental mortality in control groups was low and did not exceed 5%. However, parental mortality data were corrected according to Abbott s formula (1). The mortality of parental individuals and progeny production data were analyzed separately for each species. Prior to analysis, mortality data were arcsine transformed, while progeny production data were subjected to log (x z 1) transformation in order to achieve homogeneity of variances. The transformed mortality data were submitted to a two-way analysis of variance, with IGR and dose as main effects. Similarly, the transformed progeny production counts were submitted to a twoway analysis of variance, with IGR and dose as main effects. All analyses were done using the JMP software (47). Means were separated by the Tukey-Kramer HSD test with P values set at 0.05 (51). Control progeny data were not incorporated in the progeny production analysis because a preliminary Dunnett s test showed that progeny production in control groups was always significantly higher than that recorded in the IGR-treated wheat. RESULTS Bioassay series 1. All main effects (for IGR, F ~ 9.9, df ~ 7, 647, P, 0.01; for dose, F ~ 35.5, df ~ 2, 647, P, 0.01; for temperature, F ~ 4.4, df ~ 2, 647, P ~ 0.02) were significant, while all associated interactions (for IGR dose, F ~ 1.7, df ~ 14, 647, P ~ 0.06; for IGR temperature, F ~ 1.4, df ~ 14, 647, P ~ 0.13; for dose temperature, F ~ 1.6, df ~ 4, 647, P ~ 0.18) were not significant for mortality levels of parental P. truncatus individuals in maize treated with IGRs. Parental mortality of P. truncatus in maize treated with the tested IGRs was generally low since it did not exceed 27.5% when flufenoxuron was applied at 10 ppm at 30uC (Table 1). Temperature did not have significant influence in the parental mortality of P. truncatus with the exception of lufenuron plus fenoxycarb at 5 and 10 ppm, where significantly fewer individuals were killed at 20uC than at 30uC (Table 1). At 1 ppm, the parental mortality did not vary significantly inside each tested temperature level (Table 1). Similarly, at 5 and 10 ppm, the mortality of the parental P. truncatus individuals did not vary significantly inside 20 or 25uC and 25uC, respectively. All main effects (for IGR, F ~ 57.0, df ~ 7, 647, P, 0.01; for dose, F ~ 44.7, df ~ 2, 647, P, 0.01; for temperature, F ~ 51.1, df ~ 2, 647, P ~ 0.02) and their interactions (for IGR dose, F ~ 2.5, df ~ 14, 647, P, 0.01; for IGR temperature, F ~ 5.4, df ~ 14, 647, P, 0.01) were significant, with the exception of dose temperature (F ~ 1.6, df ~ 4, 647, P ~ 0.17), which was not significant for progeny production of P. truncatus in maize treated with IGRs. The tested IGRs caused suppression from 86.4 to 100% of progeny production of P. truncatus individuals (Table 2). Of the IGRs that were examined here, diflubenzuron was the most effective since it almost completely suppressed progeny in all of the tested cases (from 99.4 to 100%) (Table 2). The less effective tested IGRs were lufenuron, lufenuron plus fenoxycarb or triflumuron at 1 ppm, lufenuron or pyriproxifen at 5 ppm, and lufenuron plus fenoxycarb at 10 ppm, causing suppression of progeny production between 89.2 and 94.7% (Table 2). Temperature had a significant impact on the suppression of progeny production when flufenoxuron, pyriproxifen, and triflumuron were applied at 1 and 5 ppm, while it significantly affected the performance of fenoxycarb when it was applied at 1 ppm (Table 2). When fenoxycarb or pyriproxifen were applied at 1 ppm, significantly more progeny were suppressed at 20uC than at 25 or 30uC. In the case of flufenoxuron at the same dose, significantly more progeny were suppressed at 20 or 25uC than at 30uC, while for triflumuron at the same dose progeny was significantly suppressed at 20 or 30uC than at 25uC. The fact that significantly more progeny were recorded at 25uC than at 20 or 30uC was also observed when flufenoxuron or triflumuron was applied at 5 ppm, while when pyriproxifen was applied at the same dose significantly more progeny were suppressed at 20uC than at 25 or 30uC (Table 2). However, when the IGRs were applied at 10 ppm, temperature was not significantly associated with the progeny suppression in the treated maize (Table 2). Bioassay series 2. All main effects (for IGR, F ~ 18.4, df ~ 7, 215, P, 0.01; for dose, F ~ 48.9, df ~ 2, 215,

4 J. Food Prot., Vol. 75, No. 5 EFFICACY OF INSECT GROWTH REGULATORS AGAINST STORED-PRODUCT PESTS 945 TABLE 1. Mortality of P. truncatus adults after 14 days of exposure in maize treated with seven IGRs and one combination of IGRs at three doses and at three temperatures a Temp (uc): Dose (ppm) IGR Diflubenzuron A a A a A a Fenoxycarb A a A a A a Flufenoxuron A a A a A a Lufenuron A a A a A a Lufenuron z fenoxycarb A a A a A a Methoxyfenozide A a A a A a Pyriproxyfen A a A a A a Triflumuron A a A a A a 5 Diflubenzuron A a A a A bc Fenoxycarb A a A a A bc Flufenoxuron A a A a A bc Lufenuron A a A a A ab Lufenuron z fenoxycarb A a AB a B bc Methoxyfenozide A a A a A a Pyriproxyfen A a A a A ab Triflumuron A a A a A ab 10 Diflubenzuron A ab A a A ab Fenoxycarb A ab A a A ab Flufenoxuron A b A a A b Lufenuron A ab A a A ab Lufenuron z fenoxycarb A ab A a B b Methoxyfenozide A ab A a A ab Pyriproxyfen A ab A a A ab Triflumuron A a A a A a a Values are mean percentages standard errors. For each IGR or combination of IGRs, within each dose, means followed by the same uppercase letter are not significantly different; in all cases, df ~ 2, 26 (Tukey-Kramer HSD test; P ~ 0.05). Within each temperature and dose, means followed by the same lowercase letter are not significantly different; in all cases, df ~ 7, 71 (Tukey-Kramer HSD test; P ~ 0.05). P, 0.01) and associated interaction (for IGR dose, F ~ 3.2, df ~ 14, 215, P, 0.01) for mortality levels of parental individuals of R. dominica were significant in wheat treated with IGRs. Mortality of treated parental R. dominica adults ranged from 0.3 to 55.7%, and it was dose dependent. However, in the cases of fenoxycarb and methoxyfenozide the parental mortality was not significantly differentiated among the tested doses (Table 3). At all tested doses, the highest values of parental mortality were observed in wheat treated with lufenuron plus fenoxycarb but without significant difference compared with fenoxycarb or lufenuron at 1 ppm; fenoxycarb, lufenuron, or triflumuron at 5 ppm; and fenoxycarb, flufenoxuron, lufenuron, pyriproxyfen, or triflumuron at 10 ppm (Table 3). All main effects (for IGR, F ~ 14.5, df ~ 7, 215, P, 0.01; for dose, F ~ 14.0, df ~ 2, 215, P, 0.01) and associated interaction (for IGR dose, F ~ 16.7, df ~ 14, 215, P, 0.01) were significant for progeny production of R. dominica in wheat treated with IGRs. Progeny production of R. dominica in treated wheat was very low and did not exceed one adult per vial when the tested IGRs were applied at 5 or 10 ppm. Fenoxycarb, methoxyfenozide, and triflumuron were the least effective IGRs compared with the remainder of the tested IGRs when the last two were applied at 5 ppm and fenoxycarb was applied at 10 ppm (Table 4). Except for lufenuron plus fenoxycarb, which caused significantly lower suppression of R. dominica progeny (59.2%) than did the remainder of the tested IGRs at 1 ppm, suppression of progeny production of this species was not significantly differentiated inside the 1-ppm dose of the remainder of the tested IGRs, and it ranged from 88.6 to 100% (Table 4). DISCUSSION Through the use of IGRs, suppression of progeny production of stored-product insect pests can be achieved. For instance, Daglish (10) reported 99.6% suppression of progeny production of R. dominica in maize treated with 10 ml of solution of an emulsifiable concentrate of methoprene during 6 months of storage, while Kostyukovsky et al. (30) showed complete F 1 reduction of a pirimiphos-methyl susceptible strain of the same species in wheat treated with 3 ppm of methoprene. Elek and Longstaff (18) reported complete F 1 progeny production of the same species after 2 weeks of exposure of parental adults in wheat treated with 0.5 ppm of chlorfluazuron, teflubenzuron, and flufenoxuron. In light of the results of the present study, all IGR treatments were effective at doses of $5 ppm against both of the tested species since suppression of progeny production was consistently higher than 88.5%. Similar results have been reported by Thind and Edwards (53), who found that 5 ppm of fenoxycarb mixed with wheat reduced completely the adult emergence

5 946 KAVALLIERATOS ET AL. J. Food Prot., Vol. 75, No. 5 TABLE 2. Number of P. truncatus adults per vial in maize treated with seven IGRs and one combination of two IGRs at four doses (inclusive of 0 ppm), 60 days after the removal of the parental adults, and percent suppression of P. truncatus adults in comparison with controls a Mean no. SE % suppression Dose (ppm) IGR 20uC 25uC 30uC 20uC 25uC 30uC 0 Control A A B 1 Diflubenzuron A a A a A a Fenoxycarb A a B bc B b Flufenoxuron A a A b B b Lufenuron A c A d A b Lufenuron z fenoxycarb A bc A c A b Methoxyfenozide A ab A b A b Pyriproxyfen A ab B c B b Triflumuron A bc B d A b Diflubenzuron A a A a A a Fenoxycarb A a A b A b Flufenoxuron A a B b A b Lufenuron A b A c A b Lufenuron z fenoxycarb A a A bc A b Methoxyfenozide A a A b A b Pyriproxyfen A a B c B b Triflumuron A a B d A b Diflubenzuron A a A a A a Fenoxycarb A ab A abc A b Flufenoxuron A a A ab A ab Lufenuron A b A bc A b Lufenuron z fenoxycarb A b A c A b Methoxyfenozide A ab A abc A b Pyriproxyfen A ab A bc A b Triflumuron A ab A abc A b a Within a specific IGR or combination of IGRs and dose, means followed by the same uppercase letter are not significantly different. For each IGR or combination of IGRs, within dose, means followed by the same uppercase letter are not significantly different; in all cases, df ~ 2, 26 (Tukey-Kramer HSD test; P ~ 0.05). Within each dose and temperature, means followed by the same lowercase letter are not significantly different; in all cases, df ~7, 71 (Tukey-Kramer HSD test; P ~ 0.05). For all temperatures and IGRs or combination of IGRs, progeny in untreated maize was significantly higher than that recorded in treated maize; in all cases, df ~ 1, 17 (Dunnett s test; P ~ 0.05). of a strain of R. dominica susceptible to insecticides 12 weeks after exposure, whereas in the case of the resistant strain the reduction was 96%. However, for some of the tested IGRs high efficacy levels were recorded even with 1 ppm. In the case of R. dominica, 1 ppm of lufenuron or pyriproxyfen completely inhibited progeny emergence in wheat. At the same dose, diflubenzuron caused 98.5% suppression of production while in the case of P. truncatus, more than 99% suppression in progeny production was observed in maize at all three tested temperature levels. Similarly, 1 ppm of pyriproxyfen induced 99.7% reduction of F 1 adults of R. dominica in treated wheat (30), whereas 1 ppm of diflubenzuron inhibited completely the progeny production of the same species for a period of 8 months of storage (38). Furthermore, Arthur (3) found that 1 ppm of s- methoprene was also capable of suppressing completely the F 1 progeny of R. dominica in treated wheat for a period from 6 to 18 months after storage. The same reduction of progeny of R. dominica was noted by Mian and Mulla (38) when wheat was treated with 1 ppm of methoprene for a period of 10 months posttreatment. Despite the fact that lufenuron or pyriproxyfen appeared to be the most effective of the tested IGRs against R. dominica, this was not the case when the former was applied as a mixture with fenoxycarb, where progeny was suppressed only by 59.2% in wheat treated with 1 ppm. Based on our results, a negative interaction between fenoxycarb and lufenuron is apparent in the case of R. dominica when these IGRs are applied as a combination in wheat at 1 ppm since the effectiveness of this mixture was significantly lower than that recorded with either lufenuron or fenoxycarb alone. The modes of insecticidal action of lufenuron and fenoxycarb differ because the former is a CSI and the latter is a JHA. Based on the percentage of each substance to the tested mixture, this treatment comprised 7.5% fenoxycarb and 3% lufenuron, so we could assume that the mode of action of this formulation was based to a greater extent on fenoxycarb than on lufenuron. Although further investigation is required to test this hypothesis, the fact that fenoxycarb at 1 ppm showed the lowest performance of the tested IGRs against R. dominica may explain the reduced performance that was also observed in the mixture of fenoxycarb with lufenuron against this species.

6 J. Food Prot., Vol. 75, No. 5 EFFICACY OF INSECT GROWTH REGULATORS AGAINST STORED-PRODUCT PESTS 947 TABLE 3. Mortality of R. dominica adults after 14 days of exposure in wheat treated with seven IGRs and one combination of two IGRs at three doses a Mean % mortality SE at each dose IGR 1 ppm 5 ppm 10 ppm Diflubenzuron A a B ab C ab Fenoxycarb A c A bc A abc Flufenoxuron A ab A a B bc Lufenuron A bc B c B bc Lufenuron z fenoxycarb A c B c C c Methoxyfenozide A ab A ab A a Pyriproxyfen A ab A ab B abc Triflumuron A ab A abc B abc a For each IGR or combination of IGRs, means followed by the same uppercase letter are not significantly different; in all cases, df ~ 2, 26 (Tukey-Kramer HSD test; P ~ 0.05). Within each dose, means followed by the same lowercase letter are not significantly different; in all cases, df ~ 7, 71 (Tukey-Kramer HSD test; P ~ 0.05). Contrary to our results, previous studies document successful combinations of IGRs belonging to different groups against stored-product pests, and therefore, generalizations in terms of the compatibility between different IGRs should be avoided and each case should be examined separately. For instance, Daglish and Wallbank (14) reported that a mixture of diflubenzuron (1 ppm) and methoprene (1 ppm) applied in sorghum was very effective against R. dominica and S. oryzae. This finding is interesting because the JHA methoprene and the ecdysteroid agonist RH-5849 or tebufenozide applied alone in wheat have shown low performance against S. oryzae (30). Similar results were also recorded for RH-5849 and tebufenozide against R. dominica for doses of 2.5 and 5 ppm, respectively (30). Furthermore, the combination of the JHAs methoprene at 10 ppm and fenoxycarb at 5 ppm with the ecdysteroid agonists TABLE 4. Number of R. dominica adults per vial in wheat treated with seven IGRs and one combination of two IGRs at four doses (inclusive of 0 ppm), 60 days after the removal of the parental adults, and percent suppression of R. dominica adults in comparison with controls a Dose (ppm) IGR Mean no. SE % suppression 0 Control Diflubenzuron A 98.5 Fenoxycarb A 88.6 Flufenoxuron A 89.1 Lufenuron A Lufenuron z fenoxycarb B 59.2 Methoxyfenozide A 89.6 Pyriproxyfen A Triflumuron A Diflubenzuron A Fenoxycarb B 95.0 Flufenoxuron A Lufenuron A Lufenuron z fenoxycarb A 99.5 Methoxyfenozide AB 98.0 Pyriproxyfen A Triflumuron AB Diflubenzuron A Fenoxycarb B 95.0 Flufenoxuron A Lufenuron A Lufenuron z fenoxycarb A 99.5 Methoxyfenozide A 99.5 Pyriproxyfen A Triflumuron A 99.5 a Within each dose, means followed by the same letter are not significantly different; in all cases, df ~ 7, 71 (Tukey-Kramer HSD test; P ~ 0.05). For all IGRs or combination of IGRs, progeny in untreated wheat was significantly higher than that recorded in treated wheat; in all cases, df ~ 1, 17 (Dunnett s test; P ~ 0.05).

7 948 KAVALLIERATOS ET AL. J. Food Prot., Vol. 75, No. 5 tebufenozide and methoxyfenozide at 5 or 10 ppm in the cereal rearing diet of the last instar larvae of P. interpunctella increased their mortality compared with the cases when tebufenozide or methoxyfenozide was applied alone (43). IGRs act mainly against the immature developmental stages and exhibit little or no lethal effect against adults (9, 12, 30, 39, 42), but our results seem to contradict the above statement since all of the tested species exhibited some mortality levels after contact with the IGR-treated commodities, while mortality for all species was dose dependent. However, this is not the first study in which toxic effects of IGRs against adults have been reported. For instance, Letellier et al. (32) reported that mortality of Sitophilus zeamais Motschulsky after contact with fenoxycarb-treated grain ranged from 4 to 45.5% following an increase of the fenoxycarb dose from to 10 ppm. Ammar (2) reported mortality of parental S. oryzae adults of from 24 to 55% initially after their exposure in wheat treated with 0.5 to 25 ppm of flufenoxuron and falling to between 2 and 6% 3 months posttreatment. Similar trends were also noted for the IGRs chlorfluazuron and XRD-473. Although Arthur (3) found slight mortality of R. dominica adults after contact with a dust formulation of s-methoprene, the author attributed this fact to the insecticidal properties of the dust itself rather than s-methoprene, since in the same study a liquid formulation of s-methoprene appeared to have no toxic effect against adults of this species. In light of our findings, doses equal to or higher than 5 ppm of fenoxycarb, lufenuron, and lufenuron plus fenoxycarb had lethal effects against parental adults of the tested species. Furthermore, R. dominica adults were more susceptible than P. truncatus adults to the IGR treatments tested here. Although we are not able to explain the adult mortality that was observed in the present study, which in some cases, especially against R. dominica, exceeded 55%, the adulticidal properties of IGRs, if they do exist, may become a useful tool in stored-product management. Those substances may enhance the efficacy of low doses of insecticides and thus provide adequate control of parental adults, preventing also their progeny production at the same time. For example, the combination of chlorpyrifosmethyl at 5 ppm with s-methoprene at 0.6 ppm in wheat increased significantly the mortality of the parental adults of a malathion-pyrethroid resistant strain of T. castaneum compared with the mortality obtained by treatments with chlorpyrifos-methyl or s-methoprene alone (11). Also, the combination of these compounds inhibited completely the progeny production as in the case of the application of chlorpyrifos-methyl alone (11). However, the selection of each IGR X insecticide combination requires great care since compatibility between these control strategies is not always feasible, especially against insecticide-resistant strains of insects, due to cross-resistance development. Kostyukovsky et al. (30) reported that a pirimphos-methyl resistant strain of T. castaneum exhibited cross-resistance to the JHAs methoprene and pyriproxifen, while in a more recent study Daglish (11) found that a malathion-resistant strain of S. oryzae and a malathion-pyrethroid resistant strain of T. castaneum were also tolerant to s-methoprene treatment alone. Temperature is a key factor in stored-product protection since it often affects the effectiveness of control methods (25 27) while at the same time influencing the developmental parameters of stored-product pests (22). Although in a few cases in our study significant differences in the performance of IGRs were observed among the tested temperatures, temperature did not appear to affect the overall performance of the tested IGRs in terms of adult mortality or suppression of progeny production against P. truncatus in treated maize. The results of our study suggest that IGRs could be considered as temperature-compatible IPM components at least for the temperature range and for the insect species tested. However, further research is required before accepting the stability of IGRs over a broad range of temperatures and species. The tested IGRs have been proposed as potential protectants against several non stored-product insect pests belonging to different orders. For example, Mendel and Rosenberg (37) reported that fenoxycarb inhibited the development of first and second instars of Matsucoccus josephi Bodenheimer and Harpaz (Hemiptera: Margarodidae), reduced the scale settlement, and caused 98% nymphal mortality. According to Ludwig and Oetting (34), pyriproxyfen and diflubenzuron reduced the adult emergence of Frankliniella occidentalis (Pergande) (Thripidae: Thysanoptera). Diflubenzuron reduced the longevity of Riptortus clavatus (Thunberg) (Hemiptera: Alydidae) adults as well as the number of eggs laid per female (28). Lababidi (31) found that flufenoxuron caused more than 86% mortality of Agonoscena targionii (Lichtenstein) (Hemiptera: Psyllidae) nymphs in a 2-year experiment. Navarro-Llopis et al. (41) showed that continuous applications of lufenuron on several generations of Ceratitis capitata (Wiedemann) (Diptera: Tephritidae) were capable of reducing the population of the insect and the fruit damage in citrus orchards during a period of 4 years. Shamshad et al. (50) reported that triflumuron was very effective against the third instar larvae of Lycoriella ingenua Dufour (Diptera: Sciaridae) causing 93% mortality at one-half of the recommended dose. Rodríguez-Enríquez et al. (45) showed that when larvae of Spodoptera exigua (Hübner) (Lepidoptera: Noctuidae) were fed with an artificial diet mixed with methoxyfenozide, it resulted in pupae with significantly less weight than the controls, in adult deformities, and in reduction in egg hatching. In conclusion, application doses of $5 ppm of the tested IGRs were of high effectiveness against both species examined in the present study, and therefore, these compounds should be considered as potential components in stored-product IPM. Screening of the tested IGRs revealed that efficacy was dose dependent, and in several cases almost complete suppression of progeny production was achieved even with 1 ppm. In contrast, temperature did not appear to influence the efficacy of the tested doses to a great extent. Parental adult mortality was recorded for all species tested in the present study. Additional studies are required to confirm the above findings and examine mixtures of these compounds with low-risk insecticides aiming to provide long-term protection in stored grains.

8 J. Food Prot., Vol. 75, No. 5 EFFICACY OF INSECT GROWTH REGULATORS AGAINST STORED-PRODUCT PESTS 949 ACKNOWLEDGMENT This work was supported by the project Control of Prostephanus truncatus, Rhyzopertha dominica and Tribolium confusum with the use of insect growth regulators (Benaki Phytopathological Institute). REFERENCES 1. Abbott, W. S A method of computing the effectiveness of an insecticide. J. Econ. Entomol. 18: Ammar, I. M. A Residual bioactivity of insect growth regulators against Sitophilus oryzae (L.) in wheat grain. Anz. Schaedlingskd. Pflanzenschutz 61: Arthur, F. H Evaluation of methroprene alone and in combination with diatomaceous earth to control Rhyzopertha dominica (Coleoptera: Bostrichidae) on stored wheat. J. Stored Prod. Res. 40: Athanassiou, C. G., F. H. Arthur, N. G. Kavallieratos, and J. E. Throne Efficacy of pyriproxifen for control of stored-product psocids (Psocoptera) on concrete surfaces. J. Econ. Entomol. 104: Athanassiou, C. G., F. H. Arthur, and J. E. 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