CA1195922A - Diabroticite pest control - Google Patents

Abstract

ABSTRACT
Diaproticiticidal effectiveness and diabroticite specificity of insecticides is enhanced by admixture with dry cucurbitacinous products derived, preferably, from hybrid Cucurbitales plant tissue. Hybrid plants bred from (wild) bitter Cucurbitaceae species and (domesticated) non-bitter species (e.g., Cucurbita andreana x C. maxima or C. texana x C. pepo) provide high yields of plant fruits with high concen-trations of cucurbitacins, which are attractant/arrestants and feeding stimulants for diabroticites. In one embodiment, dried ground fruits of Cucurbita hybrid plants are admixed with essentially minute quantities of insecticides to provide diabroticite-specific insecticide compositions allowing for effective, economical, environmentally safe control of diabroticites such as corn rootworm beetles.

Description

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BACKGROUND
The present invention relates generally to pest control and more particularly to the enhancement of the effectiveness of insecticidal compositions in the control of leaf beetles of the genus Diabrotica.
Among the most economically significant agricultural pests in the united States are the so-called "leaf beetlesi' of the Order Coleoptera, Superfamily Chrysomeloidea~ Family Chrysomelidae. Within tribe Luperini of the subfamily Galeru-clnae are those particularly important beetles of the genusDiabrotica which are generally referred to as cucumber beetles~
The spotted cucumber beetles Diabrotica undecimpunta howardi Barber and D. u. undecimpunctata Mannerheim, the banded cucumber beetle D. balteata LeConte, the western corn rootworm D. vir~ifera LeConte, and the striped cucumber beetle Acalymma vittata (Fabricius) and its western relative A. trivittata (Mannerheim) are important pests of squash, cucumbers, and muskmelons in North America. D. undecimpunctata howardi, D. longicornis and D. virgifera are severe pests of corn as well. For the most part, the adult cucumber beetle feeds on the blossoms, fruits, and leaves of the host plants while the larval forms (developing from eggs deposited in the soil during the previous fall) feed on plant roots.
The attacks of the cucumber beetles in the U.S.
are conservatively estimated to cause damage and expenditures for control aggregating hundreds of millions of dollars annually. ~ecent reports have indicated, for example, that 25 million acres of corn land are routinely treated for corn rootworm beetle infestation. The chemical treatments --applied as sprays, but principally as 50il treatments forlarval control and to combat adults at egg laying times -~

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are organophosphorus and carbamate insecticides. Applied at the rate of one or two pounds per acre, the total cost of such treatment exceeds about $500 million per year.
The widespread distribution of such huge quantities of toxic insecticides for control of crop destructive insects has been the cause for significant concern by environmentalists.
Many insecticides used in corn rootworm beetle control (e.g., phorate, terbufos, fonofos and fensulfothion) are so potentially toxic to birds and other wildlife that there are governmental requirements that they be employed in large quantities only as subsoil treatments. Certain insecticidal compositions which could effectively control diabroticites in lesser concentrations are also undesirably lethal to all insect life in the areas in which they are broadcast and hence destroy useful insects as well as the "target" beetles.
There has existed, therefore, a need in the art for new systems for effective control of adult and larval diabroticite beetles which are ~oth more economical and less envîronmentally hazardous than those presently employed.
~dO Of interest to the background of the present inven-tion is a coevolutionaxy association or "link" between dia-brot:icite beetles and certain plants of the so-called "gourd Order", Cucurbitales, which has been the subject of intensive study by the co-inventors and their co-workers. See, Metcalf, et al~ P~N.A~S., 77, pp.3769-3772 (1980). The family~ Cucur-bitaceae, within this Order comprises about 100 genera, including some 900 species of plantsO Many domesticated species are familiar and significant food sources, including the following: Cucumis sativus (cucumber), C. anguria (gherkin);
C. melo ~muskmelon, including the cantaloupe, Persian, casaba, and honeydew melons); Citrullus lanatus (watermelon~, Cucurbita (summer squash, pumpkin, yellow-flavored gourd); C.

mixta (pumpkin squash); C7 moschata (pumpkin, winter squash);
C. maxima /pumpkin, winter squash); C~ ficifolia (fig leaf gourd); Lagenaria siceraria ~white-flowered gourd); Benincasa hispida (Chinese preserving melon); Sechium edule (chayote);
Sicana odorifera (cassabanana); Momordica balsamina (balsam apple, Chinese bitter melon); Trichosanthes cucumeroides (snake gourd); Cyclanthera pedaclita (lard fruit).
At least 100 species within about 30 genera of Cucurbitaceae are known ~o contain small quantities oE one or more of about 20 related oxygenated tetracyclic ~riterpene compounds, the cucurbitacins, which are responsible for a characteristic bitter taste in plant tissue and which therefore make the fruits useless as foodstuffs. Indeed, the cucurbitacins are among the most intensely bitter sub-stances known to man and can be detected by humans in dilutions as low as 1 part per billion. The compounds are also rela-tively toxic in high concentrations, causing severe poisoning in sheep and cattle consuming various bitter Cucumis and Cucurbita fruits.
It is currently believed that cucurbitacins triter-penes were the subject of evolutionary selection as a vehicle or protection of wild Cucurbitaceae against attack by verte-br~te and invertebrate herbivores. Remarkablyl however, certain species o North American diabroticite beetles are neither repelled nor poisoned by the cucurbitacins Rather, the substances act as kairomones which strongly attrac~
and arrest the beetles and initiate compulsive feeding res-ponses. See, e.g., Contrardi; Physis (Florence~, 18, ppO 331-347 (1939); Chamblis/ et al., Science, 153, 1392-1393 (1966);
DaCosta, et al., Hortic. Science, 6, pp. 340-342 (1971);
Howe, et al., EnvironO Entomol., 5, pp. 1043-1048 (1976), Bogwa~l et al. Indian J. Entomol., 29, ppO 349-352 (1967), and Sharma, et al., Environ. ~ntomol., 2, pp. 154 156 (1973).
As an example of the analysis of the attractance/-arrestance of cucumber beetles to bitter Cucurbitaceae, Sharma, et al., supra provided a study of the attractiveness of cut fruits of fifteen wild and domesticated plant species to D. undecipunctata howardi. Included were fruits of six genera (Citrullus, Cucumis, Curcurbita, Lagenaria, Luffa and Momordica). At~ractant/arrestant substances were prin-]0 cipally found to be present in species of Citrullus, Cucumisand Cucurbita genera and the direct arrival of beetles at cut bitter fruits was construed to indicate the ability of the beetles to detect the presence of cucurbitacins and their glycosides without direct contact. Feediny on bitter fruits continued until the fruits became dehydrated. Among the conclusions of the study were proposals to use beetles to screen for cucurbitacins in other species and to use attractant cucurbitacins "...in population assessment and in lurin~ the beetle to an insecticide or to an adhesive".
~0 The co-inventors and their co-workers provided Eurtheï analysis of attrac~ant/arrestant and compulsive eeeding ~timulation properties Eor 18 species of the Cucurbita genus vis-a-vis males and females of six species of diabroticite beetles, using pure cucurbitacins selec~ively derived from plant leaves, fruits, blossoms and roots by chloroform extrac-tion and thin layer chromatography~ (See~ Metcalf, et al., P~N~AoS~ I supra.) Specific body parts of diabroticites were characteri~ed as having cucurbitacin l'receptor" func~
tions and correlations were drawn between evolution patterns of particular beetle species and Cucurbita ;`;

species based on the cucurbitacin content of plants and cucurbitacin sensitivity of beetles.
Despite -the correlations drawn in the prior art between cucurbitacin kairomones and diabroticite beetles, there h~ve been no reports o~ any use of cucurbitacins in any aspect of the management of diabroticites. This is likely due to a number of factors, including: the absence of appreciable quantities of curcurbitacins in non-bitter domesticated Cucurbitale species; the low concentrations of the compounds in plant tissues of even the wild bitter species (ranging from trace amounts in leaves to about mg per gram in fruits of even the bitterest wild species);
the undependability of wild species of Cucurbitale as sources of bitter fruits (many species having photoperiod dependent fruiting); the consequent difficulty in providing lar5~e, constant source of fresh fruits; and the rela~ive difficulty of ~xtracting and purifying cucurbitacins from plant tissues for us in pest management programs.

BRIEF SUMMARY
According to the present invention, the diabroticiti~
cidal effectiveness and diabroticite specificity of insecticides is enhanced by admixture of the compositions with dry cucur bitacinous products derived from bitter Cucurbitale plant tissue, preferably from the fruits of hybrid plants bred by crossing bitter, wild species with domes~icated, non~
bitter species.
Dry granulated "diabroticite baited" insecticide compositions of the invention comprise a major proportion o dried bitter Cucurbitale fruit and a minor proportion of insecticide (from 0.01 to 10~ by weight). Such dry insecti-cide compositions permit effective con-trol of corn rootworm beetle populations in cornfields despite the use of approxi-mately as little as l/lOOth the quantity of insecticide chemical ordinarily employed. Remarkably, the dried insecti-cidal compositions (which contain plant tissues having less than about ten percent moisture) are effective as attractant/-arrestants and compulsive feeding stimulants despite the absence of molsture or similar volatilizing component and retain their effectiveness for long periods of time. Moreover, these dried insecticides appear to be highly selective in killing diabroticites as apposed to other, potentially useful, insects.
Preferred bitter Cucurbitale plant tissues for use in preparing the dried products o the invention include fruits of hybrid plants bred by crossing bitter, wild Cucur-bitale species with non-bitter domesticated speciesO Examples of presently preferred hybrids are those resulting from the cross of Cucurbita andreana with C. maxima and G. texana with C. pepo (zucchini).
Other aspects of the present invention will be

2.0 b~ter understood upon consideration of the following detailed description.

DETAILED DESCRIPTION
As employed herein and applied to plant tissues, the term "cucurbitacinous" shall mean containing one or more of the tetracyclic triterpenes, cucurbitacins A, B~
C, D~ E, F, G, H, I, J, K , L or R and the.ir glycosides.
Structures, formula names and physical properties Eor all the above cucurbitacins (except "R" which difers from "G'l and "H" only in the absence of an hydroxy group at position 24) are set forth in The Merck Index, 9th Edo I monographs 2609-2614 -~ 6 -~
~ ~D~ ~ ~

(Merck & Co., ~ahwayl N~J. 1976)~ As employed herein the term "insec~icide" shall mean and include a chemical compound or mixture of compounds which, alone or together with additives, ad~uvants and/or carriers, is useful in the control of insect populations either directly ~e.g., by killing) or indirectly (e.g., by causing sterility)~ As illustrative examples of insecticides there are included the highly toxic "natural"
arsenic compounds, such as Paris Green, fluorine compounds such as cryolite and the zinc and mercury compounds which cause breakdown of insect digestive tract tissue. Also included are the contact insecticides such as nicotine, rotenone, pyrethrum and its derivatives and analogs, as well as the petroleum oils. The term also comprehends such halogenated hydrocarbons as~ endrin; aldrin and its epoxide, dieldrin; heptachlor; DDT; BHC and its isomer, lindane;
chlordane; methoxychlor; DDD or TDE; and the polychlorinated biphenylsO Organophosphate compounds which may be mentioned include malathion, parathion, TEPP, schradan, demeton and dimethoate. Carbamates which may be mentioned include carbaryl ~0 and m~thyl carbamateO Also included by way of example are the orgallic thiocyanates such as Thani~e, Lethane 384 and r~lethane 6n and the chlorinated phenols such as penta- and tetrachlorophenolsO "Diabroticiticidal" shall refer to insecticide activity as applied to the control of insects oE the genus ~iabrotica.
As employed herein, the term l'hybrid'l shall designate the ofspring of different breeds, varieties, species or genera and shall include, but not be limited to, first genera-tion (Fl) offspring. Also included, therefore, are subsequent hybrid generations and inbred lines which are at least in part developed by hybridi~ation procedures.

As set forth in detail in the publication of the co~inventors and their co-worker appearing in J. AmerO SocO
Hort. Science, 105, pp. 838-842 (mailed December 11; 19~0), it has been determined that while tissues of species of wild~ bitter Cucurbitaceae of the genus Cucurbita frequently have moderate cucurbitacln contents, the fruits of such plants are frequently produced in quite low yield, both in terms of number of fruit and average fruit weigh~. Domesti-cated Cucurbita species, on the other hand, generally provide high numbers of heavy plant fruit which contain no detectable amounts of cucurbitacins. Careful hybridization of bitter species (e.g., C. andreana or C~ texana) with domesticated non-bitter species (e.g., C. maxima cv. Macre or C~ pepo cv. Zucchini) results in Fl hybrid generations of plants which substant;ally retain the high fruit yield charac~er-istics of the non-bitter parental species as well as the moderate cucurbitacin contents of the bitter parental species.
More particularly, a C. texana x C. pepo hybrid produced semibush plants with leaves, fruits and blossoms ~0 resembling C. pepo cv. Zucchini. Sample 6-plant plots produced an average of 98 fruits (average weight 0.73 Icg) while the C. texana parent produced a much lower quantity of Eruits having average weights of 0.10 kg. In a slmilar manner, a C. andreana x C. maxima hybrid produced long-vined plants with leaves, fruits and blossoms resembling C~ maxima.
A 6 plant plot produced 84 fruits with an average weight of 3.90 kg. The bitter C. andreana parent, again, produced fruit in much lower quantities with an average weight of only 0.50 kg~ Yields oE fruit of domesticated parent species were not measured.
Spectrophotometric analysis of chloroform extracts of leaves, fruits, roots and blossoms of both the parental - B -~L~Q ~

species and hybrids for cucurbi~acins B, D, E~ E-glycoside and I were carried out and the results are reported in Table I, below.

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2n TABLE I

Cucurbitacin content (mg per g fresh wt) Species Organ B D D I E-glycoside C. andreana leaf 0,15 0.12 f~uit 2.7~ 0.42 blossom Q.l9 0.12 root 0.58 0.51 o ~
C. andreana x C. maxima leaf 0.56 0.23 fruit 1.17 0.09 blossom 0.16 0.17 .
root 0.26 0.09 CO texana leaf trace trace trace fruit 0.07 U.36 0.75 blossom 0.24 0.15 0.50 root 0.18 0.08 0~3g C. texana x C. pepo leaf trace trace trace f r uit 0.15 b',ossom 0.10 0.l4 trace root 0.09 0.04 trace C. maxima no cucurbltacins detected in frui~ and rco~
C. pe~o no cucurbitacins detected in fruit and root Acre sized plots of the above-noted Fl hybrids have been grown and tested for cucurbitacin content. The C. andreana x C~ maxima hybrid fruits contained about 0.13%
by weight cucurbitacin B and D and were obtained in yields of about 30,000 pounds per acre~ The C. texana x C. pepo hybrid fruits contained about 0.06~ cucurbitacin E and its glycoside and were obtained in yields of about 15,000 pounds per acre.
The above~noted Fl hybrids of bitter Cucurbita species with non-bitter species are seen to provide, for the first time, a dependable source of large quantities of cucurbitacins. Clearly, other Fl hybrids can be e~pected to be developed using other bitter species of Cucurbita (e.g~, C. ecuadorensis Cutl. & Whit., C~ foetidissima ~IBK, C. martinenzii Bailey, C. okeechobeensis Bailey, C. palmata Wats, C. palmeri Bailey, and C. pedatifolia Bailey) and non-bitter species (e.g., F. ficifolia Bouche, C. mixta Pangalo, and C. moschata Duchesne ex Poir~ In a like manner, the problems attending provision of reliable sources of 2~ cucurbitacins may be addressed by generating Fl hybrids oE bitter species of the genera Cucumis and Citrullus with correspondin~ high fruit yielding non-bitter species. Finally, it is anticipated that among F2 and subsequent generations of the above hybridization procedures there may occur true breeding Cucurbitale populations which will provide -the advantages of the Fl hybrids without the drawback of annual hybridization processing.
The value of the present invention in the context of diabroticite management and, more particularly, in enhanc-ing the efectiveness of insecticides in the control ofdiabroticites is illustrated by the following examples oE

the preparation and use of dried granulated insecticide compositions including cucurbitacinous tissue of Cucurbitale plants.

Plant frui~s were harvested from first generation C. andreana x C. maxima and C. texana x C. pepo hybrids.
Whole fruits were sliced into 10 to 50 gram pieces and air dried at from 35 to 60C for 4 to 7 days. The dried fruits were then ground in a food processing mill to a 30 to 60 mesh size.
Granules were then surface-coated with insecticides in varying concentrations (from about 0.01 to Ool weight percent) by dissolving the insecticide in a minimal amount of acetone and mixing the acetone solution with the granules in a roller drum for about 5 minutes.
A variety of insecticide compositions were formed us.ing both hybrid fruit types and various insecticides includ-ing pyrethroids (envalerate, permethrin and de~amethr.in) ~() carbamates ~methomyl, carbaryl and carbofuran) and organophos-phates (trichlorofon, malethion, isofenfos and chlorpyrifos).

EX~MPLE 2 Dried, ~Idiabroticite baited" insecticide compositions prepared according to Example 1 were broadcast as granular baits for the control of corn rootworms in fields of sweetcornO
The compositions were applied in a variet of doses ranging from about 10 pounds per acre to 100 pounds per acre (10-100 kg/hc). Typical results of these diabroticite control pro-cedures were as follows.
A C~ texana x C. pepo frult granular compositionadmixed with O.:L percent by weight methomyl killed approximately ~ 12 -2;2 63,000 western and southern corn rootworm beetles per acre within 20 hours of appllcation at a rate of 30 pounds per acre. Total methomyl applied was 13.5 grams per acre and the result was an approximately 85 percent reduction of the pretreatment population. At a dosage of 10 pounds per acre (4.5 grams of methomyl/acre) the population reduction in 20 hours was 62 percent. Another composition providing another sweetcorn plot with 30 grams of methomyl per acre resulted in the killing of approximately 150,000 beetles over a period of 20 hoursO
Compositions containing 0.05 percent decamethrin killed large numbers of beetles when applied at dosages ranging from 0.45 to 1.35 grams of insecticide per acre.
~ pparently due to the fibrous nature of Eruit tissue of first generation hybrids of C. texana x C. pepo, compositions employing such tissue were generally more effec-tive than compositions using C. andreana x C. maxima tissue and having the same insecticide chemical content.
Insecticide compositions remained effective in ~0 stimulating compulsive Eeeding behavior with consequent killing oE beetles for two weeks or more.
Expectedly, compositions including equal quantities of pyrethroids were generally more effective at equal applica-tion dosages than compositions including carbamates. The latter were~ in turn, more effective than those including organophosphate insecticide chemicals.
Remarkably, the combination of Cucurbitale plant tissue with the insecticide chemicals appeared uniformly to increase the selectivity of the chemicals Eor diabroticites and at no time were any significant quantities of other dead insec-ts found among beetles killed by the composltionsO

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I~ is apparent from the above examples ~hat effective diabroticiticidal compositions can be readily provided without the need for laborious extraction and purification of the attractant/arrestan~ cucurbitacins from plant tissue. Nor is it necessary for effective diabroticite control to contin-uously provide cucurbitacinous plant tissue in the form of fresh cut plant parts. Owing to the remarkable ability of dried cucurbitacinous tissue to remain active as a diabroti-cite attractant/arrestant and compulsive feeding stimulant despite the absence oE appreciable amounts of water or other volatilizing components, there can now be provided stora~e stable (non-spoilingj diabroticiticidal compositions which can be prepared far in advance of use and which are easily handledr transported and applied in bulk quantities.
The amount of moisture which may be retained in the cucurbitacinous plant tissue component of insecticide compositions of the invention may be sub]ect to same degree of variation. While it i5 preerable from the standpoint o~ bulk handling and long term storage to retain less than about ten weight percent moisture content, greater moisture contents may be acceptable, especially if this facilitates uniform admixture with insecticides or iE spoi:Lage retardants can be added during composition formulation.
The insecticidal compositions of the above examples were formed by first developing the plant tissue constitllent in dried granular form and then admYxing the granules with insecticide to provide granules "coatedl' with insecticideO
It is expected that the advantages of the invention will similarly attend a more intimate mixture of plant tissue and insecticide prior to granulation with consequent greater uniformity of distribution of insecticide throughout granules.

Granular formulations may also include inert bulking agents such as dried corn cobs, if desired. The granule size oE
the insecticide composi~ions of ~he above examples ranged from about 30 to about 60 mesh. The optimal size of granules for use in diabroticite control may be subject to variation depending on particular conditions in the field treated and whether the granules are to be applied on or below the soil or as very finely divided dusts on plant surfaces.
The above examples illustrate that a significant enhancement of diabroticiticidal effectiveness of insecticides attends admixture with cucurbitacinous plant tissue when the active insecticide chemical comprises as little as 0.01 weight percent of the total insecticide composition. It should be readily understood b~ those skilled in the art that effective "diabroticite baited" compositions may include an even lesser proportion of active insecticide chemical (e.g., 0.005 weight percent) and. depending on the particular in5ecticide selec~ed and the quantity necessary to kill target beetles, could also include a substantially higher proportion (e.~.~ 50 weight percent) of insecticide chemical.
As a practical ~atter, the advantages of the use of cucur-bitaclnous Cucurbitale plant tissue are most extensive when the "major" proportion of plant tissue in the insecticide composition is significantly higher than the "minor'i proportion of insecticide chemical therein, iOe., 90 weight percent plant tissue: 10 weight percent insecticide chemical, or greater. Few advantages are presently seen to attend employing the compulsive feeding stimulant cucurbitacins to induce ingestion of higher doses oE insec-ticide than are required to kill or sterilze the insects.
While the above, driedl "diabroticite baited"

insecticide compositions were formed through use of fruits 2~

of Cucurbitale plants, it will be understood ~ha-t other cu~urbitacinous plant parts may also be employed. Use of plant fruits is, of course, highly preferred owing to the relatively ]ow quantities of cucurbitacins generally present in leaves, stems and roots, especially after fruit development has gone to completion and other plant parts are in diminished size. As a rule, it is preferred to employ Cucurbitale plant tissue having a cucurbitacin concentration in the range of 0.1 to 3.0 or more mg/gram of fresh weightO
While tissue obtained from wild, bitter Cucurbitale plants can be used in practice of the invention, the most highly preferred source of cucurbitacinous plant tissue is the fruit of hybrids bred from bitter species of Cucurbitale and especially Cucurbita genera) crossed with non-bitter species which produce large quantities of large fruits.
Especially preferred~ also, are hybrids formed using non-bitter parental species which have fruits of a relatively highly fibrous type such as zucchini fruits and those hybrids ~ormed using bitter parental species which have relatively ~0 higher cucurbitacin B and E concentrations, than, for example, cucurbitacin D or L concentrations.

Numerous modifications and variations in the inven-tion as above illustrated are expected to occur to those skilled in the art and consequently only such limitations as appear in the appended claims should be placéd on the scope of the invention.

Claims (11)

THE EMBODIMENTS OF THE INVENTION FOR WHICH
AN EXCLUSIVE PROPERTY OR PRIVILEGE IS
CLAIMED ARE DEFINED AS FOLLOWS:

1. An insecticide composition for use in control of diabroticite populations, said composition comprising the admixture of a major proportion by weight of dried cucur-bitacinous Cucurbitale plant tissue and a minor proportion by weight of a diabroticiticidal insecticide.

2. A composition according to claim 1 wherein the Cucurbitale plant tissue is hybrid plant tissue.

3. A composition according to claim 2 wherein the hybrid plant tissue is Cucurbita hybrid plant tissue.

4. A composition according to claim 3 wherein the Cucurbita hybrid plant is a first generation hybrid of the cross between C. andreana and C. maxima.

5. A composition according to claim 3 wherein the Cucurbita hybrid plant is a first generation hybrid of the cross between C. texana and C. pepo.

6. A composition according to claim 1 wherein the moisture content of said dried cucurbitacinous plant tissue is less than 10 percent by weight.

7. A composition according to claim 1 wherein the diabroticiticidal insecticide comprises less than 10 percent by weight of the total composition.

8. A composition according to claim 1 wherein the diabroticiticidal insecticide is selected from the group consisting of fenvalerate, permethrin, decamethrin, methomyl, carbaryl, carbofuran, trichlorofon, malethion, isofenfos and chlorpyrifos.

9. A composition according to claim 1 in the form of granules having a mesh size of 30 to 60.

10. A method for enhancing the diabroticiticidal effectiveness of an insecticide, said method comprising admixing said insecticide with dried cucurbitacinous Cucurbitale plant tissue.

11. A method for controlling a diabroticite beetle population, said method comprising applying to the soil an insecticide composition comprising the admixture of a major proportion by weight of dried cucurbitacinous plant tissue and a minor proportion by weight of a diabroticiticidal insecticide.