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Patent Issued for Biocontrol of Storage Maladies of Potatoes by Bacterial Antagonists Produced in Co-Culture

January 29, 2014

By a News Reporter-Staff News Editor at Journal of Engineering -- From Alexandria, Virginia, VerticalNews journalists report that a patent by the inventors Slininger, Patricia J. (Metamora, IL); Schisler, David A. (Morton, IL), filed on August 29, 2012, was published online on January 14, 2014.

The patent's assignee for patent number 8628764 is The United States of America as represented by the Secretary of Agriculture (Washington, DC).

News editors obtained the following quote from the background information supplied by the inventors: "The present invention relates to the biological control of fungal potato diseases and/or potato sprout inhibition. More particularly, this invention relates to compositions of bacteria which are effective antagonists against fungal species responsible for Fusarium dry rot and other potato diseases which occur in the field or in postharvest storage.

"The potato is the most important dicotyledonous source of human food, ranking as the fifth major food crop of the world. Fusarium-induced potato dry rot is an economically important problem of potatoes both in the field and in storage. Several species of the Fusaria induce this disease, however, Gibberella pulicaris (Fries) Sacc. (anamorph: Fusarium sambucinum Fuckel) is a major cause worldwide, especially in North America. Fusarium spp. can survive for years in field soil, but the primary inoculum is generally borne on seed tuber surfaces. The dry rot fungi infect potatoes via wounds in the periderm inflicted during harvesting or subsequent handling. In stored potatoes, dry rot develops most rapidly in high relative humidity (circa 70% and higher) and at C., but continues to advance at the coldest temperatures safe for potatoes. Although rots caused by Fusarium seldom reach epidemic proportions, the level of infected tubers in storage often reaches 60% or higher, with average losses estimated in the 10-20% range. In addition to destroying tissue, F. sambucinum can produce trichothecenes that have been implicated in mycotoxicoses of humans and animals.

"The high value of the potato crop and the significant economic losses caused by potato dry rot have led to investigations of various methods to control the disease. Success has been attained by use of the fungicides, thiabendazole and 2-aminobutane, which are applied to tubers at harvest or at preplanting [Carnegie et al. 1990. Ann App. Biol. 116: 61-72; Leach, 'Control of Postharvest Fusarium Tuber Dry Rot of White Potatoes,' pages 1-7 In ARS-NE-55, U.S. Dep. Agric., Washington, D.C.]. However, strong concerns are being raised about the potential adverse impact of these chemicals on ground and surface water reservoirs and on the health of agricultural product workers and consumers. Also, thiabendazole-resistant strains of F. sambucinum have emerged in populations from severely dry-rotted tubers in North America and in Europe. Potato breeding programs have given increased attention to development of cultivars with resistance to Fusarium but most of the reported cultivars produced by these programs are resistant to only one or two of several Fusarium strains [Leach et al. 1981. Phytopathology 71(6):623-629]

"One alternative to chemical fungicides in controlling potato rot is the use of biological agents. Postharvest biological control systems of fruit have been actively investigated since the 1980's. These include iturins as antifungal peptides in biological control of peach brown rot with Bacillus subtilis [Gueldner et al. 1988. Journal of Agricultural and Food Chemistry 36:366-370]; postharvest control of blue mold on apples using Pseudomonas spp. isolate L-22-64 or white yeast isolate F-43-31 [Janisiewicz, 1987. Phytopathology 77:481-485]; control of gray mold of apple by Cryptococcus laurentii [Roberts, 1990. Phytopathology 80:526-530]; biocontrol of blue mold and gray mold on apples using an antagonistic mixture of Pseudomonas spp. and Acremonium breve [Janisiewicz, 1988. Phytopathology 78:194-198]; control of gray mold and reduction in blue mold on apples and pears with an isolate of Pseudomonas capacia and pyrrolnitrin produced therefrom [Janisiewicz and Roitman, 1988. Phytopathology 78:1697-1700]; postharvest control of brown rot in peaches and other stone fruit with the B-3 strain of Bacillus subtilis [Pusey and Wilson. 1984, Plant Disease 68:753-756; Pusey et al. 1988. Plant Disease 72:622-626; and U.S. Pat. No. 4,764,371 to Pusey et al.]; antagonistic action of Trichoderma pseudokoningii against Botrytis cinerea Pers. which causes the dry eye rot disease of apple [Tronsmo and Raa. 1977. Phytopathol. Z. 89:216-220]; and postharvest control of brown rot and Alternaria rot in cherries by isolates of Bacillus subtilis and Enterobacter aerogenes [Utkhede and Sholberg. 1986. Canadian Journal of Microbiology 963-967]. A review of biological control of postharvest diseases of fruits and vegetables is given by Wisniewski et al. [1992. HortScience 27:94-98].

"More recently, significant progress has also been made in the isolation and development of bacteria agents for controlling diseases of potatoes. For instance, eighteen Gram-negative bacteria were originally discovered and developed as biocontrol agents to protect potatoes entering storage from Fusarium dry rot incited by Gibberella pulicaris [Schisler and Slininger. 1994. Selection and performance of bacterial strains for biologically controlling Fusarium dry rot of potatoes incited by Gibberella pulicaris. Plant Disease 8:251-255; Slininger et al. 1994 Two-dimensional liquid culture focusing: A method of selecting commercially promising microbial isolates with demonstrated biological control capability, in: M H Ryder, P M Stephens and G D Bowen (Eds.). Improving plant productivity with rhizosphere bacteria, 3rd international workshop on plant growth-promoting rhizobacteria, Adelaide, S. Australia, pp. 29-32. Glen Osmond, South Australia CSIRO Division of Soils; Slininger et al. 1996. Bacteria for the control of Fusarium dry rot of potatoes. U.S. Pat. No. 5,552,315; Schisler et al. 1997. Effects of antagonist cell concentration and two-strain mixtures on biological control of Fusarium dry rot of potatoes. Phytopathology 87:177-183; Schisler et al. 1998. Bacterial control of Fusarium dry rot of potatoes. U.S. Pat. No. 5,783,411], and these bacteria significantly reduced the level of dry rot disease in pilot trials (Schisler et al. 2000. Biological control of Fusarium dry rot of potato tubers under commercial storage conditions. American Journal of Potato Research 77:29-40). Top dry rot suppressive strains included Pseudomonas fluorescens biovar 5 (S11:P:12 NRRL B-21133 and P22:Y:05 NRRL B-21053, Pseudomonas fluorescens biovar 1 (S22:T:04 NRRL B-21102) and Enterobacter cloacae (S11:T:07 NRRL B-21050). All of these strains were also documented to suppress sprouting (Slininger et al. 2000. Biological control of sprouting in potatoes. U.S. Pat. No. 6,107,247; Slininger et al. 2003. Postharvest biological control of potato sprouting by Fusarium dry rot suppressive bacteria. Biocontrol Science and Technology 13:477-494), with Pseudomonas fluorescens S11:P:12 (NRRL B-21133) exhibiting the greatest efficacy, and E. cloacae S11:T:07 being the second most efficacious. In addition, the strains have been shown to suppress late blight incited by Phytophthora infestans US-8 mating type A2 in laboratory bioassays and small pilot simulations of commercial storage conditions with top performance shown by the following treatments: a mixture of four strains (comprised of S11:P:12+P22:Y:05+S22:T:04+ S11:T:07)>strain S22:T:04 used alone >strain S11:P:12 used alone [Slininger et al. 2007. Biological control of post-harvest late blight of potatoes. Biocontrol Science and Technology 17(5/6):647-663]. Most recently, we showed the ability of several of the dry rot antagonistic bacteria to suppress pink rot disease incited by Phytophthora erythroseptica, including S11:T:07 which exhibited the greatest efficacy and S22:T:04 exhibiting the third greatest efficacy (Schisler et al. 2007. Gram negative bacteria for reducing pink rot, dry rot, late blight, and sprouting potato tubers in storage. American Journal of Potato Research 84:115; Schisler et al. 2009. Bacterial antagonists, zoospore inoculums retention time and potato cultivar influence pink rot disease development. American Journal of Potato Research 86:102-11).

"Several researchers have reported that mixtures of other microbial strains can enhance and/or improve the consistency of biological control (Pierson and Weller 1994. Use of mixtures of fluorescent pseudomonads to suppress take-all and improve the growth of wheat. Phytopathology 84:940-947; Duffy and Weller. 1995. Use of Gaeumannomyces graminis var. graminis alone and in combination with fluorescent Pseudomonas spp. to suppress take-all of wheat. Plant Disease 79:907-911; Duffy et al. 1996. Combination of Trichoderma koningii with fluorescent pseudomonads for control of take-all on wheat. Phytopathology 86:188-194; Janisiewicz. 1996. Ecological diversity, niche overlap, and coexistence of antagonists used in developing mixtures for biocontrol of post-harvest diseases of apples. Phytopathology 86:473-479; Leeman et al. 1996. Suppression of Fusarium wilt of radish by co-inoculation of fluorescent Pseudomonas spp. and root-colonizing fungi. European Journal of Plant Pathology 102:21-31; de Boer et al. 1999. Combining fluorescent Pseudomonas spp. strains to enhance suppression of Fusarium wilt of radish. European Journal of Plant Pathology 105:201-210; Guetsky et al. 2001. Combining biocontrol agents to reduce the variability of biological control. Phytopathology 91:621-27; Krauss and Soberanis. 2001. Biocontrol of cocoa pod diseases with mycoparasite mixtures, Biological Control 22: 149-158; Hwang and Benson. 2002. Biocontrol of Rhizoctonia stem and root rot of poinsettia with Burkholderia cepacia and binucleate Rhizoctonia. Plant Disease 86:47-53; Cruz et. al. 2006. Exploiting the genetic diversity of Beauveria bassiana for improving the biological control of the coffee berry borer through the use of strain mixtures, Applied Microbiology and Biotechnology 71: 916-92. Preliminary research has shown that formulations containing multiple strains of the above-mentioned dry rot antagonists performed more consistently than individual strains did when subjected to thirty-two storage environments varying in potato cultivar, harvest year, potato washing procedure (microflora exposure), temperature, and storage time (Slininger et al. 2001. Combinations of dry rot antagonistic bacteria enhance biological control consistency in stored potatoes. Phytopathology 91:S83) However, despite these advances, the need remains for improved biological control agents for inhibiting fungal diseases of potatoes."

As a supplement to the background information on this patent, VerticalNews correspondents also obtained the inventors' summary information for this patent: "We have now discovered novel compositions of bacteria which are effective for inhibiting (reducing the incidence or severity of) fungal diseases of potatoes, and particularly fungal diseases of potatoes under storage conditions. The bacteria which comprise the compositions of this invention, specifically two or more of Pseudomonas fluorescens (NRRL B-21.13), Pseudomonas fluorescens biovar (NRRL B-21053), Pseudomonas fluorescens (NRRL B-21102) and Enterobacter cloacae (NRRL B-21050), have been previously described for the control of fungi-induced diseases of potatoes. However, we have unexpectedly discovered that when two or more of these bacteria are cultured together in the same culture medium, the resultant composition or co-culture is significantly more effective for inhibiting fungi-induced diseases of potatoes than blends of the same bacteria which have been cultured separately. In addition to their efficacy as biological control agents against fungi-induced diseases, the compositions also exhibit significantly greater control of sprouting in stored potatoes. The method of producing the bacterial antagonistic compositions comprises: a) inoculating a culture medium with at least two bacteria selected from the group of Pseudomonas fluorescens (NRRL B-21133), Pseudomonas fluorescens (NRRL B-21053), Pseudomonas fluorescens (NRRL B-21102) and Enterobacter cloacae (NRRL B-21050), and incubating the inoculated medium containing said bacteria under conditions effective for their growth, and for a period of time effective to produce a co-culture thereof; and (b) recovering the co-culture, a composition of bacteria effective for inhibiting fungal diseases of potatoes.

"In accordance with this discovery, it is an object of this invention to provide novel compositions of bacteria which are superior antagonists of fungi responsible for fungi-induced potato diseases which occur in the field or in postharvest storage.

"Another object of this invention is to provide novel compositions of bacteria which are superior antagonists of fungi responsible for Fusarium dry rot and other potato diseases which occur during postharvest storage.

"Yet another object of this invention is to provide novel compositions of bacteria which exhibit significantly increased efficacy for inhibiting sprouting of potatoes during storage.

"A further object of this invention is to provide novel method for producing compositions of bacteria which exhibit significantly increased efficacy for inhibiting fungi induced diseases of potatoes and inhibiting sprouting of stored potatoes.

"These and other objects of the invention will become readily apparent from the ensuing description."

For additional information on this patent, see: Slininger, Patricia J.; Schisler, David A.. Biocontrol of Storage Maladies of Potatoes by Bacterial Antagonists Produced in Co-Culture. U.S. Patent Number 8628764, filed August 29, 2012, and published online on January 14, 2014. Patent URL:

Keywords for this news article include: Bacillaceae, Agricultural, Plant Diseases, Bacillus subtilis, Enterobacteriaceae, Gram-Positive Rods, Enterobacter cloacae, Gram-Negative Bacteria, Gram-Positive Bacteria, Science And Technology, Endospore-Forming Bacteria, Plant Physiological Phenomena, Gram-Positive Endospore-Forming Rods.

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Source: Journal of Engineering

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