Ex Parte Nishiguchi et alDownload PDFPatent Trial and Appeal BoardMar 15, 201712978825 (P.T.A.B. Mar. 15, 2017) Copy Citation United States Patent and Trademark Office UNITED STATES DEPARTMENT OF COMMERCE United States Patent and Trademark Office Address: COMMISSIONER FOR PATENTS P.O.Box 1450 Alexandria, Virginia 22313-1450 www.uspto.gov APPLICATION NO. FILING DATE FIRST NAMED INVENTOR ATTORNEY DOCKET NO. CONFIRMATION NO. 12/978,825 12/27/2010 Naonobu NISHIGUCHI 20101815A 1927 513 7590 03/17/2017 WENDEROTH, LIND & PONACK, L.L.P. 1030 15th Street, N.W., Suite 400 East Washington, DC 20005-1503 EXAMINER ZHANG, YANZHI ART UNIT PAPER NUMBER 1617 NOTIFICATION DATE DELIVERY MODE 03/17/2017 ELECTRONIC Please find below and/or attached an Office communication concerning this application or proceeding. The time period for reply, if any, is set in the attached communication. Notice of the Office communication was sent electronically on above-indicated "Notification Date" to the following e-mail address(es): ddalecki@wenderoth.com eoa@ wenderoth. com PTOL-90A (Rev. 04/07) UNITED STATES PATENT AND TRADEMARK OFFICE BEFORE THE PATENT TRIAL AND APPEAL BOARD Ex parte NAONOBU NISHIGUCHI, HIROYUKI KAWANO, and KAZUHIDE NAKADA Appeal 2016-002252 Application 12/978,825 Technology Center 1600 Before JEFFREY N. FREDMAN, JOHN G. NEW, and TAWEN CHANG, Administrative Patent Judges. FREDMAN, Administrative Patent Judge. DECISION ON APPEAL This is an appeal1 under 35 U.S.C. § 134 involving methods of attracting a fly. The Examiner rejected the claims as obvious. We have jurisdiction under 35 U.S.C. § 6(b). We reverse and enter a new ground of rejection. 1 Appellants identify the Real Party in Interest as the SUMITOMO CHEMICAL COMPANY, LIMITED (see App. Br. 2). Appeal 2016-002252 Application 12/978,825 Statement of the Case Background “An object of the present invention is to provide a fly attractant composition which has an excellent attracting effect on flies and a fly attracting method” (Spec. 1:17-19). The Claims Claims 15, 16, 26, and 28 are on appeal. Claim 15 is representative and is reproduced below. 15. A method of attracting a fly which comprises applying an effective amount of a ligninsulfonate to an area where the fly lives, wherein the ligninsulfonate is contained in a fly attractant composition, and the fly attractant composition contains 5 to 20% by weight of the ligninsulfonate and at least 0.1% by weight but less than 5% by weight of an insecticidal active ingredient, wherein the content of the ligninsulfonate is 4 to 200 times the content of the insecticidal active ingredient. The Issues A. The Examiner rejected claims 15 and 26 under 35 U.S.C. § 103(a) as obvious over Tanio2 and Valent3 (Ans. 2-4). B. The Examiner rejected claims 16 and 28 under 35 U.S.C. § 103(a) as obvious over Tanio, Valent, and Tsutomu4 (Ans. 4-8). Because the same issue is dispositive for both of these rejections, we will consider them together. 2 Tanio et al., JP 2008-143804 A, published June 26, 2008 (“Tanio”). 3 Valent, Material Safety Data Sheet, 2009 Valent U.S.A. Corporation (“Valent”). 4 Tsutomu, N., JP 62-042903, published Feb. 24, 1987 (Tsutomu”). 2 Appeal 2016-002252 Application 12/978,825 We limit our consideration of the merits of the appealed rejection to the elected species. See Ex parte Ohsaka, 2 USPQ2d 1460, 1461 (BPAI 1987). Thus, we read the claims as limited to the use of the elected clothianidin as the active insecticide and cis-9-tricosene as the pheromone (see Response to Election/Restriction filed on June 15, 2012). The issue with respect to these rejections is: Does the evidence of record support the Examiner’s conclusion that Valent render obvious the use of clothianidin as an insecticide for flies? Findings of Fact 1. Valent teaches a composition of 0.25% clothianidin, but provides no information regarding target insects (see Valent 2). Principles of Law A prima facie case for obviousness “requires a suggestion of all limitations in a claim,” CFMT, Inc. v. Yieldup Int’l Corp., 349 F.3d 1333, 1342 (Fed. Cir. 2003) and “a reason that would have prompted a person of ordinary skill in the relevant field to combine the elements in the way the claimed new invention does.” KSR Int’l Co. v. Teleflex Inc., 550 U.S. 398, 418 (2007). Analysis We agree with Appellants that: The Valent reference does not state that clothianidin is effective against flies. [A]n insecticide effective against one insect may not necessarily be effective against all other insects. Therefore, the replacement of the active ingredient of the Tanio reference effective to treat and eliminate flies with another active ingredient, namely clothianidin, which is not taught by the Valent reference to treat flies is not a simple substitution as suggested by the Examiner. (App. Br. 5). 3 Appeal 2016-002252 Application 12/978,825 That is, Valent provides no reason to use clothianidin in a method of attracting and killing flies as taught by Tanio (FF 1) and recited in claim 15. Conclusion of Law The evidence of record does not support the Examiner’s conclusion that Valent render obvious the use of clothianidin as an insecticide for flies. New Ground of Rejection Under the provisions of 37 C.F.R. § 41.50(b), we enter the following new ground of rejection. Claims 15, 16, 26, and 28 are rejected under 35 U.S.C. § 103(a) as obvious over Miura,5 Tanio, Barry,6 and Tsutomu. Findings of Fact 2. Miura teaches a “method for controlling flies [comprising using a compound or salt thereof having an affinity for a nicotinic acetylcholine receptor of insects,] wherein the compound is one or more compounds selected from the group consisting of clothianidin (common name)” (Miura 1113,16). 3. Miura teaches that the compounds used in its method may be formulated as a preparation to which emulsifier, spreader, penetrant, and the like may be added (Miura 138). Miura teaches “surfactants to be used as 5 Miura et al., US 2001/0046986 Al, published Nov. 29, 2001 (“Miura”). 6 Barry et al., Feeding And Survivorship Of Blueberry Maggot Flies (Diptera: Tephritidae) On Protein Baits Incorporated With Insecticides, 88(3) Florida Entomologist, 268-277 (2005) (“Barry”). 4 Appeal 2016-002252 Application 12/978,825 emulsifier, spreader, penetrant, dispersant or the like include nonionic and anionic surfactants such as . . . alkylsulfonates” (Miura 142). 4. Miura teaches “it is preferred to add about 1 to 20% by weight, preferably about 1 to 15% by weight of a surfactants [sic]” (Miura 145). 5. Miura teaches the use of an insecticidal composition “in combination with one or two or more attractants” (Miura 148). 6. Miura teaches “the liquid composition has concentration of from 0.001 to 20% by weight of the active component, and attractants are added in an amount of from 1 to 20% by weight” (Miura 148). 7. Miura teaches a working example with a composition as reproduced below: Preparation of :50% ciilotManidme granular wetta'ble powder Clothianidin 50 parts. Aikcnylsulfate 10 parts and I\s1u.10ULl- tt^UU t tr^rr* !A-W,-L-L% (Miura 151). Miura further teaches a working example of an insecticidal test where clothianidin were orally administered to flies: Female Musca domestica imagoes (ten flies) of a sensitive strain kept without water for 3-4 hours were anesthetized and lightly fixed to a pressure sensitive adhesive sheet which was cut into strips. After the test insects came out from the anesthetic, 1 pL of the liquid composition (containing 0.05 jug of clothianidin) was released from an injection needle attached to a micro dropper so that the test insects inhaled the composition. After 24 hours, it was confirmed that all flies were dead. (Miura 1 58). 5 Appeal 2016-002252 Application 12/978,825 8. Tanio teaches a “control method of the flies which fly to the habitat of livestock” (Tanio 11) using a “poison bait agent” (Tanio 1 5) that comprises “(a) ten to 30 mass percent 1-methyl-2-nitro-3-[(3-tetrahydro furil) methyl] guanidine, (b) surfactant One to 20 mass percent. . . (c) water- soluble powdered carrier” (Tanio 17). 9. Tanio teaches surfactants include a lignosulfonic acid salt (Tanio 19) and exemplifies a preparation using WG-4, a ligninsulfonate surfactant (Tanio 119). 10. Barry teaches a “no-choice assay evaluated fly mortality to bait containing the following insecticides . . . clothianidin” (Barry 271, col. 1). 11. Figure 1, panel B of Barry is reproduced below: list sa 4# 2§ § wK Y x Y& m “Fig. 1. Percent of flies that were living (active + incapacitated flies) after feeding for 10 s on a droplet of a given insecticide concentration. . . . B) Clothianidin . . . (T =4 ppm, o = 40 ppm, • = 400 ppm insecticide; X = control bait without insecticide)” (Barry, p. 272). 12. Tsutomu teaches a “fly attractant obtained by mixing at least one active constituent selected from . . . cis-9-tricosene . . . The above- 6 Appeal 2016-002252 Application 12/978,825 mentioned fly attractant is capable of exhibiting improved effect by application to a toxic bait” (Tsutomu, abstract). Analysis Miura teaches a method of controlling (FF 2) and attracting (FF 5) flies that comprises the application of an effective amount of a surfactant (FF 3) in amounts ranging from 1 to 20 % by weight (FF 4) in combination with active insecticides including clothianidin (FF 2) in amounts ranging from 0.001 to 20% by weight (FF 6). Miura further provides an example using clothianidin solution as insecticide against flies (FF 7). Miura does not teach selection of a ligninsulfonate surfactant and, for claims 16 and 28, does not teach the use of a cis-9-tricosene pheromone. Tanio teaches a method of controlling flies using an insecticidal active with a surfactant (FF 8) where the surfactants include a lignosulfonic acid salt (FF 9). Barry teaches that clothianidin is active at amounts as low as 40 and 400 ppm (FF 10-11). Applying the KSR standard of obviousness to the findings of fact, we conclude that the person of ordinary skill reasonably would have found it obvious to select the ligninsulfonate surfactant of Tanio for use as a surfactant in the fly control method of Miura (FF 2-6) because Tanio teaches that ligninsulfonate surfactants may be incorporated into fly control compositions as known equivalents (FF 9). The selection of the specific surfactant, when they are demonstrated as equivalent by the prior art, is merely a “predictable use of prior art elements according to their established functions.” KSR, 550 U.S. at 417. The claims “recite[] a combination of elements that were all known in the prior art, and all that was required to 7 Appeal 2016-002252 Application 12/978,825 obtain that combination was to substitute one well-known . . . agent for another.” Wm. WrigleyJr. Co. v. Cadbury Adams USALLC, 683 F.3d 1356, 1364 (Fed. Cir. 2012). With regard to the limitation in claims 15 and 16 regarding the ratio of the ligninsulfonate surfactant to the insecticidal active, Miura teaches overlapping ranges for both the surfactant and the insecticidal active (FF 4, 6) where the starting point for the surfactant is 1 % (FF 4) while the starting point for the insecticidal active is 0.001% (FF 6). See In re Peterson, 315 F.3d 1325, 1329 (Fed. Cir. 2003) (“A prima facie case of obviousness typically exists when the ranges of a claimed composition overlap the ranges disclosed in the prior art.”) We further note that simply because claims 15 and 16 optimize amounts of surfactant and insecticide as a ratio does not render optimization of the ratio of the two components nonobvious. See In re Applied Materials, 692 F.3d 1289, 1298 (Fed. Cir. 2012) (“The mere fact that multiple result- effective variables were combined does not necessarily render their combination beyond the capability of a person having ordinary skill in the art.”) Here, Miura teaches that both components may be optimized, rendering their combined optimization obvious (FF 4, 6). We note that Barry evidences that lower amounts of clothianidin are sufficient for fly control, further supporting optimization at the lower end of Miura’s range (FF 6, 11). Finally, with regard to claims 16 and 28, Tsutomu teaches incorporation of pheromones such as cis-9-tricosene as a fly attractant (FF 12). Applying the KSR standard of obviousness to the findings of fact, we conclude that the person of ordinary skill reasonably would have found it 8 Appeal 2016-002252 Application 12/978,825 obvious to incorporate Tsutomu’s pheromones because Tsutomu teaches that the “fly attractant is capable of exhibiting improved effect by application to a toxic bait” (FF 12). That is, the pheromone would improve efficacy of the toxic bait. See DyStar Textilfarben GmbH & Co. Deutschland KG v. C.H. Patrick Co., 464 F.3d 1356, 1368 (Fed. Cir. 2006) (“[Motivation to combine exists not only when a suggestion may be gleaned from the prior art as a whole, but when the . . . combination of references results in a product or process that is more desirable, for example because it is . . . more efficient.”) We have considered Appellants arguments, but find most of them inapplicable to the rejection discussed above. While Appellants’ unexpected results argument remains applicable, we do not find the results persuasive for several reasons. First, Appellants have not compared their formulation to the closest prior art of Miura (FF 7), or even to the results of Tanio (FF 9) or Barry (FF 11). See In re Baxter TravenolLabs., 952 F.2d 388, 392 (Fed. Cir. 1991) (“[W]hen unexpected results are used as evidence of nonobviousness, the results must be shown to be unexpected compared with the closest prior art.”). Second, Appellants do not identify a teaching in the Specification or other evidence that teaches the results are unexpected. See In re Klosak, 455 F.2d 1077, 1080 (CCPA 1972) (“It is not enough to show that results are obtained which differ from those obtained in the prior art: that difference must be shown to be an unexpected difference”). In this case, there is no evidence that the difference between the formulations in Table 1 of the Specification was unexpected (see Spec. 1 52). 9 Appeal 2016-002252 Application 12/978,825 The only evidence that the results are unexpected is based on attorney argument. See In re Soni, 54 F.3d 746, 750 (Fed. Cir. 1995) (“It is well settled that unexpected results must be established by factual evidence. Mere argument or conclusory statements . . . [do] not suffice.”). Also see In re Pearson, 494 F.2d 1399, 1405 (CCPA 1974) (“Attorney’s argument in a brief cannot take the place of evidence”). Third, we note that in Examples 9 and 10, 5% clothianidin and 3% ligninsulfonate surfactant was used, while in comparative examples 11, 12, and 13, no more than 1% clothianidin was used with different amounts of ligninsulfonate surfactant, so the evidence in Table 1 of the Specification does not compare identical amounts of the formulation. Therefore, not only was no comparison performed with other surfactants, but the evidence is not commensurate in scope with the ranges of claim 1. Harris found that “showing of unexpected results is not commensurate in scope with the degree of protection sought by the claimed subject matter because ... the record does not show that the improved performance would result if the weight-percentages were varied within the claimed ranges.” In re Harris, 409 F.3d 1339, 1344 (Fed. Cir. 2005). SUMMARY In summary, we reverse the Examiner’s obviousness rejections. We reject claims 15, 16, 26, and 28 under 35 U.S.C. § 103(a) as obvious over Miura, Tanio, Barry, and Tsutomu. This decision contains a new ground of rejection pursuant to 37 C.F.R. § 41.50(b). Section 41.50(b) provides “[a] new ground of rejection 10 Appeal 2016-002252 Application 12/978,825 pursuant to this paragraph shall not be considered final for judicial review.” Section 41.50(b) also provides: When the Board enters such a non-final decision, the appellant, within two months from the date of the decision, must exercise one of the following two options with respect to the new ground of rejection to avoid termination of the appeal as to the rejected claims: (1) Reopen prosecution. Submit an appropriate amendment of the claims so rejected or new Evidence relating to the claims so rejected, or both, and have the matter reconsidered by the examiner, in which event the prosecution will be remanded to the examiner. The new ground of rejection is binding upon the examiner unless an amendment or new Evidence not previously of Record is made which, in the opinion of the examiner, overcomes the new ground of rejection designated in the decision. Should the examiner reject the claims, appellant may again appeal to the Board pursuant to this subpart. (2) Request rehearing. Request that the proceeding be reheard under § 41.52 by the Board upon the same Record. The request for rehearing must address any new ground of rejection and state with particularity the points believed to have been misapprehended or overlooked in entering the new ground of rejection and also state all other grounds upon which rehearing is sought. No time period for taking any subsequent action in connection with this appeal may be extended under 37 C.F.R. § 1.136(a)(l)(iv). REVERSED: 37 C.F.R, $ 41.50(b) 11 Application/Control No. Applicant(s)/Patent Under Patent Notice of References Cited 12/978,825 Appeal No. 2016-002252 Administrative Patent Judge 1 Art Unit Page 1 of 1 Jeffrey N. Fredman 1600 U.S. PATENT DOCUMENTS * Document Number Country Code-Number-Kind Code Date MM-YYYY Name Classification A US-2001/0046986 A1 11/2001 Miura et al. B US- C US- D US- E US- F US- G US- H US- 1 US- J US- K US- L US- M US- FOREIGN PATENT DOCUMENTS * Document Number Country Code-Number-Kind Code Date MM-YYYY Country Name Classification N O P Q R S T NON-PATENT DOCUMENTS * Include as applicable: Author, Title Date, Publisher, Edition or Volume, Pertinent Pages) U Barry et al., Feeding And Survivorship Of Blueberry Maggot Flies (Diptera: Tephritidae) On Protein Baits Incorporated With Insecticides, 88(3) Florida Entomologist, 268-277 (2005) V w X *A copy of this reference is being furnished with the associated Board decision from this appeal.. Dates in MM-YYYY format are publication dates. Classifications may be US or foreign. U.S. Patent and Trademark Office PTO-892 (Rev. 01-2001) Notice of References Cited Part of Paper No. 268 Florida Entomologist 88(3) September 2005 FEEDING AND SURVIVORSHIP OF BLUEBERRY MAGGOT FLIES (DIPTERA: TEPHRITIDAE) ON PROTEIN BAITS INCORPORATED WITH INSECTICIDES James D. Barry1 and Sridhar Polavarapu2 Blueberry and Cranberry Research and Extension Center, Rutgers University 125A Lake Oswego Road, Chatsworth, NJ 08019 Current Address: DuPont Crop Protection, Stine-Haskell Research Center, 1090 Elkton Road, Newark DE 19714; james.d.barry@usa.dupont.com 2Deceased, 7 May 2004 Abstract Laboratory feeding trials evaluated fly survivorship on six insecticides (acetamiprid, clothianidin, deltamethrin, fipronil, imidacloprid, and spinosad) incorporated at 4, 40, and 400 ppm in protein baits. Higher concentrations of insecticides resulted in increased fly mor- tality. At all concentrations of insecticides in baits, except those on deltamethrin, there was a significantly higher mortality 4 d after the initial feeding, compared with flies that fed on a control bait. The presence of clothianidin or imidacloprid in baits led to significantly less feeding compared with a control bait without insecticide. There were no feeding deterrent ef- fects of bait containing either fipronil or spinosad compared with a control bait without in- secticide. Exposure of flies to fresh bait containing 40 ppm of acetamiprid, clothianidin, or imidacloprid, resulted in significantly more flies becoming knocked down than the control. Baits containing 40 ppm of fipronil or spinosad resulted in higher levels of fly mortality than baits containing either neonicotinoids (acetamiprid, clothianidin, or imidacloprid) or no in- secticide for trials with fresh and 1-d-old bait with unlimited exposure. At the rates tested baits containing deltamethrin resulted in no fly knockdown and always had the lowest mor- tality of any insecticide treatment. The tradeoffs between insecticides capable of knockdown and mortality are discussed as they relate to management of R. mendax. Key Words: Rhagoletis mendax, acetamiprid, clothianidin, fipronil, imidacloprid, spinosad Resumen La sobrevivencia de la mosca, Rhagoletis mendax, contra seis insecticidas (acetamiprid, clo- thianidin, deltamethrin, fipronil, imidacloprid, y spinosad) incorporados a 4, 40, y 400 ppm en cebos de proteina fue evaluada en pruebas de alimentacion en el laboratorio. Las concen- traciones mas altas de insecticidas resultaron en un aumento de la mortalidad de las mos- cas. En todas las concentraciones de los insecticidas en cebo, menos aquellas tratadas con deltamethrin, hubo una mortalidad significativamente mas alta 4 d despues de la alimenta- cion inicial, comparada con las moscas que se alimentaron sobre el cebo de control. La pre- sencia de clothianidin o imidacloprid en el cebo resulto en una alimentacion significativamente menor comparada con el cebo de control sin insecticida. No hubo ningun efecto detrimental en la alimentacion del cebo que tenia fipronil o spinosad comparado con el cebo de control sin insecticida. La exposicion de las moscas al cebo fresco con 40 ppm de acetamiprid, clothianidin o imidacloprid, resulto en significativamente mas moscas derriba- das que en el control. Los cebos con 40 ppm de fipronil o spinosad resultaron en un nivel mas alto de la mortalidad de moscas que en los cebos con neonicotinoids (acetamiprid, clothiani- din, o imidacloprid) o sin insecticida para las pruebas con cebo fresco y de cebo de un dia con exposicion sin limite. A las tasas de insecticidas probadas, ningun mosca fue afectada en la prueba de cebo con deltamethrin y siempre tenian la menor mortalidad que cualquier otro tratamiento de insecticidas. Se commentan sobre los factores de los insecticidas con la capa- cidad para un efecto de noqueo de las moscas versus un insecticida que mata las moscas en relacion al manejo de R. mendax. The blueberry maggot fly, Rhagoletis mendax Curran, is a serious pest of lowbush and highbush blueberries, Vaccinium angustifolium Aiton and V. corymbosum L., respectively, in the northeast- ern United States and Atlantic Provinces of Can- ada. In areas not infested with R. mendax there is zero-tolerance for maggot presence. As a result, growers exporting fruit to non-infested areas of Scientific names italics only 269 Canada must participate in a Blueberry Certifica- tion Program (Canadian Food Inspection Agency 1999). This certification program mandates following either a calendar-based or an integrated pest management (IPM) spray program. A calendar- based approach requires growers to start spray- ing insecticides within 10 d of the first detection of an adult fly in the area, and continue spraying at 7- to 10-d intervals until the end of harvest. An IPM-spray program requires growers to monitor the presence of adults with traps baited with am- monium acetate. A recommended insecticide should be applied within 5 d of the date of capture of a single fly in any one of the monitoring traps, followed by a second spray 7-10 d later. This spray sequence should be repeated for each subsequent fly detection until the end of harvest. Many blue- berry growers use one of these spray regimens, but there are several alternative strategies that have been investigated. Rhagoletis flies can be controlled and managed by a variety of insecticides and application meth- ods. Broad-spectrum insecticides, such as organo- phosphates and carbamates, have been applied in ultra low volume sprays, where contact and feed- ing toxicity of small droplets can cause fly mortal- ity (Mohammad & Aliniazee 1989; Hu et al. 2000). The enactment of the Food Quality Protection Act (FQPA) (1996) has placed severe restrictions on the use of these broad-spectrum insecticides, and future management of Rhagoletis flies will in- volve the use of insecticides that are not impacted by FQPA reassessment. Many of these new com- pounds have little or no contact toxicity; there- fore, they are often incorporated into a bait sta- tion or bait spray, in which mortality results after flies ingest significant quantities of insecticide. Painted spheres baited with ammonia com- pounds are highly attractive to tephritids and have been developed as bait stations. Spheres were first coated with a sticky material to trap flies (Prokopy 1975), but the need for decreased deployment and handling time necessitated find- ing an insecticide replacement (Duan & Prokopy 1995b). Studies evaluating the effects of insecti- cides, which were incorporated into the paint and sugar matrix that coated the surface of spheres, have been performed for Anastrepha ludens (Loew) (Prokopy et al. 2000b); Ceratitis capitata (Wiedemann) (Hu et al. 1998); and R. mendax and R. pomonella (Walsh) (Duan & Prokopy 1995b; Liburd et al. 1999; Ayyappath et al. 2000; Stelin- ski et al. 2001). Comparisons between baited spheres and azinphos-methyl sprays in a com- mercial apple orchard showed similar reductions in populations of R. pomonella (Prokopy et al. 2000a). However, in commercial blueberry fields insecticidal spheres are not currently used be- cause of the deployment density, lack of attractive selective lure, associated costs of products (i.e., spheres and residue extending agents), and labor requirements (i.e., monitoring and applying in- secticides to spheres) (Barry et al. 2004). Another alternative to broad-spectrum sprays are protein bait sprays which contain ammonia- based attractants, a feeding stimulant such as su- crose, and an insecticide. Protein bait sprays have been used to control outbreaks of Anastrepha ludens, Bactrocera dorsalis (Hendel), and Cerati- tis capitata since the 1950s in the United States (Steiner 1952; Moreno & Mangan 2003). However, development and evaluation of protein bait sprays on R. mendax has begun only recently. Pro- tein and ammonia-based attractants have been evaluated on Anastrepha spp. (Moreno & Mangan 2003), B. cucurbitae (Coquillett) (Fabre et al. 2003), B. dorsalis (Cornelius et al. 2000), R. cerasi (L.) (Katsoyannos et al. 2000), and R. pomonella (Duan & Prokopy 1992). Different concentrations of sugar feeding stimulants have been tested on A. suspensa (Loew) (Sharp & Chambers 1984), R. pomonella (Duan & Prokopy 1993), and R. mendax (Barry & Polavarapu 2004). Dowell (1994) outlined alternatives to a malathion bait spray, which had been the pre- ferred method in eradicating incipient infesta- tions of C. captitata. Further development of a re- placement has led to evaluation of insecticides classified as reduced risk. One compound that has already been incorporated into a bait spray for tropical and sub-tropical tephritid pests is spi- nosad, which was developed from the bacterium Saccharopolyspora spinosa Mertx and Yao. Feed- ing on baits containing spinosad has resulted in high mortality for A. ludens (Prokopy et al. 2000b), A. suspensa (King & Hennessey 1996), B. cucurbitae (Prokopy et al. 2003), and C. capitata (Peck & McQuate 2000; Vargas et al. 2001; Barry et al. 2003). Trials assessing toxicity have oc- curred for several of the non-organophosphate and non-carbamate compounds, such as delta- methrin, imidacloprid, and spinosad, on R. pomonella (Duan & Prokopy 1995a; Hu et al. 2000; Bostanian & Racette 2001; Reissig 2003) and acetamiprid, deltamethrin, fipronil, and imi- dacloprid on R. mendax (Barry et al. 2004). The reduced risk insecticide clothianidin, a neonicoti- noid, has not been evaluated on any tephritid spe- cies. Our goal was to identify the most effective con- centrations of insecticides present in bait that re- sulted in knockdown, mortality, and had the least feeding deterrence on R. mendax. Materials and Methods Insects Infested blueberries were collected near Chat- sworth, NJ, in the summer of 2002 and 2003. The rearing procedures of Ayyappath et al. (2000) 270 Florida Entomologist 88(3) September 2005 were used to obtain adult R. mendax. Briefly, in- fested berries were placed over moist sand for lar- vae to drop and pupate. Puparia were sifted from sand three-five weeks later and kept in a screen- house. Puparia were transferred to an incubator on 1 November 2002 and 2 November 2003, at 6°C with a photoperiod of 12:12 (L:D) to complete dia- pause. On 27 March 2003 and 30 March 2004 pu- paria were placed at 8°C. Periodically groups of puparia were transferred from 8 to 15°C for ap- proximately 8 d and then transferred to an incu- bator at 25°C with a photoperiod of 16:8 (L:D) un- til adult emergence, which occurred 25-45 d later. Adult flies were kept at 22°C and were provided a diet of sucrose and water (i.e., protein-starved). Flies used in assays were 7-13 d-old and allowed to acclimatize to experimental conditions in the laboratory for several hours before trials com- menced. Feeding Assay-Feeding for 10 s In the laboratory (21-23°C), a no-choice feeding test was used to evaluate survivorship of R. mendax on a control bait with baits containing three concentrations (4, 40, and 400 ppm or 0.0004, 0.004, and 0.04% [AI], respectively) of six insecticides: acetamiprid (technical, 30% [AI]; Cerexagri, King of Prussia, PA), clothianidin (technical, 49.17% [AI]; Arvesta, San Francisco, CA); deltamethrin and imidacloprid (technical 99.1, and 98.9% [AI], respectively; Bayer, Kansas City, MO); fipronil (technical 88% [AI]; Aventris Crop Science, Research Triangle Park, NC); and spinosad (technical, 90.4% [AI]; Dow Agro- Sciences, Indianapolis, IN). Solutions of each in- secticide concentration were prepared by weigh- ing the appropriate amount of technical and then adding it to the corresponding l:3-mixture of Sol- Bait (prepared as a 2x concentrate, USDA-ARS, Weslaco, TX) and water. (A l:3-mixture corre- sponds to a 1:4 mixture of GF-120 Fruit Fly Bait [Dow AgroSciences] to water.) After preparing the highest concentration, serial dilutions with a 1:3- mixture of SolBait were used to obtain mixtures with lower concentrations of insecticides. The con- trol was a l:3-mixture of SolBait to water contain- ing no insecticide. One 10-pl droplet of bait was placed on a white plastic lid (5.5 cm in diameter) located on top of a plastic cylinder (4 cm in diameter, 4 cm in height) in the center of a Plexiglas cage (30 cm X 30 cm X 30 cm). A fly was transferred to this lid and placed next to the droplet. After feeding on a droplet for 10 seconds, the fly was removed from the lid and placed inside a plastic cylinder (5 cm in diameter, 8.5 cm in height) containing water and sucrose. Flies that fed less than 10 seconds were dis- carded, unless it was determined that after the initial feeding a fly became incapable of feeding as a result of the insecticide (i.e., knockdown). A total of 30 flies were evaluated with the control and for each of three concentrations of insecticide (except deltamethrin which was not evaluated at 4 ppm). Flies were assessed for knockdown (i.e., immo- bile or incapable of walking) 1 h after the 10-s feeding. The number of dead, active, and incapac- itated flies was recorded after 1, 2, 3 and 4 d. Flies were characterized as dead if there was no pres- ence of visible body movement (i.e., no leg twitch), active if able to walk, and incapacitated if incapa- ble of walking (Hu et al. 2000; Reissig 2003). The number of living flies is represented by the sum of active and incapacitated flies. Feeding Assay-Feeding for 5 min In the laboratory (21-23°C), a no-choice test was used to evaluate feeding propensity of female R. mendax on protein bait containing 40 ppm of insecticide. Treatments were prepared by the methods described in the 10-s assay and included a control bait (without insecticide), clothianidin, fipronil, imidacloprid, and spinosad. One 10-pl droplet of a treatment was placed on a silk ficus leaf (Michaels, Irving, TX) that was placed on top of a plastic cylinder (4 cm in diameter, 4 cm in height) in the center of a Plexiglas cage (30 X 30 X 30 cm). Silk leaves were preferred to blueberry leaves because of the presence of chemical cues in the latter. One fly was transferred to the leaf within 1 cm of the droplet. Each feeding trial ended after 5 min if a fly was still present on a leaf or when a fly left a leaf after 5 s. (Flies that left a leaf in less than 5 s were not counted because they were believed to be in an agitated state.) In addi- tion, all flies had to feed a minimum of 1 s on the droplet. The amount of time that a fly spent feeding on a droplet was recorded. Flies were assessed for knockdown 1 h after feeding. Mortality was re- corded 1 and 4 d after feeding. Each fly was tested only once. A total of 28 replicates were completed for fly feeding and 20 replicates were completed for knockdown and mortality. One replicate was completed after a female fly had been tested on four protein baits incorporated with different in- secticides and the control bait. Exposure-4 h Survivorship and knockdown of flies was as- sessed to blueberry bushes treated with insecti- cidal baits. A control bait and four baits contain- ing 40 ppm insecticide of clothianidin, fipronil, imidacloprid, and spinosad, were prepared by the methods described in the 10-s Feeding Assay. Bait was applied with a handheld sprayer (30 Gunjet; Spraying Systems Co., Wheaton, IL) to deliver three 1-ml squirts at 30 psi to each of four three- yr-old blueberry bushes. This rate is equivalent to 9 liters/ha (0.95 gallons/acre). Three branches Scientific names italics only 271 (10-15 cm in length) were removed from each bush and placed inside a 250-ml Erlenmeyer flask in a Plexiglas cage (30 X 30 X 30 cm) that con- tained 20 flies (10 male, 10 female). The flask and branches were removed after 4 h. Flies were as- sessed for knockdown 3 h after introduction of treated branches and for mortality after 24 and 48 h. A total of 4 replicates were completed. Unlimited Access-Fresh and 1-d-old bait A no-choice assay evaluated fly mortality to bait containing the following insecticides: ace- tamiprid, clothianidin, fipronil, imidacloprid, and spinosad. Bait was prepared by adding enough technical insecticide to obtain 40 ppm [AI] in a mixture with GF-120 Fruit Fly Bait blank that did not contain spinosad (Dow AgroSciences). The control contained bait without the addition of in- secticide. For each treatment, one 10-pl droplet of bait was applied to 60 highbush blueberry leaves. Half of these leaves were removed within 10 min of application for use in fresh assays and the other half remained on bushes for 24 h before being col- lected. Three treated leaves of the same bait were placed inside each of ten 1-liter plastic containers with a screened lid, which contained a moist cot- ton ball. Five flies were then placed in each con- tainer, which constituted a replicate. Flies were assessed for knockdown after 1 h and mortality 24 and 48 h after the start of exposure. Ten replicates were completed for fresh and 1-d-old bait. This ex- periment occurred in the laboratory where tem- perature was 21-23°C. Statistical Analyses Knockdown and survivorship data from the 10- s Feeding Assay are presented in tabular and graphical form, respectively. For this feeding as- say, comparisons also were made between the control and each treatment with multiple chi- square tests after Bonferroni correction for the number of flies living versus dead after 4 d. In the 5-min assay feeding, duration was log trans- formed and analyzed by Fisher’s least significant different (LSD) tests (P = 0.05). Knockdown and mortality were analyzed by multiple chi-square tests after Bonferroni correction. Prior to analysis of variance (ANOVA), mortality and knockdown were arcsine-square root transformed in both the 4-h exposure assay and the unlimited access as- says (SAS Institute 1999). Means were separated by Fisher’s LSD tests (P = 0.05). Results Feeding Assay-10 s Insecticide type and concentration resulted in different survivorship of living (active + incapaci- tated) and active flies (Figs. 1 and 2, respectively). After 4 d, 97% of flies fed the control bait were liv- ing, which was significantly higher than all treat- ments except 4 ppm of acetamiprid, clothianidin, and imidacloprid, and 40 and 400 ppm of delta- methrin (%2, with Bonferroni correction, P = 0.05). Greater than 90% of flies were living 4 d after feeding on bait containing 4 ppm of acetamiprid, clothianidin, and imidacloprid (Fig. 1A-C); whereas less than 10% were living after feeding on baits with the same concentration of fipronil and spinosad (Fig. IE, F). Four days after feeding on bait containing 400 ppm of insecticide, there were 13, 43, and 87% flies categorized as living for treatments of clothianidin, acetamiprid, and deltamethrin, re- spectively (Fig. 1A, B, D). At 400 ppm of fipronil, spinosad, and imidacloprid in baits it took 1, 3, and 4 d after treatment to reach 0% survivorship, respectively (Fig. 1C, E, F). Large decreases in survivorship occurred between 1 and 4 d for all concentrations of spinosad and 4 ppm fipronil. Comparison of survivorship curves of living (active + incapacitated) with active flies appeared similar for treatments of clothianidin, delta- methrin, and fipronil (compare Fig IB with 2B, ID with 2D, IE with 2E, respectively), but dif- fered for the other three insecticides. The number of active flies increased from 1 to 4 d after feeding on bait containing 400 ppm acetamiprid, which indicated that some flies which had been incapac- itated were now active (Fig. 2A). The percent of living flies compared with active flies was 40 and 3%, respectively, 1 d after feeding on bait contain- ing 400 ppm imidacloprid, indicating that most (>90%) living flies were incapacitated (Fig. 1C and 2C, respectively). A large proportion of flies that fed on bait containing 40 and 400 ppm of spi- nosad were incapacitated, resulting in signifi- cantly fewer active than living flies 1-2 d after feeding (compare Fig. IF with 2F). More than 80% of flies were knocked down af- ter 1 h on baits containing 400 ppm of acetami- prid, clothianidin, and imidacloprid, with 30% knocked down for fipronil and spinosad (Table 1). Flies exposed to treatments of deltamethrin and control bait were not affected. Compared with the control bait there were significant higher knock- down effects after 1 h for baits containing 40 ppm of acetamiprid (20%), clothianidin (80%), and im- idacloprid (63%). Feeding Assay-5 min Protein baits containing insecticide had a sig- nificant effect on feeding duration (F = 65.79; df = 4,135; P < 0.0001; Fig. 3). Compared with the con- trol bait flies fed significantly less on baits con- taining imidacloprid and clothianidin, and fly feeding was not significantly different for baits containing fipronil and spinosad. Bait containing 272 Florida Entomologist 88(3) September 2005 m II 80 30 # HHJ Iva1 so m 48 21) 0 vX yX tX *.... ........ ...... ............... * A. ace&miiprid *X X X * .... ...»................• i FI iMmmihrki ▼X.............TX.............▼x .... ... . X X X V *■. B *........ ....•................. * O clothianidin E fiproail w...............w.................................. >* W 1r 40 HI 1) c tmidaeiopnd * X ▼ X X FVspinosad x X ! 2 3 4 't'sttw (days) 2 j 4 t'jftne (days) Fig. 1. Percent of flies that were living (active + incapacitated flies) after feeding for 10 s on a droplet of a given insecticide concentration. A) Acetamiprid, B) Clothianidin, C) Imidacloprid, D) Deltamethrin, E) Fipronil, F) Spi- nosad; (▼ = 4 ppm, O = 40 ppm, • = 400 ppm insecticide; X = control bait without insecticide) (Deltamethrin was not evaluated at 4 ppm.) imidacloprid was the only treatment that resulted in knockdown after 1 h that was significantly higher than the control (%2 with Bonferroni correc- tion, P = 0.05; Table 2). Flies that fed on fipronil were dead after one day and flies that fed on spi- nosad were all dead after four days; and both re- sults were significantly higher than the fly mor- tality in the control (%2 with Bonferroni correction, P = 0.05; Table 2). Exposure-4 h All insecticide treatments resulted in signifi- cantly higher knockdown than the control except spinosad after 3 h (F = 4.47; df = 4, 15; P = 0.014; Table 3). After 24 h, treatments had a significant effect on fly mortality (F = 3.58; df = 4, 15; P = 0.031; Table 3), with all insecticide treatments re- sulting in significantly higher mortality than the Scientific names italics only 273 I1 MO 8(1 m a SM s* Si} i 1“ "3 1 4(1 * w T*-- -- --- ~ - *x -- -- ~ -vX ~ .................. .* * =* .... ...... ..... #•......................♦ .... ..............*.......................* .A. acetamiprid ▼X *x B do i lua bi din • ♦........... so Ir 40 16 iniidacloprid ¥X X * E SproHjl x i ¥ siptnosad *• ■■■¥" Time (days) 1 8 Time ninys) Fig. 2. Percent of flies that were active (with incapacitated flies excluded) after feeding for 10 s on a droplet of a given insecticide concentration. A) Acetamiprid, B) Clothianidin, C) Imidacloprid, D) Deltamethrin, E) Fipronil, F) Spinosad; (▼ = 4 ppm, O = 40 ppm, • = 400 ppm insecticide; X = control bait without insecticide) (Deltamethrin was not evaluated at 4 ppm.) control except imidacloprid. After 48 h, there were no differences among the treatments including the control (F = 1.12; df = 4, 15; P = 0.385; Table 3). Survivorship Unlimited Access-Fresh bait Feeding on fresh bait containing insecticide re- sulted in significant fly knockdown after 1 h (F = 7.45; df= 5, 54; P < 0.0001; Table 4). Significantly more flies were knocked down on treatments of bait containing acetamiprid, clothianidin, and im- idacloprid compared with the control or treat- ments containing fipronil and spinosad. After 24 and 48 h, treatments had a significant effect on fly mortality (F = 14.86; df = 5, 54; P < 0.0001; and F = 15.65; df= 5, 54; P < 0.0001, respectively). After 24 h, fly mortality was significantly higher on fipronil and spinosad baits compared with the other three insecticide treatments, all of which 274 Florida Entomologist 88(3) September 2005 Table 1. Knockdown of flies 1 h after feeding for 10 S ON INSECTICIDAL BAIT. Treatment Fly knockdown (%) 400 ppm 40 ppm 4 ppm Acetamiprid 83 a 20 b 0 Clothianidin 96 a 79 a 0 Deltamethrin 0 c 0b - Fipronil 30 b 0b 0 Imidacloprid 100 a 63 a 3 Spinosad 30 b 0b 0 Control 0 c 0b 0 NS Values in the same column having the same letter are not significantly different (multiple chi-square tests after Bonfer- roni corrections; P = 0.05). n = 600 flies. were significantly higher than the control. These relative treatment relationships were the same after 48 h. Survivorship Unlimited Access-1-d old bait Treatments of 1-d old bait containing insecti- cides had a significant effect on fly knockdown (F = 8.49\df= 5, 54; P < 0.0001; Table 4). The highest numbers of flies were knocked down on acetami- prid, followed by imidacloprid, with both signifi- cantly higher than the control. The other three in- secticides were not different from the control in fly knockdown. After 24 and 48 h, treatments had a significant effect on fly mortality (F = 8.88; df = 5, 54; P < 0.0001; and F = 9.9; df = 5, 54; P < 0.0001, respectively). After 24 h, fly mortality was S 4I* ■; « ..... :------S...................... ..............:....... ....... ................ ..............: lapftiml Sunils;..! ClmfeiftitKlirt Cwrtmt Fig. 3. Duration of fly feeding (mean ± SE) on a bait containing 40 ppm insecticide. Flies were allowed to feed a maximum of 5 min on a 10-pl droplet. The control was bait without insecticide. Vertical bars with the same letter are not significantly different. (Fisher’s LSD test with log transformed data). (F = 65.79; df = 4, 135; P< 0.0001). Table 2. Fly Mortality and knockdown after 5 min EXPOSURE TO BAIT CONTAINING 40 PPM INSEC- TICIDE. Knockdown (%) Mortality (%) Treatment lh Id 4d Clothianidin 55 ab 15 ab 25 ab Fipronil 5 ab 100 a 100 a Imidacloprid 80 a 20 ab 50 ab Spinosad 5 ab 40 ab 100 a Control 0b 0b 5b Values in the same column having the same letter are not significantly different (multiple chi-square tests after Bonfer- roni corrections; P < 0.05). n = 100 flies. significantly higher on fipronil and spinosad baits compared with the control and the other insecti- cide baits. After 48 h, baits containing fipronil and spinosad resulted in significantly higher mortal- ity than baits containing either acetamiprid or clothianidin, with the latter two baits resulting in significantly higher mortality than either the con- trol bait or bait containing imidacloprid. Discussion Novel compounds were initially evaluated to find replacements for organophosphates and car- bamates. Results of several insecticides warrant future field trials to determine the efficacy of dif- ferent bait spray formulations for controlling R. mendax. Compounds differed in their ability to in- capacitate and kill flies. Depending on the insec- ticide chosen for inclusion in protein baits, the modes of action can be predominantly knockdown (acetamiprid, clothianidin, and imidacloprid) or kill (fipronil and spinosad). Table 3. Fly knockdown and mortality after 4 h EXPOSURE TO BAIT CONTAINING 40 PPM INSEC- TICIDE. Knockdown Mortality (%) (%) Treatment 3 h 24 h 48 h1 Clothianidin 11.3 ± 1.3 a 13.8 ± 3.8 a 25.0 ± 6.5 Fipronil 8.8 ± 4.3 a 26.3 ± 9.4 a 37.5 ± 14.4 Imidacloprid 5.0 ± 2.0 a 12.5 ± 4.3 ab 23.8 ± 7.2 Spinosad 3.8 ± 1.3 ab 16.3 ± 2.4 a 35.0 ± 7.4 Control 0.0 ± 0.0 b 3.8 ± 2.4 b 16.3 ± 5.9 Values in the same column having the same letter are not significantly different (Fisher’s LSD test, P = 0.05). ’NS, ANOVA, P > 0.05. n = 400 flies. Scientific names italics only 275 Table 4. Fly knockdown and mortality after Exposure to blueberry leaves containing bait droplets WITH 40 PPM INSECTICIDE. Experiment Treatment Knockdown (%) Mortality (%) lh 24 h 48 h Fresh1 Acetamiprid 12.0 ± 4.4 a 42.0 ± 6.9 b 68.0 ± 4.4 b Clothianidin 24.0 ± 8.3 a 58.0 ± 8.6 b 78.0 ± 4.6 b Fipronil 0.0 ± 0.0 b 80.0 ± 4.2 a 96.0 ± 2.6 a Imidacloprid 16.0 ± 4.0 a 44.0 ± 4.9 b 68.0 ± 8.5 b Spinosad 0.0 ± 0.0 b 80.0 ± 6.6 a 94.0 ± 3.0 a Control 0.0 ± 0.0 b 14.0 ± 2.1 c 30.0 ± 9.1 c 1-d-old1 Acetamiprid 16.0 ± 4.0 a 30.0 ± 7.4 b 56.0 ± 10.2 b Clothianidin 2.0 ±2.0 be 28.0 ± 8.0 b 58.0 ± 9.1b Fipronil 0.0 ± 0.0 c 60.0 ± 6.6 a 86.0 ± 5.2 a Imidacloprid 6.0 ± 3.0 b 10.0 ± 4.4 b 30.0 ± 7.4 c Spinosad 0.0 ± 0.0 c 68.0 ± 6.8 a 88.0 ± 4.4 a Control 0.0 ± 0.0 c 16.0 ± 10.2 b 22.0 ± 10.5 c For each experiment, values in the same column having the same letter are not significantly different (Fisher’s LSD test, P = 0.05). 1n = 300 flies. Bait sprays containing feeding stimulants (e.g., sucrose) have several advantages to conventional sprays. Lower concentrations of insecticide are needed in bait sprays than conventional sprays be- cause mortality is primarily from oral toxicity, which has lower LC50 thresholds than dermal tox- icity, and more insecticide is consumed because of the presence of feeding stimulants (e.g., sucrose) (Hu et al. 2000; Reissig 2003; Barry & Polavarapu 2004). Therefore, baits sprays can be applied at a lower rate of active ingredient per hectare than conventional sprays. The attraction and feeding re- sponses of flies to bait sprays have led to evalua- tions assessing their potential use as border sprays (Prokopy et al. 2003; Prokopy et al. 2004). Fly survivorship differed based on concentra- tion and type of insecticide used. As expected, higher concentrations of insecticide resulted in higher mortality of flies. At 400 ppm the shortest lag time between feeding and 100% mortality re- sulted from bait containing fipronil, followed by bait containing spinosad. The pyrethroid delta- methrin did not result in fly knockdown or mor- tality that was significant enough to warrant fur- ther evaluation on R. mendax, which was also the finding of Barry et al. (2004) investigating insec- ticidal coatings for spheres used in attract and kill of R. mendax. The neonicotinoids (acetami- prid, clothianidin, and imidacloprid) resulted in intermediate survivorship, performing better than deltamethrin, but not as well as spinosad or fipronil. Our findings are in agreement with Reis- sig (2003), who found the LC50 (with flies unable to walk considered dead) of imidacloprid and spi- nosad for R. pomonella to be approximately 11 ppm and between 3-10 ppm, respectively. Many published insecticide assays involve ex- posing flies to an insecticide treatment for several days in a small container to determine mortality. These conditions are likely to underestimate the concentration of insecticide needed for fly mortal- ity in the field. The importance of such studies is to determine the suitable type and range of activ- ity for insecticides to be further evaluated. In the current study we used three types of assays to evaluate the effects of insecticides: a variable feeding duration (up to 5 min), a fixed short dura- tion (Feeding Assay-10 s), and a fixed long dura- tion (Survivorship Unlimited Access). Each of these assays has limitations, but taken together supports the findings of the other assays. Sub-lethal effects of insecticides are known to manifest as a reduction in fecundity, measured in- directly from oviposition punctures by R. pomonella (Reissig 2003). In most of the assays in the current study, observations for knockdown oc- curred 1 h after a fly fed, but flies feeding on the neonicotinoids were often in that state much ear- lier and later, as evidenced by some flies being un- able to feed for the duration of the 10-s trial from becoming incapacitated. Liburd et al (2003) found insecticide-fed flies have lower levels of activity compared with a control. In our study flies that were knocked down often died, but some of the flies in this condition appeared no different than control flies after 1-2 d, apparently recovering from exposure to the insecticide. This finding leads us to suggest that there may be an optimal concentration for consumption to achieve the de- sired mortality. Measuring fly mortality in the context of field evaluations of insecticides contained in bait 276 Florida Entomologist 88(3) September 2005 sprays is one way to determine the effectiveness of knockdown. This would provide a realistic set- ting in which the effects of natural enemies could be evaluated on flies that are not completely dead, as well as other sub-lethal effects associated with a reduction in oviposition and larval presence. The results of future field trials can determine the effectiveness of bait sprays containing insecti- cides with different modes of action. Acknowledgments We thank Elizabeth Bender for rearing flies; Linda Tran-Barry for assisting with laboratory work; Rob Holdcraft, Andy Kyryczenko, and Jenn Hall for collecting flies, the late Daniel Moreno for supplying SolBait; and Arvesta, Bayer and Dow AgroSciences for providing insecticides. This re- search was funded in part by a USDA-CS- REES-Risk Avoidance and Mitigation Program and Hatch grant. References Cited Ayyappath, R., S. Polavarapu, and M. R. McGuire. 2000. Effectiveness of thiamethoxam-coated spheres against blueberry maggot flies (Diptera: Tephriti- dae). J. Econ. Entomol. 93: 1473-1479. 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Field tests of en- vironmentally friendly malathion replacements to suppress wild Mediterranean fruit fly (Diptera: Te- phritidae) populations. J. Econ. Entomol. 93: 280- 289. PROKOPY, R. J. 1975. Apple maggot control by sticky spheres. J. Econ. Entomol. 68: 197-198. Scientific names italics only 277 Prokopy, R. J., S. E. Wright, J. L. Black, X. P. Hu, AND M. R. MCGUIRE. 2000a. Attracticidal spheres for controlling apple maggot flies: Commercial-orchard trials. Entomol. Exp. Appl. 97: 293-299. Prokopy, R. J., I. Jacome, J. Pinero, L. Guillen, F. D. Fleischer, X. Hu, and M. Aluja. 2000b. Post- alighting responses of Mexican fruit flies (Dipt., Te- phritidae) to different insecticides in paint on attrac- tive spheres. J. Appl. Entomol. 124: 239-244. Prokopy, R. J., N. W. Miller, J. C. Pinero, J. D. Barry, L. C. Tran, L. Oride, and R. I. Vargas. 2003. Effectiveness of GF-120 fruit fly bait spray ap- plied to border area plants for control of melon flies (Diptera: Tephritidae). J. Econ. Entomol. 96: 1485- 1493. Prokopy, R. J., N. W. Miller, J. C. Pinero, L. Oride, N. Chaney, H. Revis, and R. I. Vargas. 2004. How effective is GF-120 fruit fly bait spray applied to border area sorghum plants for control of melon flies (Diptera: Tephritidae). Fla. Entomol. 87: 354- 360. REISSIG, W. H. 2003. Field and laboratory tests of new insecticides against the apple maggot, Rhagoletis pomonella (Walsh) (Diptera: Tephritidae). J. Econ. Entomol. 96: 1463-1472. SAS INSTITUTE. 1999. User’s manual, version 8.0. SAS Institute, Cary, NC. Sharp, J. L., and D. L. Chambers. 1984. Consumption of carbohydrates, proteins and amino acids by Anas- trepha suspensa (Diptera: Tephritidae) in the labora- tory. Environ. Entomol. 13: 768-773. STEINER, L. F. 1952. Fruit fly control in Hawaii with poi- soned sprays containing protein hydrolysate. J. Econ. Entomol. 45: 838-843. Stelinski, L. L., O. E. Liburd, S. Wright, R. J. Prokopy, R. Behle, and M. R. McGuire. 2001. Comparison of neonicotinoid insecticides for use with biodegradable and wooden spheres for control of key Rhagoletis species (Diptera: Tephritidae). J. Econ. Entomol. 94: 1142-1150. Vargas, R. I., S. L. Peck, G. T. McQuate, C. G. Jack- son, J. D. Stark, and J. W. Armstrong. 2001. Po- tential for areawide integrated management of Mediterranean fruit fly (Diptera: Tephritidae) with a braconid parasitoid and a novel bait spray. J. Econ. Entomol. 94: 817-825. US 20010046986A1 (i9) United States (12) Patent Application Publication (io) Pub. No.: US 2001/0046986 Al Miura et al. (43) Pub. Date: Nov. 29,2001 (54) METHOD FOR CONTROLLING FI TES (76) Inventors: Hiroyuki Miura, Nishitokyo-shi (JP); Atsuo Akayama, Tsukuba-shi (JP) Correspondence Address: WENDEROTH, LIND & PONACK, L.L.P. 2033 K STREET N. W. SUITE 800 WASHINGTON, DC 20006-1021 (US) (21) Appl. No.: 09/840,820 (22) Filed: Apr. 25, 2001 (30) Foreign Application Priority Data Apr. 26, 2000 (JP)................................... 131562/2000 Publication Classification (51) Int. Cl.7 ...................... A01N 43/54; A01N 47/28; A01N 43/78 (52) U.S. Cl................ 514/211.03; 514/212.08; 514/241; 514/269; 514/365; 514/351; 514/595 (57) ABSTRACT There is provided a method for controlling flies, that live in or come flying to livestock pens or poultry houses, by using a compound or salt thereof with an affinity for a nicotinic acetylcholine receptor of insects. Since having a very high activity against flies, the compound or salt thereof helps quickly and efficiently to control flies that live in or come flying to livestock pens or poultry houses. Also, since having an excellent control effect on flies that are resistant to conventional chemicals including a organo-phosphoric pes- ticide and a pyrethroid, the compound or salt thereof helps to prevent propagation, caused by flies, of various diseases to livestock, poultry and humans, and reduce discomfort of workers engaged in the livestock and poultry farming and a community. Still, it is possible with the present invention to keep livestock pens or poultry houses in a hygienic condi- tion, which may greatly contribute to the development of the livestock and poultry farming. 2/15/2017, EAST Version: 3.0.1.1 US 2001/0046986 A1 1 Nov. 29, 2001 METHOD FOR CONTROLLING FLIES FIELD OF THE INVENTION [0001] The present invention relates to a method for controlling flies, that live in or come flying to livestock pens (including bams, etc.,) or poultry houses such as chicken houses, by using one or more compounds or salts thereof having an affinity for a nicotinic acetylcholine receptor (hereinafter, sometimes abbreviated to as “NACR”) of insects. The present invention also relates to a composition for controlling flies that live in or come flying to livestock pens or poultry houses comprising the compounds or salts thereof. [0002] The compounds having an affinity for NACR used herein include insecticidal compounds with an agonist activ- ity or antagonist activity for NACR (hereinafter, sometimes abbreviated to as “NACRAtype insecticidal compounds”). DISCLOSURE OE THE PRIOR ART [0003] Examples of conventional agents for controlling flies in livestock pens or poultry houses include organo- phosphoric pesticides, synthetic pyrethroids, IGR (chitin- biosynthesis inhibitor), JHM (materials for juvenile hor- mone) and the like. However, repeated use of these limited chemicals results in various problems including the occur- rence of flies having resistance to the chemicals and delayed effect of, in particular, IGR and JHM on the flies. Thus, this is not a satisfactory situation. Also, synthetic pyrethroids are known to have a repellent activity. Therefor, even though flies are apparently reduced after chemical treatment, in fact they move to the surroundings of livestock pens or poultry houses. In some cases, it may make a community uncom- fortable. [0004] On the other hand, examples of the NACRA type insecticidal compounds include compounds described in EP-A 135956, EP-A 136636, EP-A 154178, EP-A 163855, EP-A 189972, EP-A 192060, EP-A 212600, EP-A 235725, EP-A 254859, EP-A 259738, EP-A 302389, EP-A 302833, EP-A 303570, EP-A 306696, EP-A 315826, EP-A 364844, EP-A 375907, EP-A 383091, EP-A 386565, EP-A 425978, EP-A 428941, EP-A 455000, EP-A 464830, EP-A 471372, EP-A 580553; DE-A 3639877, DE-A 3712307; JP-A 63-287764, JP-A 63-307857, JP-A 2-171, JP-A 2-207083, JP-A 3-128361, JP-A 3-157308, JP-A 3-169861, JP-A 3-218370, JP-A 3-220176, JP-A 3-246283, JP-A 3-255072, JP-A 3-279359, JP-A 4-9371, JP-A 4-154741, JP-A 5-271207, JP-A 6-183918, JP-A 7-179448, JP-A 7-278094; U.S. Pat. No. 4,918,086, U.S. Pat. No. 4,948,798, U.S. Pat. No. 5,034,404, U.S. Pat. No. 5,034,524, U.S. Pat. No. 5,039,686; WO 91/4965, WO 91/17659; FR-A 2611114; Brazil Patent Application No. 8803621 and the like. These NACRA type insecticidal compounds are sometimes referred to as neonicotinoid compounds, nitromethylene compounds, chloronicotinyl compounds and thianicotinyl compounds. Some of these compounds are marketed or under development. [0005] Some of these patent publications disclose that the compounds have the effect on agricultural insect pests and insanitary insects (e.g., JP-A 3-157308). However, none of them refer to control of flies in livestock pens or poultry houses. [0006] Moreover, a method for directly treating host ani- mals with these NACRA type compounds for extermination of fleas in mainly dogs and cats are described in, for example, JP-A8-92091 and JP-A2000-63271. However, no description is given with respect to control of flies that live in or come flying to livestock pens or poultry houses. [0007] Examples of the flies in livestock pens or poultry houses include flies such as Hypoderma bovis whose larva parasitizing the skin, or stinging and biting Stomoxys that directly inflict pain on livestock, and flies such as Musca domestics, Muscina stabulaus and Fannia canicularis that produce indirect detrimental effects on livestock. Such indi- rect detrimental effects include malodor caused by evolution of ammonia resulting from softening of feces by the larva and propagation of various diseases by the imagoes to livestock, poultry or humans. Examples of the diseases propagated by adult flies include Newcastle disease and influenza in chickens, infectious gastroenteritis and cholera in pigs, mastitis and epizootic stomatitis in cows, dysentery, cholera and alimentary intoxication (e.g., alimentary intoxi- cation caused by Salmonella, pathogenic Escherichia coli 0-157 and the like) in humans. Also, flies are considered to be filthy insect pests. Therefor, when flies come flying to houses surrounding livestock pens or poultry houses, troubles with a community may occur as a problem. Thus, it is essential for livestock farmers to achieve control of these flies. OBJECTS OF THE INVENTION [0008] However, according to the present situation involv- ing the above-described problem of the development of resistance, it is desired that a new type of agents having high activities for controlling flies are developed. [0009] The main object of the present invention is to provide a method for controlling flies, that live in or come flying to livestock pens or poultry houses. [0010] This objective as well as other objectives and advantages of the present invention will become apparent to those skilled in the art from the following description. SUMMARY OF THE INVENTION [0011] Under the above circumstances, the present inven- tors have studied intensively for searching a new type of an agent for controlling flies that live in or come flying to livestock pens or poultry houses. As a result, surprisingly, the present inventors have found that an NACRA type insecticidal compound has very high activity against these flies. As a result of the present inventors’ further study, the present invention has been completed. [0012] That is, according to the present invention, there is provided: [0013] (1) A method for controlling flies, that live in or come flying to livestock pens or poultry houses, comprising using a compound or salt thereof having an affinity for a nicotinic acetylcholine receptor of insects; [0014] (2) The method for controlling flies according to the above (1), wherein the compound or the salt thereof is a compound of the formula I, II or III: 2/15/2017, EAST Version: 3.0.1.1 US 2001/0046986 A1 2 Nov. 29, 2001 R1 I A CH2------N. .R2Y X (CH2)n I I A-----CHo------NY^YT A-----CH,------ N. N------R3 T X [0015] wherein A represents 6-chloro-3-pyridyl, 2-chloro- 5-thiazolyl, tetrahydrofuran-3-yl, 5-methyltetrahydrofuran- 3-yl, 3-pyridyl, 6-bromo-3-pyridyl, 3-cyanophenyl, 2-me- thyl-5-thiazolyl, 2-phenyl-5-thiazolyl or 2-bromo-5- thiazolyl; R1 represents hydrogen atom, methyl, ethyl, propyl, propenyl, propynyl, formyl, acetyl or methoxycar- bonyl; represents methyl, ethyl, amino, methylamino, N,N-dimethylamino, ethylamino, N,N-diethylamino, N-me- thyl-N-ethylamino, 1-pyrrolidinyl, N-methylformamide, N-methylacetamide or N-methyl-N-(methoxycarbony- l)amino; R3 represents a hydrogen atom, methyl, ethyl, propyl, propenyl, propynyl, formyl, acetyl or methoxycar- bonyl; X represents nitromethylene, nitroimino, cyanoimino or trifluoroacetylimino; Y represents a group represented by N (R4) (R4 is as defined with respect to R1) or sulfur atom; Z represents a group represented by N (R5) (R5 is as defined with respect to R1) or oxygen atom; and n is an integer of 2 or 3, or a salt thereof; [0016] (3) The method for controlling flies according to the above (1), wherein the compound is one or more compounds selected from the group consisting of clothiani- din (common name), nitenpyram (common name), imida- cloprid (common name), thiacloprid (common name), acetamiprid (common name), thiamethoxam (common name) and dinotefuran (common name); [0017] (4) The method for controlling flies according to the above (1), wherein the compound is chlothianidine (trade name); [0018] (5) The method for controlling flies according to the above (1), wherein the compound or salt thereof having an affinity for a nicotinic acetylcholine receptor of insects is sprinkled or sprayed in livestock pens or poultry houses; [0019] (6) The method for controlling flies according to the above (1), wherein the compound or salt thereof with an affinity for a nicotinic acetylcholine receptor of insects is applied to the inside of livestock pens or poultry houses; [0020] (7) The method for controlling flies according to the above (1), wherein a poisoned bait containing the compound or salt thereof with an affinity for a nicotinic acetylcholine receptor of insects is placed in livestock pens or poultry houses; [0021] (8) A composition for controlling flies, that live in or come flying to livestock pens or poultry houses, com- prising a compound or salt thereof having an affinity for a nicotinic acetylcholine receptor of insects; [0022] (9) The composition for controlling flies according to the above (8), wherein the compound or the salt thereof is a compound of the formula I, II or III shown in the above (2) or a salt thereof; [0023] (10) Use of a compound or salt thereof having an affinity for a nicotinic acetylcholine receptor of insects for manufacturing a composition for controlling flies that live in or come flying to livestock pens or poultry houses; and [0024] (11) Use according to the above (10), wherein the compound or the salt thereof is a compound of the formula I, II or III as shown in the above (2) or a salt thereof. DETAILED DESCRIPTION OF THE INVENTION [0025] Examples of the compounds having an affinity for NACR include the compounds described in the above patent publications. They can be produced according to the meth- ods described in the above patent publications, or their modifications. [0026] Among NACRA type insecticidal compounds, preferably, an insecticidal compound having an agonist activity for NACR are used. [0027] It is preferred to use a compound of the formula I, II or III: I R1 I A-----CH2------Nv .R2 T A-----CHo------NYYiT A-----CH2------ \. N----- R3 T X [0028] wherein the symbols are as defined above, or a salt thereof. [0029] It is more preferred to use clothianidin [(E)-l-(2- chloro-l,3-thiazol-5-ylmethyl)-3-methyl-2-nitroguanidine], nitenpyram [(E)-N-(6-chloro-3-pyridylmethyl)-N-ethyl-N'- methyl-2-nitrovinylidenediamine], imidacloprid [1-(6- chloro-3-pyridylmethyl)-2-nitroiminoimidazolidine], thia- cloprid [3-(6-chloro-3-pyridylmethyl)-2-cyanoimino-l,3- thiazolidin], acetamiprid [(E)-N4-(6-chloro-3- pyridylmethyl) -N2-cyano-N1-methylacetoamidine], thiamethoxam [3-(2-chloro-l,3-thiazol-5-ylmethyl)-5-me- 2/15/2017, EAST Version: 3.0.1.1 US 2001/0046986 A1 3 Nov. 29, 2001 thyl-4-nitroimino-l,3,5-perhydroxadiazine], and dinotefu- ran [1 -methyl-2-nitro-3-(3-tetrahydrofurylmethyl)guani- dine]. The structural formulae thereof are as follows: CH2----- N Me----- N I H •nno2 chlothianidine El Cl-------i 7------CII2------N Me----- N I H CHNOo Cl- nitenpirame ,r~Y_ r\<' ')-CR. N N Y imidacloprid ^ A ............. nno2 CH2----- N. thiacloprid Cl-------\ V------ CH2------N' Me Y ,0^ / acetamiprid r -CH2----- N. .N-Me T nno2 thiametoxane II CH2----- N Y:NNQ2 H dinotefuran [0030] Among the above, chlothianidine (trade name) is the most preferred. [0031] These compounds having an affinity for NACR sometimes react with acidic substances or basis substances, to form salts. These salts may be agricultural chemicals acceptable salts. More specifically, examples of the basic substances include alkali metals such as sodium, potassium, lithium, etc.; alkaline earth metals such as calcium, magne- sium, etc.; inorganic bases such as ammonia, etc.; and organic bases such as pyridine, collidine, triethylamine, triethanolamine, etc.; and the like. Examples of the acidic substances include inorganic acid such as hydrochloric acid, hydrobromic acid, hydroiodic acid, phosphoric acid, sulfuric acid, perchloric acid,etc.; and organic acid salts such as formic acid, acetic acid, tartaric acid, malic acid, citric acid, oxalic acid, succinic acid, benzoic acid, picric acid, meth- anesulfonic acid, p-toluenesulphonic acid, etc. [0032] These compounds or salts thereof having an affinity for NACR have low toxicity against mammals including livestock and humans and low fish toxicity, and thus are excellent eco-friendly fly-controlling agents. [0033] Flies targeted for control by the present invention in livestock pens or poultry houses refer to all flies that live in or come flying to livestock pens or poultry houses. Examples of the flics include those of Muscidac such as Musca domestics, Fannia canicularis, Muscina stabulans, Stomoxys calcitrans, etc.; Calliphoridae such as CaUiphora lata, Callophora vicina, Aldrichina grahami, Lucilia illus- Iris, etc.; and Sarcophagidae such as Boellcherisca pereg- rina, etc. The most troublesome flies in the livestock farming are flies of Museidae mentioned above. [0034] These flies targeted for control include every stage of generation, i.e., eggs, larvae, pupas and imagoes. Particu- larly, the compound or a salt thereof used in the present invention exhibits a high activity against larvae and ima- goes. That is, the preferred embodiment of the present invention include a method for controlling larvae and ima- goes of flies that live in livestock pens or poultry houses, or imagoes of flies that come flying thereto. [0035] The compounds or salts thereof having an affinity for NACR described above exhibits a strong insecticidal activity against Diptera in general, enabling simultaneous control of mosquitoes and gnats of Nematocera, and horse- flies of Brachycera. [0036] Typical examples of livestock pens or poultry houses to which the compounds or salts thereof having an affinity for NACR described above arc used, include a cow house, a pig pen and a poultry farm. Examples thereof also include places for bleeding other livestock and pets (e.g., horses, sheep, goats, camels, buffalo, donkeys, rabbits, deer, reindeer, minks, dogs and cats) and poultry (e.g., ducks, turkeys and quails). Further, “the livestock pens or poultry houses” of the present invention includes sites for accumu- lating feces and urine, garbage, spilt feed and the like discharged from these facilities, which may be located indoor or outdoor around livestock pens or poultry houses. [0037] For using the compounds or salts thereof having an affinity for NACR described above for controlling flies in livestock pens or poultry houses, they can be formulated in the form of conventional agrochemical formulations. That is, one or more (preferably, one or more but three or less) of the compounds or salts thereof having an affinity for NACR as active components are dissolved or dispersed in an appropriate liquid carrier, or mixed or absorbed with an appropriate solid carrier to prepare emulsion, solution, microemulsion, flowable preparation, oily preparation, water soluble powder, wettable powder, granular wettable powder, powder, granule, microgranule, smoking agent, tablet, microcapsule, spray, EW, ointment, poisoned bait, and the like. 2/15/2017, EAST Version: 3.0.1.1 US 2001/0046986 A1 4 Nov. 29, 2001 [0038] If necessary, to these preparations may be added emulsifier, suspension, spreader, penetrant, humecant, muci- lage, stabilizer and the like. They are prepared by a per se known method. [0039] Examples of the liquid carrier (solvent) to be used include water, alcohols (e.g., methyl alcohol, ethyl alcohol, n-propyl alcohol, isopropyl alcohol, ethylene glycol, etc.), ketones (e.g., acetone, methyl ethyl ketone, etc), ethers (e.g., dioxane, tetrahydrofuran, ethylene glycol monomethyl ether, diethylene glycol monomethyl ether, propylene glycol monomethyl ether, etc.), aliphatic hydrocarbons (e.g., kero- sene, lamp oil, fuel oil, machine oil, etc.), aromatic hydro- carbons (e.g., benzene, toluene, xylene, solvent naphtha, methylnaphthalene, etc.), halogenated hydrocarbons (e.g., dichloromethane, chloroform, carbon tetrachloride, etc.), acid amides (e.g., N,N-dimcthylformamidc, N,N-dimcthy- lacetamide, etc.), esters (e.g., ethyl acetate, butyl acetate, glycerol fatty acid ester, etc.), nitrites (e.g., acetonitrile, propionitrile, etc.) and the like. One or more of these carriers (preferably, one or more, but three or less) may be mixed at a suitable ratio according to particular used. [0040] Examples of the solid carriers (diluent, extender) include vegetable powder (e.g., soy flour, tobacco powder, wheat flour, wood flour, etc.), mineral powder (e.g., clay such as kaolin, bentonite, acid clay, etc.; talc such as talc powder, pyrophyllite powder, etc.; silica such as diatom earth, mica powder, etc.; and the like), alumina, sulfur powder, activated carbon and the like. One or more of these carrier (preferably, one or more, but three of less) may be mixed at a suitable ratio according to particular use. [0041] Examples of ointment bases include polyethylene glycol; pectin; polyol ester of higher fatty acid such as glyceryl monostearate ester, etc.; cellulose derivative such as methyl cellulose, etc.; sodium alginate; bentonite; higher alcohol; polyol such as glycerin, etc.; vaseline; white vase- line; liquid paraffin; lard; various vegetable oils; lanolin; anhydrous lanolin; hardened oil; resin and the like. These may be used alone or in combination thereof (preferably, one or more, but three or less). If necessary, various surfactants can be appropriately added. [0042] Examples of the surfactants to be used as emulsi- fier, spreader, penetrant, dispersant or the like include non- ionic and anionic surfactants such as soaps, polyoxyethylene alkylaryl ethers [e.g., Noigen (trade name) and E*A 142 (trade name) manufactured by Daiichi Kogyo Seiyaku Co., Ltd.; Nonal (trade name) manufactured by Toho Chemical], alkylsulfates [e.g., Emal 10 (trade name) and Emal 40 (trade name) manufactured by Kao Corporation], alkylsulfonates [e.g., Neogen (trade name) and Neogen T (trade name) manufactured by Daiichi Kogyo Seiyaku Co., Ltd.; Neopellex (trade name) manufactured by Kao Corporation], polyethylene glycol ethers [e.g., Nonipol 85 (trade name), Nonipol 100 (trade name) and Nonipol 160 (trade name) manufactured by Sanyo Chemical Industries, Ltd.], polyhy- dric alcohol esters [e.g., Tween 20 (trade name) and Tween 80 (trade name) manufactured by Kao Corporation] and the like. They may be used whenever they are needed. Further, the compounds or salts thereof having an affinity for NACR may be used in combination with one or more other insec- ticides (e.g., pyrethroid insecticides, organophosphorus insecticides, carbamate insecticides, phenylpyrazole insec- ticides, phenylpyrrole insecticides, benzoylurea insecticides, dibenzoylhydrazine insecticides, Nereistoxin insecticides, juvenile hormone substances, natural insecticides, and the like), miticides (acaricides), nematocides, fungicides (e.g., copper fungicides, organochlorine fungicides, organosulfur fungicides, phenolic fungicides, benzimidazole fungicides, EBI, melanin-biosynthesis inhibitors and acrylate fungi- cides), synergists, attractants (e.g., sugar, vinegar, molasses, milk powder, etc.), repellents, pigments, and the like accord- ing to particular use. [0043] Normally, the compounds or salts thereof having an affinity for NACR are formulated in an amount of about 0.01 to 95% by weight, preferably about 0.1 to 80% by weight based on the total weight of the composition for controlling flies of the present invention. More specifically, when the composition to be used is in the form of emulsion, solution, wcttablc powder, granular wcttablc powder or the like, suitably, the compounds or salts can be formulated in an amount of about 1 to 80% by weight, preferably about 1 to 20% by weight based on the total weight of the compo- sition. When the composition to be used is in the form of oily solution, powder or the like, suitably, the compounds or salts can be formulated in an amount of about 0.1 to 50% by weight, preferably about 0.1 to 20% by weight based on the total weight of the composition. When the composition to be used in the form of granule or the like, suitable, the com- pounds or salts can be formulated in an amount about 5 to 50% by weight, preferably about 1 to 20% by weight based on the total weight of the composition. [0044] Normally, other active components to be combined in the composition for controlling flies of the present inven- tion (e.g., insecticides, acaricides (miticides) and/or fungi- cides) may be used in an amount ranging from about 0.01 to 80% by weight, preferably about 1 to 20% by weight based on the total weight of the composition. [0045] The content of additives other than the above active components varies depending on a particular kind or content of the active component, a particular form of the composi- tion and the like. Normally, they are formulated in an amount of about 0.001 to 99.9% by weight, preferably about 1 to 99% by weight based on the total weight of the composition. More specifically, normally, it is preferred to add about 1 to 20% by weight, preferably about 1 to 15% by weight of a surfactants, about 1 to 20% by weight of a fluidization aid, and about 1 to 90% by weight, preferably about 1 to 70% by weight of a carrier based on the total weight of the composition, respectively. Still more specifi- cally, for producing a solution, normally, it is preferred to about 1 to 20% by weight, preferably about 1 to 10% by weight of a surfactant and about 20 to 90% by weight of water based on the total weight of the composition. Upon use, the emulsion, wettable powder (e.g., granular wettable powder) or the like may be diluted with water and the like to a suitable concentration (e.g., about 100 to 5,000-fold) before sprinkling or spraying. [0046] Representative examples of insecticides, miticides (acaricides) and fungicides, which may be used by mixing or combining with the compounds or salts thereof having an affinity for NACR in the method for controlling flies of the present invention, are as follows: EPN, acephate, isoxathion, isofenphos, isoprocarb, etrimfos, oxydeprofos, quinalphos, cadusafos, chlorethoxyfos, chlorpyrifos, chlorpyrifos-me- thyl, chlorofenvinphos, salithion, cyanophos, disulfoton, 2/15/2017, EAST Version: 3.0.1.1 US 2001/0046986 A1 5 Nov. 29, 2001 dimethoate, sulprofos, diazinon, thiometon, tetrachlorvin- phos, tebupirimfos, trichlorphon, naled, vamidothion, pyra- clophos, pyridafenthion, pirimiphos-methyl, fenitrothion, fenthion, phenthoate, fosthiazate, butathiofos, prothiofos, propaphos, profenofos, phosalone, fosthiazate, malathion, methidathion, metolcarb, monocrotophos, BPMC, XMC, alanycarb, ethiofencarb, carbaryl, carbosulfan, carbofuran, xylylcarb, cloethocarb, thiodicarb, triazamate, pirimicarb, fenoxycarb, fenothiocarb, furathiocarb, propoxur, bendio- carb, bcnfuracarb, mcthomyl, acrinathrin, imiprothrin, ethofenprox, cycloprothrin, sigma-cypermethrin, cyhalo- thrin, cyflutyrin, cypermethrin, silatiuofen, teiluthrin, delta- methrin, tralomethrin, fenvalerate, fenpropathrin, flucythri- nate, fluvalinate, Ilufenoprox, Iluproxyfen, llumethrin, prallethrin, beta-cyfluthrin, benfluthrin, permethrin, cartap, thiocyclam, bensultap, avermectin, emamectin-benzoate, clofentezine, chlorfluazuron, cyromazine, diafenthiuron, dienochlor, dichlorvos, diflubenzuron, spynosyn, sulflura- mid, teflubenzuron, tebufenozide, tebufenpyrad, hydro- prene, vaniliprole, pymetrozine, pyridaben, pyriproxyfen, pyrimidifen, fipronil, fenazaquin, fenpyroximate, fluazuron, flucycloxuron, flufenoxuron, buprofezin, hexaflumuron, hexythiazox, milbemycin, metoxadiazone, lufenuron, levamisol, chlorphenapyr, NC-184, etoxazole, IBP, ampro- pylfos, edifenphos, chlorthiophos, tolclofos-methyl, fosetyl, ipconazole, imazalil, imibenconazole, etaconazole, epoxi- conazole, cyproconazole, diniconazole, difenoconazole, tet- raconazole, tebuconazole, triadimenol, triadimefon, triti- conazole, triforine, bitertanol, viniconazole, fenarimol, fenbuconazole, fluotrimazole, furconazole-cis, flusilazole, flutriafol, bromuconazole, propiconazole, hexaconazole, pefurazoate, penconazole, myclobutanil, metconazole, cabendazin, debacarb, prothiocarb, benomyl, maneb, TPN, isoprothiolane, iprodione, iminoctadine-albesil, iminocta- dine-triacetate, ethirimol, etridiazole, oxadixyl, oxycar- boxin, oxolinic acid, ofurace, kasugamycin, carboxin, cap- tan, clozylacon, chlobenthiazone, cyprodinil, cyprofuram, diethofencarb, dichloflnanid, diclomezine, zineb, dimethiri- mol, dimethomorph, dimefluazole, thiabendazole, thiopha- nate-methyl, tliifluzamide, tecloftalam, triazoxide, tricla- midc, tricyclazolc, tridcmorph, triflumizolc, validamycin A, hymexazol, pyracarbolid, pyrazophos, pyrifenox, pyrimethanil, pyroquilon, ferimzone, fenpiclonil, fenpropi- din, fenpropimorph, fthalide, furametpyr, furalaxyl, fluazi- nam, furcarbanil, fluquinconazole, fludioxonil, flusulfamide, flutolanil, butiobate, prochloraz, procymidone, probenazole, benalaxyl, benodanil, pencycuron, myclozolin, metalaxyl, metsulfovax, methfuroxam, mepanipyrim, mepronil, kresoxim-methyl, azoxystrobin, SSF-126 and carpropamid. [0047] For controlling flies in livestock pens or poultry houses by using the compounds or salts thereof having an affinity for NACR, the above-described composition com- prising the compounds or salts thereof having an affinity for NACR can be sprinkled or sprayed over habitats and gath- ering places of flies in livestock pens or poultry houses by a per se known method, whereby the composition is allowed to come into contact with flies or to be ingested by flies to exterminate them. The composition may be sprinkled or sprayed in the air to exterminate flying adult flies in live- stock pens or poultry houses, or sprinkled or sprayed over places where mainly larvae live (e.g., feces, spilt feed, etc.). The preferred composition includes that in the form of water soluble powder, solution, wettable powder, granular wet- table powder, powder or emulsion. The amount of the composition to be applied may be varied broadly according to particular application time, application place, application method, or the like. In general, the composition is preferably applied so that the amount of the active component (the compounds or salts thereof having an affinity for NACR) per 1 m2 is in the range of about 0.1 mg to 10 g, preferably about 1 mg to 1 g. In the case where the composition is usually diluted by water upon use, the composition may be diluted so that the final concentration of the active component ingredient is in the range of about 0.1 to 10,000 ppm, preferably about 10 to 500 ppm. [0048] Alternatively, it is possible to exterminate flies by applying the above composition to the inside of livestock pens or poultry houses, or by placing the composition as poisoned bait therein. More specifically, for example, the above composition may be applied to or sprinkled or sprayed over a wall, door, pillar, window, floor, ceiling, beam and the like of livestock pens or poultry houses, if this is convenient, in combination with one or two or more attractants (e.g., sugar, vinegar, molasses, milk powder, bran, etc.) (preferably, one or more, but three or less). Also, plates treated with the chemicals (e.g., 10x2(1 cm to 1 x2 m) may be taken in livestock pens or poultry houses, and placed or suspended. Preferably, the liquid composition has con- centration of from 0.001 to 20% by weight of the active component, and attractants are added in an amount of from 1 to 20% by weight. As poisoned bait, there can be used a mixture of attractants including sugar, vinegar, molasses, milk powder and bran (one or two or more, preferably, one or more but three of less), and the above composition containing one or more compounds or salts thereof having an affinity for NACR (preferably, water soluble powder, wettable powder, granular wettable powder and the like) diluted by water. The preferred concentration of the active component and attractants is the same as in the case of the application to the inside of livestock pens or poultry houses. Poisoned bait thus prepared can be used by putting it in, for example, a tray of an appropriate size (e.g. 10x10 cm to 1x2 m). It may be effective to lay paper or cloth in the tray for attracting the imagoes. Also, for example, a bottle made of PET provided with lateral holes may be used as a simple trap by putting an adequate amount of the poisoned bait (50 to 500 ml) therein. [0049] Since having a very high activity against flies, the compound or salt thereof having an affinity for NACR of the present invention helps quickly and efficiently to control flies that live in or come flying to livestock pens or poultry houses. Also, since having an excellent control effect on flies that show resistance to conventional chemicals including a organo-phosphoric pesticide and a pyrethroid, the com- pound or salt thereof helps to prevent propagation, caused by flies, of various diseases to livestock, poultry and humans, and reduce discomfort of workers engaged in the livestock and poultry farming and a community. Still, it is possible with the present invention to keep livestock pens or poultry houses in a hygienic condition, which may greatly contrib- ute to the development of the livestock and poultry farming. [0050] The following preparation example, examples and test example further illustrate the present invention in detail but not to be construed to limit the scope of the present invention. In the following preparation example, examples and test example, all the parts and percents are by weight unless otherwise stated. 2/15/2017, EAST Version: 3.0.1.1 US 2001/0046986 A1 6 Nov. 29, 2001 1 R1 I A-----CH2------NL .R2T Preparation Example 1 [0051] Preparation of 50% chlothianidine granular wettable powder Clothianidin Alkenylsulfate Radiolite #200 (trade name) were added 50 parts, 10 parts and 100 parts (total) (CHj),, [0052] The above materials were thoroughly mixed, added with 25 parts of tap water, and kneaded in a mortar. Then, the resultant kneaded materials were granulated into col- umn-shaped granules by a pellet mill (Kikusui Seisakusho Ltd., RG-5W) using a screen with a diameter of 0.8 mm. The resultant granules were dried at 60° C. for 1 hour to obtain clothianidin 50% granular wettable powder. EXAMPLE 1 [0053] Preparation of liquid composition for application to livestock pen and poultry house Ten grams of clothianidin 50% granular wettable powder and 10 g of sugar were mixed. To the mixture was added water and the total volume was adjusted to 100 ml. EXAMPLE 2 [0054] Preparation of liquid composition for poisoned bait [0055] To 100 g of a 1:5:10 mixture of sugar, vinegar and milk powder (feed for weaning baby pigs) and 10 g of clothianidin 50% granular wettable powder was added water and the mixture was adjusted to the total volume of 500 ml to obtain a liquid composition. Test Example 1 [0056] Insecticidal test by oral administration to Musca domestica [0057] A clothianidin solution in alcohol was diluted with 5% granulated sugar solution to prepare a 50 ppm liquid composition of clothianidin. [0058] Female Musca domestica imagoes (ten flies) of a sensitive strain kept without water for 3-4 hours were anesthetized and lightly fixed to a pressure sensitive adhe- sive sheet which was cut into strips. After the test insects came out from the anesthetic, 1 ^L of the liquid composition (containing 0.05 fag of clothianidin) was released from an injection needle attached to a micro dropper so that the test insects inhaled the composition. After 24 hours, it was confirmed that all flies were dead. What is claimed is: 1. A method for controlling flies, that live in or come flying to livestock pens or poultry houses, comprising using a compound or salt thereof having an affinity for a nicotinic acetylcholine receptor of insects. 2. The method for controlling flies according to claim 1, wherein the compound or the salt thereof is a compound of the formula I, II or III: A-----CH,------\. AT A-----CH2------N. N-----R3 T X wherein A represents 6-chloro-3-pyridyl, 2-chloro-5-thiaz- olyl, tetrahydrofuran-3-yl, 5-methyltetrahydrofuran-3-yl, 3-pyridyl, 6-bromo-3-pyridyl, 3-cyanophenyl, 2-methyl-5- thiazolyl, 2-phcnyl-5-thiazolyl or 2-bromo-5-thiazolyl; R1 represents hydrogen atom, methyl, ethyl, propyl, propenyl, propynyl, formyl, acetyl or methoxycarbonyl; R2 represents methyl, ethyl, amino, methylamino, N,N-dimethylamino, ethylamino, N,N-diethylamino, N-methyl-N-ethylamino, l-pyrrolidinyl, N-methylformamide, N-methylacetamide or N-methyl-N-(methoxycarbonyl)amino; R3 represents a hydrogen atom, methyl, ethyl, propyl, propenyl, propynyl, formyl, acetyl or methoxycarbonyl; X represents nitrometh- ylene, nitroimino, cyanoimino or trifluoroacetylimino; Y represents a group represented by N (R4) (R4 is as defined with respect to R1) or sulfur atom; Z represents a group represented by N (R5) (R5 is as defined with respect to R1) or oxygen atom; and n is an integer of 2 or 3, or a salt thereof. 3. The method for controlling flies according to claim 1, wherein the compound is one or more compounds selected from the group consisting of clothianidin (common name), nitenpyram (common name), imidacloprid (common name), thiacloprid (common name), acetamiprid (common name), thiamethoxam (common name) and dinotefuran (common name). 4. The method for controlling flies according to claim 1, wherein the compound is clothianidin (common name). 5. The method for controlling flies according to claim 1, wherein the compound or salt thereof having an affinity for a nicotinic acetylcholine receptor of insects is sprinkled or sprayed in livestock pens or poultry houses. 6. The method for controlling flies according to claim 1, wherein the compound or salt thereof with an affinity for a nicotinic acetylcholine receptor of insects is applied to the inside of livestock pens or poultry houses. 7. The method for controlling flies according to claim 1, wherein a poisoned bait containing the compound or salt thereof with an affinity for a nicotinic acetylcholine receptor of insects is placed in livestock pens or poultry houses. 8. A composition for controlling flies, that live in or come flying to livestock pens or poultry houses, comprising a compound or salt thereof having an affinity for a nicotinic acetylcholine receptor of insects. 2/15/2017, EAST Version: 3.0.1.1 US 2001/0046986 A1 7 Nov. 29, 2001 9. The composition for controlling flies according to claim 11. Use according to claim 10, wherein the compound or H, wherein the compound or the salt thereof is a compound the salt thereof is a compound of the formula I, II or III: of the formula I, II or III: A-----CH2------N. .R'Y A-----CH2------NL -R;Y (CH2)1 I I A-----CH2------V ,YT X (CH2)n I I A-----CH2------NY/Yr A-----CH2------Nv -N-----R-Y A-----CH2------ ISL .N-----R-Y wherein A represents 6-chloro-3-pyridyl, 2-chloro-5-thiaz- olyl, tetrahydrofuran-3-yl, 5-methyltetrahydrofuran-3-yl, 3-pyridyl, 6-bromo-3-pyridyl, 3-cyanophcnyl, 2-mcthyl-5- thiazolyl, 2-phenyl-5-thiazolyl or 2-bromo-5-thiazolyl; R1 represents hydrogen atom, methyl, ethyl, propyl, propenyl, propynyl, formyl, acetyl or methoxycarbonyl; R2 represents methyl, ethyl, amino, methylamino, N,N-dimelhylamino, ethylamino, N,N-diethylamino, N-methyl-N-ethylamino, 1-pyrrolidinyl, N-methylformamide, N-methylacetamide or N-methyl-N-(methoxycarbonyl)amino; R3 represents a hydrogen atom, methyl, ethyl, propyl, propenyl, propynyl, formyl, acetyl or methoxycarbonyl; X represents nitrometh- ylene, nitroimino, cyanoimino or trifluoroacetylimino; Y represents a group represented by N (R4) (R4 is as defined with respect to R1) or sulfur atom; Z represents a group represented by N (R5) (R5 is as defined with respect to R1) or oxygen atom; and n is an integer of 2 or 3, or a salt thereof. 10. Use of a compound or salt thereof having an affinity for a nicotinic acetylcholine receptor of insects for manu- facturing a composition for controlling flies that live in or come flying to livestock pens or poultry houses. wherein A represents 6-chloro-3-pyridyl, 2-chloro-5-thiaz- olyl, tetrahydrofuran-3-yl, 5-methyltetrahydrofuran-3-yl, 3-pyridyl, 6-bromo-3-pyridyl, 3-cyanophenyl, 2-methyl-5- thiazolyl, 2-phenyl-5-thiazolyl or 2-bromo-5-thiazolyl; R1 represents hydrogen atom, methyl, ethyl, propyl, propenyl, propynyl, formyl, acetyl or methoxycarbonyl; R2 represents methyl, ethyl, amino, methylamino, N,N-dimethylamino, ethylamino, N,N-diethylamino, N-methyl-N-ethylamino, 1-pyrrolidinyl, N-methylformamide, N-methylacetamide or N-methyl-N-(methoxycarbonyl)amino; R3 represents a hydrogen atom, methyl, ethyl, propyl, propenyl, propynyl, formyl, acetyl or methoxycarbonyl; X represents nitrometh- ylene, nitroimino, cyanoimino or trifluoroacetylimino; Y represents a group represented by N (R4) (R4 is as defined with respect to R1) or sulfur atom; Z represents a group represented by N (R5) (Rs is as defined with respect to R1) or oxygen atom; and n is an integer of 2 or 3, or a salt thereof. * * * * 4= 2/15/2017, EAST Version: 3.0.1.1 Copy with citationCopy as parenthetical citation