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WO 2017/184673 Al 26 October 2017 (26.10.2017) W !P O PCT

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WO 2017/184673 Al 26 October 2017 (26.10.2017) W !P O PCT (12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization International Bureau (10) International Publication Number (43) International Publication Date WO 2017/184673 Al 26 October 2017 (26.10.2017) W !P O PCT (51) International Patent Classification: (72) Inventors: LU, Albert; 7250 NW 62ND AVE, PO BOX CI2N 15/82 (2006.01) 552, Johnston, Iowa 50131-0552 (US). WU, Gusui; 7250 NW 62ND AVE, PO BOX 552, Johnston, Iowa 5013 1-0552 (21) International Application Number: (US). PCT/US2017/028258 (74) Agent: BECKER, James J.; PIONEER HI-BRED INTER (22) International Filing Date: NATIONAL, INC., 7100 N.W. 62nd Avenue, P.O. Box 19 April 2017 (19.04.2017) 1014, Johnston, Iowa 50131-1014 (US). (25) Filing Language: English (81) Designated States (unless otherwise indicated, for every (26) Publication Language: English kind of national protection available): AE, AG, AL, AM, AO, AT, AU, AZ, BA, BB, BG, BH, BN, BR, BW, BY, BZ, (30) Priority Data: CA, CH, CL, CN, CO, CR, CU, CZ, DE, DJ, DK, DM, DO, 62/324,652 19 April 2016 (19.04.2016) US DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, HN, (71) Applicants: PIONEER HI-BRED INTERNATIONAL, HR, HU, ID, IL, IN, IR, IS, JP, KE, KG, KH, KN, KP, KR, INC. [US/US]; 7100 N.W. 62nd Avenue, P.O. Box 1014, KW, KZ, LA, LC, LK, LR, LS, LU, LY, MA, MD, ME, MG, Johnston, Iowa 5013 1-1014 (US). E I DU PONT DE MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, OM, NEMOURS AND COMPANY [US/US]; Chestnut Run PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SA, SC, Plaza, 974 Centre Road, P.O. Box 2915, Wilmington, SD, SE, SG, SK, SL, SM, ST, SV, SY,TH, TJ, TM, TN, TR, Delaware 19805 (US). TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW. (54) Title: INSECTICIDAL COMBINATIONS OF POLYPEPTIDES HAVING IMPROVED ACTIVITY SPECTRUM AND USES THEREOF El = IIiilii 00 (57) Abstract: The disclosure provides nucleic acids, and variants and fragments thereof, derived from strains of Bacillus thuringiensis encoding variant polypeptides having increased pesticidal activity against insect pests, including Lepidoptera and Coleopteran. Particu lar embodiments of the disclosure provide isolated nucleic acids encoding pesticidal proteins, pesticidal compositions, DNA constructs, o and transformed microorganisms and plants comprising a nucleic acid of the embodiments. These compositions find use in methods for controlling pests, especially plant pests. o [Continued on nextpage] WO 2017/184673 Al llll II II 11III I II I II I i Illl III II I II (84) Designated States (unless otherwise indicated, for every kind of regional protection available): ARIPO (BW, GH, GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, ST, SZ, TZ, UG, ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, RU, TJ, TM), European (AL, AT, BE, BG, CH, CY, CZ, DE, DK, EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, LU, LV, MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK, SM, TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, GN, GQ, GW, KM, ML, MR, NE, SN, TD, TG). Published: — with international search report (Art. 21(3)) — before the expiration of the time limit for amending the claims and to be republished in the event of receipt of amendments (Rule 48.2(h)) — with sequence listing part of description (Rule 5.2(a)) INSECTICIDAL COMBINATIONS OF POLYPEPTIDES HAVING IMPROVED ACTIVITY SPECTRUM AND USES THEREOF CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to United States Provisional Application No. 62/324652, filed April 19, 2016, which is hereby incorporated herein in its entirety by reference. REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY A sequence listing having the file name "71 16WOPCT_Sequencel_isting.txt" created on April 15, 2016 and having a size of 266 kilobytes is filed in computer readable form concurrently with the specification. The sequence listing is part of the specification and is herein incorporated by reference in its entirety. FIELD The present disclosure relates to recombinant nucleic acids that encode pesticidal polypeptides having insecticidal activity against corn earworm and/or fall armyworm and/or an improved spectrum of pesticidal activity against insect pests. Compositions and methods of the disclosure utilize the disclosed nucleic acids, and their encoded pesticidal polypeptides, to control plant pests. BACKGROUND Insect pests are a major factor in the loss of the world's agricultural crops. For example, armyworm feeding, black cutworm damage, or European corn borer damage can be economically devastating to agricultural producers. Insect pest-related crop loss from European corn borer attacks on field and sweet corn alone has reached about one billion dollars a year in damage and control expenses. Traditionally, the primary method for impacting insect pest populations is the application of broad-spectrum chemical insecticides. However, consumers and government regulators alike are becoming increasingly concerned with the environmental hazards associated with the production and use of synthetic chemical pesticides. Because of such concerns, regulators have banned or limited the use of some of the more hazardous pesticides. Thus, there is substantial interest in developing alternative pesticides. Biological control of insect pests of agricultural significance using a microbial agent, such as fungi, bacteria, or another species of insect affords an environmentally friendly and commercially attractive alternative to synthetic chemical pesticides. Generally speaking, the use of biopesticides presents a lower risk of pollution and environmental hazards, and biopesticides provide greater target specificity than is characteristic of traditional broad- spectrum chemical insecticides. In addition, biopesticides often cost less to produce and thus improve economic yield for a wide variety of crops. Certain species of microorganisms of the genus Bacillus are known to possess pesticidal activity against a broad range of insect pests including Lepidoptera, Diptera, Coleoptera, Hemiptera, and others. Bacillus thuringiensis {Bt) and Bacillus papilliae are among the most successful biocontrol agents discovered to date. Insect pathogenicity has also been attributed to strains of B. larvae, B. lentimorbus, B. sphaericus (Harwook, ed., ((1989) Bacillus (Plenum Press), 306), and B. cereus (WO 96/1 0083). Pesticidal activity appears to be concentrated in parasporal crystalline protein inclusions, although pesticidal proteins have also been isolated from the vegetative growth stage of Bacillus. Several genes encoding these pesticidal proteins have been isolated and characterized (see, for example, U.S. Patent Nos. 5,366,892 and 5,840,868). Microbial insecticides, particularly those obtained from Bacillus strains, have played an important role in agriculture as alternatives to chemical pest control. Recently, agricultural scientists have developed crop plants with enhanced insect resistance by genetically engineering crop plants to produce pesticidal proteins from Bacillus. For example, corn and cotton plants have been genetically engineered to produce pesticidal proteins isolated from strains of Bt (see, e.g., Aronson (2002) Cell Mol. Life Sci. 59(3):41 7-425; Schnepf et al. (1998) Microbiol Mol Biol Rev. 62(3)775-806). These genetically engineered crops are now widely used in American agriculture and have provided the farmer with an environmentally friendly alternative to traditional insect-control methods. In addition, potatoes genetically engineered to contain pesticidal Cry toxins have been sold to the American farmer. While they have proven to be very successful commercially, these genetically engineered, insect- resistant crop plants provide resistance to only a narrow range of the economically important insect pests. Accordingly, there remains a need for new Bt toxins with an improved spectrum of insecticidal activity against insect pests, e.g., toxins which are improved active against insects from the order Lepidoptera and/or Coleoptera. In addition, there remains a need for biopesticides having activity against a variety of insect pests and for biopesticides which have improved insecticidal activity. SUMMARY Compositions and methods are provided for impacting insect pests. More specifically, the embodiments of the present disclosure relate to methods of impacting insects utilizing nucleotide sequences encoding insecticidal peptides to produce transformed microorganisms and plants that express an insecticidal polypeptide of the embodiments. In some embodiments, the nucleotide sequences encode polypeptides that are pesticidal for at least one insect belonging to the order Lepidoptera. In some aspects nucleic acid molecules and fragments and variants thereof are provided, which encode polypeptides that possess pesticidal activity against insect pests (e.g. SEQ ID NO: 4 , SEQ ID NO: 6, SEQ ID NO: 8 , SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, SEQ ID NO: 28, SEQ ID NO: 30, SEQ ID NO: 32, SEQ ID NO: 34, SEQ ID NO: 36, SEQ ID NO: 38, SEQ ID NO: 40, SEQ ID NO: 42, SEQ ID NO: 44, and SEQ ID NO: 46, and encoding the polypeptide of SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 2 1, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, SEQ ID NO: 29, SEQ ID NO: 3 1, SEQ ID NO: 33, SEQ ID NO: 35, SEQ ID NO: 37, SEQ ID NO: 39, SEQ ID NO: 4 1, SEQ ID NO: 43 or SEQ ID NO: 45, respectively). The wild-type {e.g., naturally occurring) nucleotide sequence of the embodiments, which was obtained from Bt, encodes an insecticidal peptide. The embodiments further provide fragments and variants of the disclosed nucleotide sequence that encode biologically active {e.g., insecticidal) polypeptides.
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