Bacillus thuringiensis containing compositions

Details Bacillus thuringiensis containing compositions Bacillus thuringiensis containing compositions

1998-04-16

2001-07-24

Marx, Irene (Department: 1651)

Drug, bio-affecting and body treating compositions

Whole live micro-organism, cell, or virus containing

Bacteria or actinomycetales

C435S252500, C435S252100

Reexamination Certificate

active

06264942

ABSTRACT:

TECHNICAL FIELD
The present invention relates to organisms producing biological pesticides. More particularly, it pertains to novel strains of the bacterium
Bacillus thuringiensis
which are effective against certain insect species, as well as to methods for the preparation and use thereof.
BACKGROUND OF THE INVENTION
Insecticides have enjoyed widespread use in commercial agriculture, and have enabled an enormous increase in crop yields and product quality. Pesticides are also routinely used to control various insects, as for example flies or mosquitoes, when pest populations pose a nuisance or health hazard to humans or livestock. There is, however, an increasing awareness of environmental risks associated with the use of certain synthetic pesticides, including concern over the bioaccumulation of pesticides in the food chain or their detrimental effects on non-target organisms. Biological pesticides and especially natural biopesticides have therefore been of considerable interest to those seeking environmentally acceptable means of pest control.
The microorganism
B. thuringiensis
has long been recognized to be useful in the control of insect pests. The sporulating
B. thuringiensis
cell produces a class of compounds, formerly regarded as a single &dgr;-endotoxin but now understood to comprise several distinct toxin proteins, which are concentrated in a crystalline protein inclusion body found in the endospore. Upon ingestion of the inclusion body by a susceptible insect larva and proteolysis in the insect gut, the endotoxin proteins are converted into active compounds which destroy the gut epithelium and ultimately the pest itself.
B. thuringiensis
&dgr;-endotoxins have accordingly been found to be useful as pesticides when applied in the form of lysates or other fermentation extracts of cultures of the microorganism. These toxins show remarkable activity against a variety of Lepidoptera species and other insects. However,
B. thuringiensis
preparations have proved to be of only limited value in combating insects such as those of the genera Spodoptera and Plutella, as well as various other lepidopteran pests. This toxicity of
B. thuringiensis
preparations against only certain pest species, or differential toxicity, is believed to be due to the expression of only certain endotoxin genes in any given
B. thuringiensis
variant, each toxin contributing in an unpredictable manner to the overall toxicity profile.
Numerous researchers have attempted to identify
B. thuringiensis
strains which have a broader or different spectrum of pesticidal activity, or to manipulate the
B. thuringiensis
genome to promote the expression of particular &dgr;-endotoxins. Efforts directed to screening individual isolates for toxicity have led to the isolation of some previously unknown strains such as
B. thuringiensis
var.
tenebrionis
, dislosed by Krieg et al. in U.S. Pat. No. 4,766,203, issued Aug. 23, 1988, which strain is reportedly effective for combating Coleoptera. The use of conventional screening procedures to identify strains with new pesticidal activity, however, is time- and labor-intensive given the innumerable variants of
B. thuringiensis
that occur in nature.
Another approach has been to clone genes coding for
B. thuringiensis
&dgr;-endotoxins and to re-arrange the
B. thuringiensis
genome in a beneficial way, or to introduce the cloned genes into new microbial hosts for expression. Herrnstadt et at., in U.S. Pat. No. 4,771,131, issued Sep. 13, 1988, describe the cloning of an M-7 toxin gene said to be suitable for expression in other microbes such as Pseudomonas and to thereby confer the ability to control beetles of the order Coleoptera. These methods, however, suffer from the drawback that the resulting organisms are subjected to increased regulatory scrutiny relative to organisms which occur naturally.
There is, therefore, a continued need for the identification of
B. thuringiensis
strains which display a broader or different spectrum of pesticidal activity. Ideally, such strains would be identified from among naturally occurring variants without the use of random screening methods.
SUMMARY OF THE INVENTION
Accordingly, the present invention comprises novel bacterial isolates of
B. thuringiensis
which have enhanced toxicity with respect to previously resistant or insufficiently susceptible insect species. These isolates can be employed against several insect species resistant to treatment with
B. thuringiensis
including, but not limited to,
Plutella xylostella
(diamondback moth),
Spodoptera frugiperda
(fall armyworm) and
Spodoptera exigua
(beet armyworm), as well as certain secondary pests such as
Trichoplusia ni
(cabbage looper). The bacterial isolates of the invention may be characterized by their possession of a particular subset of the genes coding for the various
B. thuringiensis
endotoxin proteins and by a characteristic plasmid profile or array known to be associated therewith.
The present invention also comprises a method for the efficient identification of such bacterial isolates, in which a generalized nucleotide probe may be used to establish, in a strain already known to have some toxicity to a particular insect pest, the identity of that strain's endotoxin genes. Next, gene-specific DNA probes are prepared for the detection of those endotoxin genes; these probes are then used to pre-screen
B. thuringiensis
strains to identify a set of variants which are capable of synthesizing the corresponding endotoxins. The variants thus selected may ultimately be subjected to further screening of a more conventional nature to obtain particular variants which display even higher levels of pesticidal activity.
Further comprised by the present invention are cloned or synthesized nucleotide sequences which are useful in the preparation of the gene-specific DNA probes of the invention. These sequences are determined by sequence analysis of the target genes of interest and in accordance with the degree of specificity desired. Where a generalized probe (i.e., one capable of recognizing multiple endotoxin genes) is required, a nucleotide sequence may be constructed which is based on a conserved region of the
B. thuringiensis
toxin genes. Alternatively, sequences which are unique to the various toxin genes may be used to prepare highly specific probes.
The present invention additionally comprises the use of bacterial isolates of the invention in the control or eradication of insect pests, as well as compositions containing an insecticidally effective amount of such an isolate in combination with an acceptable carrier.


REFERENCES:
patent: 4206281 (1980-06-01), Goldberg
patent: 4652628 (1987-03-01), Walfield et al.
patent: 4695455 (1987-09-01), Barnes
patent: 4766203 (1988-08-01), Krieg et al.
patent: 4771131 (1988-09-01), Hernstadt et al.
patent: 4853331 (1989-08-01), Hernstadt et al.
patent: 4902507 (1990-02-01), Morris
patent: 4935353 (1990-06-01), Burges
patent: 5039523 (1991-08-01), Payne et al.
patent: 5045469 (1991-09-01), Payne et al.
patent: 5063055 (1991-11-01), Burges
patent: 5080897 (1992-01-01), Gonzalez
patent: 5366892 (1994-11-01), Foncerrada et al.
patent: 5407825 (1995-04-01), Payne et al.
patent: 5424409 (1995-06-01), Ely et al.
patent: 5426049 (1995-06-01), Sick et al.
patent: 5427786 (1995-06-01), Payne et al.
patent: 5430137 (1995-07-01), Gaertner et al.
patent: 0256553 (1988-02-01), None
patent: 0401979 (1990-12-01), None
Shimizu et al.,Agric. Biol. Chem. vol. 52, pp. 1565-1573, 1988.*
Haider, M.Z., et al., “Cloning and heterologous expression of an insecticidal delta-endotoxin gene fromBacillus thuringiensisvar. aizawai ICI toxic to both lepidoptera and deptera”,Gene, 52:285-290 (1987).
Prefontaine, G., et al., “Use of Oligonucleotide Probes to Study and Relatedness of Delta-Endotoxin Genes amongBacillus thuringiensisSubspecies and Strains”,Applied and Environmental Microbiology, 53(12):2808-2814 (1987).
Klier, A., et al., “Cloning and Expression inEscherichia coliof the Crystal Protein Gene fromBacillus thuringiensisStrain

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