Bacillus thuringiensis isolates for controlling acarides

Disclosed are Bacillus thuringiensis isolates designated B.t. PS45B1, B.t. PS24J, B.t. PS94R3 B.t. PS17, B.t. PS62B1 and B.t. PS74G1 which Make novel .delta.

-endotoxins active against acarid pests. Thus, these isolates, or mutants thereof, can be used to control such pests. Claimed are genes encoding these novel .delta. -endotoxins, that can be taken out from such isolates and moved to other host germs , or plants. Expression of those toxins in microbe hosts results in the control of acarid pests, whereas altered plants become immune to acarid pests.


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The soil microbe Bacillus thuringiensis (B.t.) is a Gram-positive, spore-forming bacterium characterized by parasporal crystalline protein inclusions. These inclusions frequently appear microscopically as distinctively shaped crystals. The B.t.microbes produce many different toms. A mouse-lethal ct-exotoxin is produced by specific isolates of this species, as well as two kinds of hemolysins, a nucleotide .beta. -exotoxin and .delta. -endotoxins. Delta-endotoxins are derived from the parasporalcrystalline protein inclusions and are toxic to pests and specific in their poisonous action. The B.t. toxins have been commercially used for their use in pest management. Particular B.t. endotoxin genes have been isolated and sequenced, andrecombinant DNA-based B.t. goods have been produced and accepted for use. In addition, by means of genetic engineering techniques, new methods of delivering B.t. endotoxins to agricultural environments are under development, including theuse of plants genetically engineered using endotoxin genes for insect resistance and using stabilized intact cancerous cells as B.t. endotoxin delivery vehicles (Gaertner, F.H., L. Kim [1988]TIBTECH 6:S4-S7).

Until the past ten decades, commercial use of B.t. pesticides has been largely restricted to a narrow range of lepidopteran (caterpillar) pests. Preparations of the spores and crystals of B. thudngiensis subsp. Kurstaki have been used for manyyears as commercial insecticides for lepidopteran pests. For example, B. thuringiensis var. Kurstaki HD-1 creates a delta-endotoxin which is toxic to the larvae of a range of lepidopteran pests.

In recent decades, however, investigators have found B.t. pesticides with specificities for a much wider array of pests. For instance, other species of B.t., specifically israelensis and san diego (a.k.a.B.t. tenebdonis, a.k.a. M-7), have beenused commercially to control pests of the orders Diptera and Coleoptera, respectively (Gaertner, F.H. [1989]”Cellular Delivery Systems for Insecticidal Proteins: Living and Non-Living Microorganisms,” in Controlled Delivery of Crop Protection Agents,R.M. Wilkins, ed., Taylor and Francis, New York and London, 1990, pp. 245-255). See also Couch, T.L. (1980) “Mosquito Pathogenicity of Bacillus thudngiensis var. israelensis,” Developments in Industrial Microbiology 22:61-76; Beegle, C.C., (1978)”Use of Entomogenous Bacteria in Agroecosystems,” Developments in Industrial Microbiology 20:97-104. Krieg, A., A.M. Huger, G.A. Langenbruch, W. Sctmetter (1983) Z. ang. Ent. 96:500-508, clarify a B.t. isolate called Bacillus thuringiensis var.tenebrionis, which is reportedly active against two beetles from the order Coleoptera. These will be the Colorado potato beetle, Leptinotarsa decemlineata, and Agelastica alni.

There have been reports concerning the use of Bacillus thudngiensis preparations for its control of acarid insects or pests. These books are as follow:


Royalty, R.N., F.R. HaH, R.A.J. Taylor (1990)”Effects of thuringiensin on Tetranychus urticae (Acari: Tetranychidae) mortality, fecundity, and Ingesting,” J. Econ.

Entomot 83:792-798.


Neal, J.W., R.K. Lindquist, K.M. Gott, M.L. Casey (1987)”Action of the themostable beta-exotoxin of Bacillus thudngiensis Berliner on Tetranychus urticae and Tetranychus cinnabarinus,”J. Agric.

Entomol. 4:33-40.


Vlayen, P., G. Impe, R. Van Semaille (1978)”Effect of a commercial preparation of Bacillus thuringiensis about the spider mite Tetranychus urticae Koch.

(Acari: Tetranychidae),” Mededelingen 43:471-479.

In the aforementioned published studies, the active ingredient from the B.t. trainings was .beta. -exotoxin (also referred to as thuringiensin).

The major focus for commercial usage of B.t. radicals is about the .delta. -endotoxins in the parasporal crystalline protein inclusions. Recently, new subspecies of all B.t. have been identified, and enzymes responsible for busy .delta. -endotoxinproteins have been isolated (Hofte, H., H.R. Whiteley [1989]Microbiological Reviews 52(2):242-255). Hofte and Whiteley categorized B.t. crystal protein genes to 4 main classes. The courses have been CryI (Lepidoptera-specific), CrylI (Lepidoptera- andDiptera-specific), CrylII (Coleoptera-specific), and CryIV (Diptera-specific). The discovery of breeds specifically hazardous to other pests was reported. (Feitelson, J.S., J. Payne, L. Kim [1992]Bio/Technology 10:271-275).

The cloning and expression of a B.t. crystal protein gene in Escherichia coli has been described in the published literature (Schnepf, H.E., H.R. Whitely [1981]Proc. Natl. Acad. Sci. USA 78:2893-2897). U.S. Pat. No. 4,448,885 and U.S.Pat. No. 4,467,036 both reveal the expression of B.t. crystal proteins in E. coli. U.S. Pat. Nos. 4,797,276 and 4,853,331 reveal B.thuringiensis strain san diego (a.k.a.B.t. tenebrionis, a.k.a. M-7) which may be employed to control coleopteranpests in various environments. U.S. Pat. No. 4,849,217 discloses isolates of B.t. which have activity against a coleopteran pests, the alfalfa weevil, while U.S. Pat. No. 4,948,734 discloses particular temptations of B.t. which have activity againstnematodes. U.S. Pat. No. 5,093,120 discloses using these B.t. isolates to control nematode diseases in plants and animals. A number of other patents have issued for new B.t. isolates and new uses of B.t. isolates. The discovery of new B.t.isolates and new uses of known B.t. isolates remains an empirical, unpredictable art.

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