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WO1989000375A1 - Souches de trichoderma competentes dans la rhizosphere - Google Patents

Souches de trichoderma competentes dans la rhizosphere Download PDF

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Publication number
WO1989000375A1
WO1989000375A1 PCT/US1988/002011 US8802011W WO8900375A1 WO 1989000375 A1 WO1989000375 A1 WO 1989000375A1 US 8802011 W US8802011 W US 8802011W WO 8900375 A1 WO8900375 A1 WO 8900375A1
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WO
WIPO (PCT)
Prior art keywords
rhizosphere
trichoderma
atcc
group
spp
Prior art date
Application number
PCT/US1988/002011
Other languages
English (en)
Inventor
R. Ralph Baker
Sayed J. Ahmad
Original Assignee
Colorado State University Research Foundation
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Colorado State University Research Foundation filed Critical Colorado State University Research Foundation
Publication of WO1989000375A1 publication Critical patent/WO1989000375A1/fr

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Classifications

    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01CPLANTING; SOWING; FERTILISING
    • A01C1/00Apparatus, or methods of use thereof, for testing or treating seed, roots, or the like, prior to sowing or planting
    • A01C1/06Coating or dressing seed
    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01NPRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N63/00Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/30Microbial fungi; Substances produced thereby or obtained therefrom
    • A01N63/38Trichoderma
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/14Fungi; Culture media therefor
    • C12N1/145Fungal isolates
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12RINDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00Microorganisms ; Processes using microorganisms
    • C12R2001/645Fungi ; Processes using fungi
    • C12R2001/885Trichoderma

Definitions

  • This invention generally relates to new biotypes of soilborne biocontrol agents. More specifically it relates to rhizosphere-competent biocontrol agents.
  • rhizosphere competence has been employed to describe an attribute of rhizobia characterized by their consistent association with legume root nodules.
  • rhizosphere-competence is used to describe the ability of a microorganism to grow and function in the developing rhizosphere. In application, this means that the microorganism can be placed on the seed, it can grow and it can colonize the rhizosphere of the developing root.
  • Trichoderma spp. such as Trichoderma harzanium Rifai act as biological control agents against certain plant diseases. Nonetheless, use of Trichoderma spp., as biocontrol agents has been rather limited. This is mainly because seed treatment with Trichoderma spp. generally does not provide continued protection for the emerging root system of the maturing plant. Such seed treatment does serve to reduce preemergence damping-off, but the root system is left unprotected. It is generally believed that this failure to protect the root system is because Trichoderma spp.
  • Trichoderma species are Trichoderma polvsporua, ATCC number 20852 and Trichoderma viride, ATCC number 20853. They are particularly useful in controlling diseases caused by Phythium spp. , Sclerotium spp. , and Rhizoctonia solani in such varied plants as beans, maize, tomato, radish, cucumber, wheat, barley, lettuce and carnations. Applicants also have found that N- methyl-N-nitro-N-nitrosoguanidine was a particularly useful mutagenic chemical in producing Trichoderma species which possess the desired rhizosphere- competence characteristic. Other useful mutagenic agents would include, but not limited to ultraviolet light and x-ray radiation.
  • Trichoderma e.g., the Trichoderma polysporum and Trichoderma viride species disclosed herein
  • Trichoderma are particularly effective when they are mutated for both rhizosphere- competence and benomyl tolerance.
  • the herein disclosed strains are grown in a nutrient medium where cellulose represents a major part of the microorganisms source of carbon, or more preferably its only source of carbon. Representative sources of carbon would include, but not be limited to carboxy-methyl-cellulose, cotton linters, solka floe, and Sigma Cell 20 (Sigma Chemicals, St. Louis) .
  • Figure 1 is a bar graph depicting the dry weight of mycelium of strains of two Trichoderma spp. grown for 6 days in Czapek Dox broth with 37. Cellulose as the sole source of carbon.
  • Figures 2A and 2B depict the population densities of Trichoderma spp. in the rhizosphere soil of cucumber plants without benomyl added to the soil.
  • Figure 2 shows the result for T_j_ polysporum ATCC 20475 and ATCC 20853.
  • Figure 3 is a bar graph depicting the rhizosphere competence (RC) index of T_j_ polvsporum T.P. 18, T_i_ viride ATCC 20476, and Tj. viride T.V. 62.
  • RC rhizosphere competence
  • Rhizosphere Competence Various methods have been employed to test rhizosphere competence. These were primarily based on a comparison of the number of colony forming units ("cfu") of microorganisms in the soil associated with roots when compared to population densities in non-rhizosphere soil.
  • the rhizosphere competence assay used in this research effort was also directed toward improving measurement in time and space of the activity of potential rhizosphere inhabitants. Certain criteria were demanded by the experimental questions to be examined. Quantitative analysis of population densities at each depth of root was also necessary. No water was added during incubation obviating the possibility of propagules being washed into the rhizosphere.
  • the rhizosphere competence assay provides the best bioassay yet developed for the rhizosphere nitrous statues at root tips. It has potential for use in a wide variety of experimental problems related to ecological and nutritional interactions in the rhizosphere.
  • Trichoderma spp. disclosed herein were tested for rhizosphere competence by coating seeds with the respective isolates and following population densities of the fungus to a root depth of 8 cm. Both grew to a depth of 8 cm. Again Trichoderma spp. which are benomyl tolerant as well as rhizosphere competent are preferred embodiments of this invention.
  • Our particular rhizosphere- competence assays were conducted as follows. Polypropylene centrifuge tubes (28.6 by 103.6 mm) were sliced longitudinally into two halves. Each half was filled with moistened soil (-3 bars) and pre-incubated for 48 hours in plastic bags. One treated seed was placed on the half-tube 1 cm below the rim.
  • the unseeded half-tube was placed on the first half and secured with rubber bands. Tubes were completely randomized and lots in portions of six each were placed vertically in 10 cm diameter plastic pots. Soil, previously moistened to -0.03 bars and of the same pH as in the tubes, was added to the pots so that the length of the tube was surrounded by the soil, with the top 1 cm of each tube being uncovered. No water was added to the tubes or the pots after seed were sown. Pots were covered with plastic bags to maintain constant metric potential leaving enough space above the tubes for the plants to grow. Pots were placed under constant illumination supplied by 10 white, 40-watt, 120 cm long fluorescent lamps (approximately 5000 lux) , at desired temperatures.
  • MATERIALS Soil Nunn sandy loam was used in these investigations. Water content of 43.2 kg portions was adjusted to -0.03 bars and the soil was stored for 48 hours before use. The soils had the following characteristics: pH 7.0, conductivity 0.4 mmhos, lime low, organic matter 1.4%, No 3 -N 1 hg/g, P 9 hg/g. K 198 hg/g, Zn 0.5 hg/g, Fe 3.2 hg/g. Its pH was measured- by the CaCl 2 method.
  • Trichoderma polvsporum ATCC 20475 and Trichoderma viride ATCC 20476 were exposed to 100 mg/ml of N- methyl-N-nitro-N-nitrosoguanidine (Sigma Chemicals Co., St. Louis, Mo.) for 30 minutes. The conidia were centrifuged at 2500 g for 15 minutes and resuspended in sterile water three times. Seeds of cucumber (Cucumis sativus L. "straight Eight") , were surface disinfected for 10 minutes in 1.1% sodium hypochlorite solution and 5% ethanol, washed in distilled water, and air dried. Seeds were treated with conidial suspensions of each of the two Trichoderma spp. in water containing 2% (v/w) Pelgel (The Nitragen Co., Milwaukee, WI) as spreader or sticker. Conidial density was adjusted to 10 6 per seed. Controls were treated with Pelgel alone.
  • Trichoderma spp. in liquid culture. Strains of each of the two subject Trichoderma spp. were grown in 250 ml Erlenmeyer flasks containing 50 ml Czapek Dox broth on a rotary shaker at 100 rpm at 26°C for 6 days. Cellulose (Sigma Cell 20) was used as sole source of carbon. Each flask was seeded with a 4 mm diameter disk of PDA on which the strains had been grown for 2 days. After 6 days the hyphal mat was removed aseptically and dried for 2 days at 60°C to obtain the weight of mycelium. There were six replicates per strain. Rhizosphere competence (RC) index.
  • RC Rhizosphere competence
  • T ⁇ polysporum ATCC 20853
  • T_j_ viride ATCC 20476
  • ⁇ . polvsporum ATCC 20475
  • T ⁇ . viride ATCC 20476
  • the mutation of these species by the mutagen, mentioned above, and selection on Trichoderma-selective medium has resulted in T ⁇ polysporum (ATCC 20852) and ⁇ viride (ATCC 20853) tolerant to at least 10 mg/ l a.i. benomyl.

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  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Health & Medical Sciences (AREA)
  • Zoology (AREA)
  • Biotechnology (AREA)
  • Chemical & Material Sciences (AREA)
  • Wood Science & Technology (AREA)
  • General Health & Medical Sciences (AREA)
  • Microbiology (AREA)
  • Organic Chemistry (AREA)
  • Virology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Mycology (AREA)
  • Genetics & Genomics (AREA)
  • Environmental Sciences (AREA)
  • Botany (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Medicinal Chemistry (AREA)
  • Biomedical Technology (AREA)
  • Agronomy & Crop Science (AREA)
  • Pest Control & Pesticides (AREA)
  • Plant Pathology (AREA)
  • Dentistry (AREA)
  • Soil Sciences (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)

Abstract

Le Trichoderma polysporum (ATCC 20852) compétent dans la rhizosphère et le Trichoderma viride (ATCC 20853) compétent dans la rhizosphère sont décrits. Ce sont respectivement, des mutants du Trichoderma polysporum ATCC 20475 incompétent dans la rhizosphère et du Trichoderma viride ATCC 20476 incompétent dans la rhizosphère. Les souches compétentes dans la rhizosphère sont des agents biorégulateurs particulièrement efficaces pour combattre les maladies des plantes et/ou des arbres associées à divers champignons p.ex., Pythium spp., Sclerotium, spp., Rhizoctonia solani et des basidiomycètes tels que le Chondrostereum purpureum.
PCT/US1988/002011 1987-07-10 1988-06-13 Souches de trichoderma competentes dans la rhizosphere WO1989000375A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US7234787A 1987-07-10 1987-07-10
US072,347 1987-07-10

Publications (1)

Publication Number Publication Date
WO1989000375A1 true WO1989000375A1 (fr) 1989-01-26

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1990003732A1 (fr) * 1988-10-04 1990-04-19 Novo Nordisk A/S Preparations a action fongicide
WO1990013219A1 (fr) * 1989-05-05 1990-11-15 Allelix Crop Technologies Amelioration de la croissance de plants de coniferes
US8716001B2 (en) 2009-02-06 2014-05-06 Cornell University Trichoderma strains that induce resistance to plant diseases and/or increase plant growth

Non-Patent Citations (12)

* Cited by examiner, † Cited by third party
Title
PHYTOPATHOLOGY, Volume 69, issued May 1979, (Saint Paul, Minnesota, USA), LOCKE et al., "Biological control of Pythium seed rot of peas with Trichoderma Conidia", page 536. *
PHYTOPATHOLOGY, Volume 71, issued January 1981, (Saint Paul, Minnesota, USA), G. PAPAVIZAS, "Survival of Trichoderma Harzianum in soil and in pea and bean Rhizospheres", pages 121-125, see page 123 in particular. *
PHYTOPATHOLOGY, Volume 72, issued January 1982, (Saint Paul, Minnesota, USA), PAPAVIZAS et al., "Evaluation of new biotypes of Trichoderma Harzianum for tolerance to Benomyl and enhanced biocontrol capabilities", pages 126-132, see pages 130 and 131 in particular. *
PHYTOPATHOLOGY, Volume 75, issued November 1985, (Saint Paul, Minnesota, USA), AHMAD et al., "Induction of rhizosphere competence in Trichoderma Harzianum", page 1302. *
PHYTOPATHOLOGY, Volume 76, issued May 1986, (Saint Paul, Minnesota, USA), CHAO et al., "Colonization of the rhizosphere by biological control agents applied to seeds", pages 60-65, see pages 64 and 65 in particular. *
PHYTOPATHOLOGY, Volume 77, issued February 1987, (Saint Paul, Minnesota, USA), AHMAD et al., "Competitive saprophytic ability and cellulolytic activity of rhizosphere-competent mutants of Trichoderma harzianum", pages 358-362, see pages 361 and 362 in particular. *
PHYTOPATHOLOGY, Volume 77, issued February 1987, (Saint Paul, Minnesota, USA), AHMAD et al., "Rhizosphere competence of Trichoderma harzianum", pages 182-189, see pages 188 in particular. *
PLANT DISEASE, Volume 69, issued May 1985, (Saint Paul, Minnesota, USA), LIFSHITZ et al., "Decrease in incidence of Rhizoctonia preemergence damping-off by use of integrated chemical and biological controls", pages 431-434, see page 432 in particular. *
PLANT DISEASE, Volume 70, issued February 1986, (Saint Paul, Minnesota, USA), CHANG et al., "Increased growth of plants in the presence of the biological control agent Trichoderma Harzianum", pages 145-148, see pages 147 in particular. *
PROCEEDINGS OF THE INDIAN NATIONAL SCIENCE ACADEMY PART B, Volume 41, issued November 1975, (Bangladore, India), S. MALL, "Rhizoctonia diseases of legume crops as affected by Trichoderma Viride", pages 559-563, see page 561 in particular. *
R. COOK, "Ultrastructure of the root-soil interface", published 1983, by FOSTER et al., (Saint Paul, Minnesota, USA), page v. *
S. GARRETT, "Pathogenic root-infecting fungi", published 1970, by S. GARRETT, (Cambridge, England), page 115. *

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1990003732A1 (fr) * 1988-10-04 1990-04-19 Novo Nordisk A/S Preparations a action fongicide
WO1990013219A1 (fr) * 1989-05-05 1990-11-15 Allelix Crop Technologies Amelioration de la croissance de plants de coniferes
US8716001B2 (en) 2009-02-06 2014-05-06 Cornell University Trichoderma strains that induce resistance to plant diseases and/or increase plant growth
US8877481B2 (en) 2009-02-06 2014-11-04 Cornell University Trichoderma strains that induce resistance to plant diseases and/or increase plant growth
US8877480B2 (en) 2009-02-06 2014-11-04 Cornell University Trichoderma strains that induce resistance to plant diseases and/or increase plant growth

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