Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Peptide containing doai
Reexamination Certificate
1999-05-14
2001-09-18
Tate, Christopher R. (Department: 1651)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Peptide containing doai
C514S002600, C514S011400, C424S093462, C424S405000
Reexamination Certificate
active
06291426
ABSTRACT:
FIELD OF THE INVENTION
The present invention is in the field of biopesticides. More particularly, this invention relates to the finding that a novel strain of
Bacillus subtilis
, AQ713, can inhibit a broad range of fungal and bacterial plant diseases and also have activity against corn rootworm. The invention also relates to fungicidal, bactericidal, and insecticidal compositions comprising this novel Bacillus strain and the antibiotics and metabolites produced by this strain either alone, or in combination with other chemical and biological pesticides.
BACKGROUND OF THE INVENTION
For a number of years, it has been known that various microorganisms exhibit biological activity so as to be useful to control plant diseases. Although progress has been made in the field of identifying and developing biological pesticides for controlling various plant diseases of agronomic and horticultural importance, most of the pesticides in use are still synthetic compounds. Many of these chemical fungicides are classified as carcinogens by the EPA, are toxic to wildlife and other non-target species. In addition, pathogens may develop resistance to chemical pesticides (see, e.g., Schwinn et al., p. 244, ADVANCES IN PLANT PATHOLOGY:
PHYTOPHTHORA INFESTANS
, THE CAUSE OF LATE BLIGHT OF POTATO (Academic Press, San Diego 1991).
Every year 250-300 million dollars of chemical pesticides are used to control corn rootworm infestations. Many of these chemical pesticides are toxic to humans, wildlife and other nontarget species. Also some have been found in the ground water. New chemical insecticides cost $100 million to develop.
Biological control offers an attractive alternative to synthetic chemical fungicides. Biopesticides (living organisms and the naturally produced compounds produced by these organisms) can be safer, more biodegradable, and less expensive to develop.
Screening programs have identified certain Bacillus spp. (Bacillus spp. includes
B. subtilis, B. cereus, B. mycoides, B. thuringiensis
) strains that exhibit antifungal activity. (See e.g Stabb et al. (1990)
Applied Environ. Microbiol
. 60: 4404-4412). These strains have been shown to produce zwittermicin-A and or kanosamine (Milner et al. (1996)
Appl. Environ. Microb
. 62: 3061-3066), two antibiotic agents that are effective against the soil borne disease damping off, caused by
Phytophthora medicaginis, P. nicotianae, P. aphanidermatum
or
Sclerotinia minor
(See Stabb et al., supra). Zwittermicin-A is a water soluble, acid stable linear aminopolyol molecule (see, He et al, (1994)
Tetra. Lett
. 35 (16) 2499-2502.
U.S. Pat. No. 5,049,379 to Handelsman et al. describes how zwittermicin-A produces damping off in alfalfa and soybeans. When the seed was coated with
B. cereus
ATCC 53522, the pathogenic activity of root rot fungus is inhibited. Similarly application of spore-based formulations of certain
B. cereus
strains to soybean seeds or the soil surrounding the seeds has been shown to improve soybean yield at field sites. (See, Osburne et al (1995)
Am. Phytopathol. Soc
. 79(6): 551-556). Methods of applying biopesticides are well known in the art and include, for example, wettable powders, dry flowables, microencapsulation of effective agents, liquid or solid formulations of antibiotic fractions from suitable cultures. (See e.g., U.S. Pat. No. 5,061,495 to Rossall or U.S. Pat. No. 5,049,379 to Handelsman).
Smith et al. (1993)
Plant Disease
77(2) 139-142 report that the activity of the soil-borne fungus,
Pythium aphanidermatum
, that causes cottony cucumber leak can be suppressed using zwittermicin-producing
B. cereus
strain UW85. Leifert et al. (1995)
J. Appl. Bacteriol
. 78: 97-108 report that the production of anti-Botrytis and anti-Alternaria antibiotics by two Bacillus strains,
B. subtilis
CL27 and
B. pumilis
CL 45. The whole broth and cell-free filtrates were active against Botrytis and Alternaria in in vitro tests and were active against Botrytis in in vivo small plant tests on Astilbe. Leifert et al. (1997) U.S. Pat. No. 5,597,565 disclose
B. subtilis, B. pumilis
, and
B. polymyxa
that are particularly effective at inhibiting post harvest disease causing fungi. They also disclose the presence of antibiotics produced in the cell-free culture filtrate and their activity at different pH values, but they do not identify these compounds.
Rossall (1994) U.S. Pat. No. 5,344,647 discloses
Bacillus subtilis
strains with broad anti-fungal activity. Sholberg et al. (1995)
Can. J. Microbiol
. 41: 247-252, Swinburne et al. (1975)
Trans. Brit. Mycol. Soc
. 65: 211-217, Singh and Deverall (1984)
Trans. Br. Mycol. Soc
. 83: 487-490, and Ferreira, et al. (1991)
Phytopathology
81: 283-287. Baker et al. (1983)
Phytopathology
73: 1148-1152 disclose the use of Bacillus spp. and
Bacillus subtilis
as biocontrol agents of fungal plant pathogens. Baker et al. (1983)
Phytopathology
73: 1148-1152 also report on an antifungal
Bacillus subtilis
for use on plant pathogens. Pusey et al. (1988)
Plant Dis
. 72: 622-626, Pusey and Robins (U.S. Pat. No. 5,047,239), and McKeen et al. (1986)
Phytopathology
76: 136-139 disclose control of post harvest fruit rot using
B. subtilis
. McKeen et al, supra, have shown that antibiotics similar to the low molecular weight iturin cyclic polypeptides contribute to this fungicidal activity of
B. subtilis.
Liu et al. (1995) U.S. Pat. No. 5,403,583 disclose a
Bacillus megaterium
, ATCC 55000 and a method to control the fungal plant pathogen,
Rhizoctonia solani
. Islam and Nandi (1985)
Journal of Plant Diseases and Protection
92(3): 241-246 disclose a
Bacillus megaterium
with antagonism to
Drechslera oryzae
, the causal agent of rice brown spot. The same authors, Islam and Nandi (1985)
Journal of Plant Diseases and Protection
92(3) 233-240 also disclose in-vitro antagonism of
B. megaterium
against
Drechslera oryzae, Alternaria alternata
and
Fusarium roseum
. They discuss three components in the culture filtrate. The most active antibiotic was highly soluble in water and methanol with a UV peak at 255 nm and a shoulder at 260 nm, which proved to be a polyoxin-like lipopeptide. Cook ((1987)
Proceedings Beltwide Cotton Production—Mechanization Research Conference
, Cotton Council, Memphis, p. 43-45) discloses the use of a suspension of
Bacillus megaterium
to reduce the number of cotton plants killed by
Phymatotrichum omnivorum
, a cause of cotton root rot.
Antibiotic production of
B. megaterium
has been recorded by Berdy (CRC Handbook of Antibiotic Compounds, Vols. I-XIV, (CRC Press, Inc. Boca Raton, Fla. 1980-87) who reports production of low-mammalian toxic peptide antibiotics such as ansamitocin-PDM-O, bacimethrin, megacin, pentapeptide, homopeptides.
Bacilli are known to produce antifungal and antibacterial secondary metabolites (Korzybski et al. (1978)). University of Wisconsin and Cornell researchers have identified a novel fungicidal compound, zwittermicin A, produced by Bacillus sp. (He et al. (1994)
Tetra. Lett
. 35(16):2499-2502). A second fungicidal metabolite produced by the same strain was recently identified as the known amino-sugar, kanosamine (Milner et al. (1996)
Appl. Environ. Microb
. 62:3061-3065).
Another group of previously described Bacillus metabolites are the cyclic lipopeptides of the iturin class, some of which are potent fingicidal agents. These agents consist of a cyclic octapeptide with seven &agr;-amino acids and one &bgr;-amino acid with an aliphatic side chain. There are several groups of iturins that differ in order and content of the amino acid sequence. These are shown in Table 1 below. Generally, a suite of related molecules is produced with differences in the length and branching of the aliphatic amino acid residue. When tested against
Saccharomyces cerevesiae
, mycosubtilin was found to be the most active agent (LC50=10 &mgr;g/mL) followed by iturin-A and bacillomycin L (both haviing an LC50=30 &mgr;g/mL) (Beeson et al. (1979)
J. Antibiotics
32(8):828-833). The mode of action of these cyclic lipopeptides has b
Heins Sherry Darlene
Jimenez Desmond Rito
Manker Denise Carol
Marrone Pamela Gail
McCoy Randy Jay
Agraquest, Inc.
Baker & McKenzie
Konski Antoinette F.
Tate Christopher R.
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