WO2023219483A1 - Probiotic biofungicidal mixture and method, for controlling mycotoxigenic fungi - Google Patents

Probiotic biofungicidal mixture and method, for controlling mycotoxigenic fungi Download PDF

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WO2023219483A1
WO2023219483A1 PCT/MX2023/050016 MX2023050016W WO2023219483A1 WO 2023219483 A1 WO2023219483 A1 WO 2023219483A1 MX 2023050016 W MX2023050016 W MX 2023050016W WO 2023219483 A1 WO2023219483 A1 WO 2023219483A1
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mixture
spp
isolated strain
biofungicide
fungi
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PCT/MX2023/050016
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Spanish (es)
French (fr)
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Luis Rodrigo BECERRA CARRANZA
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Becerra Carranza Luis Rodrigo
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    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION 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
    • A01N63/00—Biocides, 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
    • A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
    • A01N63/22—Bacillus
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION 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
    • A01N63/00—Biocides, 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
    • A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
    • A01N63/22—Bacillus
    • A01N63/23—B. thuringiensis
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION 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
    • A01N63/00—Biocides, 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
    • A01N63/30—Microbial fungi; Substances produced thereby or obtained therefrom
    • A01N63/38—Trichoderma

Definitions

  • Mycotoxins are toxic compounds produced naturally by some types of fungi and/or molds; which grow in numerous foods, such as cereals, dried fruits, nuts and spices. Its growth can take place before or after harvest, during storage or in the food itself in warm and humid environments. Most mycotoxins are chemically stable and persist after food processing. Several hundred mycotoxins have been identified, but the most frequent ones that pose a problem for human and livestock health are aflatoxins, ochratoxin A, patulin, fumonisins, zearalenone and nivalenol and deoxynivalenol. Mycotoxins appear in the food chain as a result of the infection of crops by fungi and/or molds, either before or after harvest. Exposure to mycotoxins can occur directly by eating infected foods, or indirectly, from animals. fed contaminated food, and particularly from milk.
  • Patent document EP2011713789A refers to combinations of active compounds, in particular within a fungicidal composition, comprising (A) a dithiyne-tetracarboximide of formula (I) and at least one agriculturally beneficial biological control agent (B); It also refers to a method for curatively or preventively controlling phytopathogenic fungi of plants or crops, by applying a combination according to the invention for the treatment of seeds. It also refers to compositions and methods for reducing general damage to plants and plant parts, as well as losses in harvested fruits or vegetables caused by plant pathogenic fungi or other unwanted microorganisms. In particular, it relates to compositions and methods for protecting fruits, vegetables, potatoes and grapes during the growth phase, around harvest and after harvest.
  • Patent document EP2011156571A describes a method for controlling phytopathogenic diseases in useful plants or in their propagation material, which comprises applying a combination of the components to the useful plants, their place or their propagation material.
  • A) and B) in a synergistically effective amount wherein component A) is a compound of formula (I), wherein R1 is di fluoromethyl or tri fluoromethyl; Y is -CHR2- or formula (DD) and R2 is hydrogen or C1-C6 alkyl, or a tautomer of such a compound; and component B) is a compound selected from compounds known for their fungicidal and/or insecticidal activity, among which some strains of microorganisms are mentioned, it is particularly effective for controlling or preventing fungal diseases of useful plants.
  • component A) is a compound of formula (I), wherein R1 is di fluoromethyl or tri fluoromethyl
  • Y is -CHR2- or formula (DD) and R2 is hydrogen or C1-C6 al
  • patent document CA2928860 discloses a biopesticide comprising an agriculturally suitable carrier, an effective amount of a fungal pesticide, at least one sorbitan fatty acid surfactant, and at least one sorbitol ethoxylate ester surfactant.
  • fungicides Alternaria cassiae, Fusarium lateritum, Fusarium solani, Verticillium lecanii, Aspergillus parasiticus, Metarhizium anisopliae or Beauveria bessiana are mentioned.
  • a probiotic mixture a biofungicide and a method have been developed for the control of mycotoxigenic fungi in plant tissues used in the preparation of food mixtures.
  • plant tissues can preferably be grains and seeds.
  • Figure 1 is a graph that illustrates the behavior of mycotoxin control in corn grains, during 3 and 6 months.
  • a further embodiment of the mixture according to the present invention is when the isolated strain of Trichoderma harzianum is the ARS TH-19 strain deposited with the deposit number CM-CNRG TB174.
  • the isolated strain of Trichoderma harzianum is the ARS TH-19 strain deposited with the deposit number CM-CNRG TB174.
  • any other isolated strain of Trichoderma harzianum could be useful.
  • the isolated strain of Bacillus subtilis is the ARS BS-19 strain deposited with the deposit number CM-CNRG TB175. It is also possible that any other isolated strain of Bacillus subtilis could be useful.
  • Another preferred embodiment of said mixture is when the isolated strains are in a 1:2:2 ratio.
  • a second object of this invention is a biofungicide for the control of mycotoxigenic fungi, said biofungicide comprising a probiotic mixture in accordance with the present invention.
  • a third object of the present invention is a method for the control of mycotoxigenic fungi, which comprises applying an effective amount of the biofungicide proposed by the present invention to plant tissues infected with mycotoxigenic fungi; preferably plant tissues useful in the preparation of food mixtures; Therefore, the food mixture can be forage for cattle, goats, pigs, horses, etc.
  • An embodiment of the method according to the present invention is when the effective amount is 0.3 mL per 1 Kg of plant tissue.
  • Example 1 Evaluation of the control effect of the biofungicide containing the probiotic mixture in the control of mycotoxigenic fungi in corn (Zea mays L.)
  • Forage corn grains (Zea mays L.) were used, to which the amount of mycotoxins was quantified: Aflatoxin, Ochratoxin, DON, T2 and Fumonisins present in the grains; and then inoculate them with the phytopathogen Fusarium spp. and Aspergillus spp., to ensure the presence of mycotoxins.
  • the inoculated corn grains were separated into two parts, where one part was treated with the biofungicide that includes the probiotic mixture that in turn contains: the RVP BTI-20 (Bacillus thuringiensis var israelensis) r ARS TH-19 (Metarhizium) strains. anisopliae) and ARS BS-19 (Bacillus subtilis). Two measurements of mycotoxins were carried out, at 3 and 6 months.
  • RVP BTI-20 Bacillus thuringiensis var israelensis
  • ARS TH-19 Trichoderma harzianum
  • ARS strains BS-19 Bacillus subtilis

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  • Life Sciences & Earth Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Zoology (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Microbiology (AREA)
  • Pest Control & Pesticides (AREA)
  • Biotechnology (AREA)
  • Agronomy & Crop Science (AREA)
  • Plant Pathology (AREA)
  • Virology (AREA)
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  • Wood Science & Technology (AREA)
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  • Mycology (AREA)
  • Agricultural Chemicals And Associated Chemicals (AREA)

Abstract

The invention relates to a probiotic mixture containing an isolated strain of Bacillus thuringiensis var israelensis; an isolated strain of Trichoderma harzianum; and an isolated strain of Bacillus subtilis. The invention also relates to a biofungicide for controlling mycotoxigenic fungi, which comprises an effective amount of the probiotic mixture and at least one diluent. The invention further relates to a method for controlling mycotoxigenic fungi, which comprises applying an effective amount of the biofungicide to plant tissues useful in the preparation of alimentary mixtures.

Description

MEZCLA PROBIÓTICA, BIOFUNGICIDA Y MÉTODO, PARA EL CONTROL DE HONGOS MICOTOXIGÉNICOS PROBIOTIC MIXTURE, BIOFUNGICIDE AND METHOD, FOR THE CONTROL OF MYCOTOXIGENIC FUNGI
CAMPO TÉCNICO DE LA INVENCIÓN TECHNICAL FIELD OF THE INVENTION
La presente invención se relaciona con el campo técnico de la Biotecnología y con la Tecnología de los alimentos , debido a que aporta una mezcla probiótica, un biofunguicida y un método , para el control de hongos micotoxigénicos , en tej idos vegetales utili zados en la preparación de mezclas alimenticias , tales tej idos vegetales pueden ser preferentemente , granos y semillas . The present invention is related to the technical field of Biotechnology and Food Technology, because it provides a probiotic mixture, a biofungicide and a method for the control of mycotoxigenic fungi, in plant tissues used in the preparation of food mixtures, such plant tissues may preferably be grains and seeds.
ANTECEDENTES DE LA INVENCIÓN BACKGROUND OF THE INVENTION
Las micotoxinas son compuestos tóxicos producidos de forma natural por algunos tipos de hongos y/o mohos ; los cuales crecen en numerosos alimentos , tales como cereales , frutas desecadas , frutos secos y especias . Su crecimiento puede tener lugar antes o después de la cosecha, durante el almacenamiento o en el mismo alimento en entornos cálidos y húmedos . La mayoría de las micotoxinas son químicamente estables y persisten tras el procesamiento de los alimentos . Se han identi ficado varios cientos de micotoxinas , pero las más frecuentes que suponen un problema para la salud humana y del ganado con las aflatoxinas , ocratoxina A, patulina, fumonisinas , zearalenona y el nivalenol y desoxinivalenol . Las micotoxinas aparecen en la cadena alimentaria a consecuencia de la infección de los cultivos por hongos y/o mohos , sea antes o después de la cosecha . La exposición a las micotoxinas puede producirse directamente al comer alimentos infectados , o indirectamente , a partir de animales alimentados con comida contaminada, y en particular a partir de la leche . Mycotoxins are toxic compounds produced naturally by some types of fungi and/or molds; which grow in numerous foods, such as cereals, dried fruits, nuts and spices. Its growth can take place before or after harvest, during storage or in the food itself in warm and humid environments. Most mycotoxins are chemically stable and persist after food processing. Several hundred mycotoxins have been identified, but the most frequent ones that pose a problem for human and livestock health are aflatoxins, ochratoxin A, patulin, fumonisins, zearalenone and nivalenol and deoxynivalenol. Mycotoxins appear in the food chain as a result of the infection of crops by fungi and/or molds, either before or after harvest. Exposure to mycotoxins can occur directly by eating infected foods, or indirectly, from animals. fed contaminated food, and particularly from milk.
De ahi la importancia de generar productos químicos y biológicos , metodologías , para el control de las micotoxinas , como se puede ver en los siguientes documentos : Hence the importance of generating chemical and biological products, methodologies, for the control of mycotoxins, as can be seen in the following documents:
El documento de patente EP2011713789A se refiere a combinaciones de compuestos activos , en particular dentro de una composición fungicida, que comprende (A) una ditiino- tetracarboximida de fórmula ( I ) y al menos un agente de control biológico agricolamente beneficioso (B ) ; además se refiere a un método para controlar curativa o preventivamente los hongos f itopatógenos de plantas o cultivos , mediante la aplicación de una combinación según la invención para el tratamiento de semillas . También se refiere a composiciones y métodos para reducir el daño general de plantas y partes de plantas , asi como las pérdidas en frutas o verduras cosechadas causadas por hongos patógenos de plantas u otros microorganismos no deseados . En particular, se refiere a composiciones y métodos para proteger frutas , verduras , patatas y uvas durante la fase de crecimiento , alrededor de la cosecha y después de la cosecha . Patent document EP2011713789A refers to combinations of active compounds, in particular within a fungicidal composition, comprising (A) a dithiyne-tetracarboximide of formula (I) and at least one agriculturally beneficial biological control agent (B); It also refers to a method for curatively or preventively controlling phytopathogenic fungi of plants or crops, by applying a combination according to the invention for the treatment of seeds. It also refers to compositions and methods for reducing general damage to plants and plant parts, as well as losses in harvested fruits or vegetables caused by plant pathogenic fungi or other unwanted microorganisms. In particular, it relates to compositions and methods for protecting fruits, vegetables, potatoes and grapes during the growth phase, around harvest and after harvest.
El documento de patente EP2011156571A describe un método para controlar enfermedades f itopatogénicas en plantas útiles o en material de propagación de las mismas , que comprende aplicar a las plantas útiles , el lugar de las mismas o material de propagación de las mismas una combinación de los componentes A) y B ) en una cantidad sinérgicamente eficaz ; en el que el componente A) es un compuesto de fórmula ( I ) , en el que R1 es di f luorometilo o tri f luorometilo ; Y es -CHR2- o fórmula ( DD) y R2 es hidrógeno o alquilo C1-C6, o un tautómero de tal compuesto; y el componente B) es un compuesto seleccionado de compuestos conocidos por su actividad fungicida y/o insecticida, entre los que se mencionan algunas cepas de microorganismos, es particularmente eficaz para controlar o prevenir enfermedades fúngicas de plantas útiles. Patent document EP2011156571A describes a method for controlling phytopathogenic diseases in useful plants or in their propagation material, which comprises applying a combination of the components to the useful plants, their place or their propagation material. A) and B) in a synergistically effective amount; wherein component A) is a compound of formula (I), wherein R1 is di fluoromethyl or tri fluoromethyl; Y is -CHR2- or formula (DD) and R2 is hydrogen or C1-C6 alkyl, or a tautomer of such a compound; and component B) is a compound selected from compounds known for their fungicidal and/or insecticidal activity, among which some strains of microorganisms are mentioned, it is particularly effective for controlling or preventing fungal diseases of useful plants.
Finalmente, el documento de patente CA2928860 divulga un biopesticida que comprende un portador agricolamente adecuado, una cantidad efectiva de un pesticida fúngico, al menos un tensioactivo de ácido graso de sorbitán, y al menos un tensioactivo de éster de etoxilato de sorbitol. Entre los fungicidas se mencionan a Alternaría cassiae , Fusarium lateritum, Fusarium solani, Verticillium lecanii , Aspergillus parasiticus , Metarhizium anisopliae o Beauveria bessiana . Finally, patent document CA2928860 discloses a biopesticide comprising an agriculturally suitable carrier, an effective amount of a fungal pesticide, at least one sorbitan fatty acid surfactant, and at least one sorbitol ethoxylate ester surfactant. Among the fungicides, Alternaria cassiae, Fusarium lateritum, Fusarium solani, Verticillium lecanii, Aspergillus parasiticus, Metarhizium anisopliae or Beauveria bessiana are mentioned.
Si bien es cierto que los tres documentos de patentes antes citados, proponen o divulgan mezclas, composiciones, etc., las cuales comprenden en todos los casos, un ingrediente químico o biológico (biopesticida) ; y de los ingredientes biopesticidas , ninguna de las mezclas o composiciones de tales documentos de patentes citados, comprenden las cepas RVP BTI-20 Bacillus thuringiensi s var israelensis, ARS TH- 19 Trichoderma harzianum y ARS BS-19 Bacillus subtilis, para el control de hongos micotoxigénicos , tales como: Aspergillus spp . , Fusarium spp . , Penicillium spp . , Alternaría spp. y Claviceps spp.; los cuales tienen la capacidad de producir diferentes micotoxinas como aflatoxina, ocratoxina, zearalenona, DON, T2, fumonisina entre otras; las cuales son dañinas para los animales de ganado como vacuno, ovinos, etc., causándoles daños que van desde vómito y diarrea causando deshidratación hasta daño hepático o cáncer; por lo cual debe de ser controlada su concentración en los granos y/o semillas vegetales utilizados para forraje alimenticio. Although it is true that the three patent documents mentioned above propose or disclose mixtures, compositions, etc., which in all cases include a chemical or biological ingredient (biopesticide); and of the biopesticide ingredients, none of the mixtures or compositions of said patent documents cited, comprise the strains RVP BTI-20 Bacillus thuringiensi s var israelensis, ARS TH- 19 Trichoderma harzianum and ARS BS-19 Bacillus subtilis, for the control of mycotoxigenic fungi, such as: Aspergillus spp. , Fusarium spp. , Penicillium spp. , Alternaria spp. and Claviceps spp.; which have the capacity to produce different mycotoxins such as aflatoxin, ochratoxin, zearalenone, DON, T2, fumonisin among others; which are harmful to livestock animals such as cattle, sheep, etc., causing damage ranging from vomiting and diarrhea causing dehydration to damage liver or cancer; Therefore, its concentration in grains and/or vegetable seeds used for food forage must be controlled.
Es por ello que, con la finalidad de contribuir a la solución de los inconvenientes antes mencionados, se ha desarrollado una mezcla probiótica, un biofungicida y un método, para el control de hongos micotoxigénicos , en tejidos vegetales utilizados en la preparación de mezclas alimenticias, tales tejidos vegetales pueden ser preferentemente, granos y semillas . That is why, in order to contribute to the solution of the aforementioned drawbacks, a probiotic mixture, a biofungicide and a method have been developed for the control of mycotoxigenic fungi in plant tissues used in the preparation of food mixtures. Such plant tissues can preferably be grains and seeds.
Los detalles característicos de la presente invención, se muestran claramente en la siguiente descripción detallada, ejemplo y figuras que se acompañan meramente con la finalidad de ilustrar la concepción y algunas realizaciones preferentes de la invención. The characteristic details of the present invention are clearly shown in the following detailed description, example and figures that are accompanied merely for the purpose of illustrating the conception and some preferred embodiments of the invention.
BREVE DESCRIPCIÓN DE LAS FIGURAS BRIEF DESCRIPTION OF THE FIGURES
La figura 1 es una gráfica donde se ilustra el comportamiento del control de micotoxinas en granos de maiz, durante los 3 y 6 meses. Figure 1 is a graph that illustrates the behavior of mycotoxin control in corn grains, during 3 and 6 months.
DESCRIPCIÓN DE LA INVENCIÓN DESCRIPTION OF THE INVENTION
Un primer objeto de la presente invención es una mezcla probiótica, que comprende: i) una cepa aislada de Bacillus thuringiensis var israelensis; ii) una cepa aislada de Trichoderma harzianum; y iii) una cepa aislada de Bacillus subtilis . Una realización de la mezcla de la presente invención es cuando la cepa aislada de Bacillus thuringiensis var israelensis es la cepa RVP BTI-20 depositada con el número de depósito CM-CNRG TB193. Aunque no se descarta la posibilidad de que pudiera ser útil cualquier otra cepa aislada de Bacillus thuringiensis var israelensis. A first object of the present invention is a probiotic mixture, which comprises: i) an isolated strain of Bacillus thuringiensis var israelensis; ii) an isolated strain of Trichoderma harzianum; and iii) an isolated strain of Bacillus subtilis. One embodiment of the mixture of the present invention is when the isolated strain of Bacillus thuringiensis var israelensis is the RVP BTI-20 strain deposited with the deposit number CM-CNRG TB193. Although the possibility that any other isolated strain of Bacillus thuringiensis var israelensis could be useful is not ruled out.
Una realización más de la mezcla de acuerdo con la presente invención, es cuando la cepa aislada de Trichoderma harzianum es la cepa ARS TH-19 depositada con el número de depósito CM-CNRG TB174. Sin embargo, cabe la posibilidad de que pudiera ser útil cualquier otra cepa aislada de Trichoderma harzianum. A further embodiment of the mixture according to the present invention is when the isolated strain of Trichoderma harzianum is the ARS TH-19 strain deposited with the deposit number CM-CNRG TB174. However, it is possible that any other isolated strain of Trichoderma harzianum could be useful.
En una modalidad de la mezcla en cuestión, es cuando la cepa aislada de Bacillus subtilis es la cepa ARS BS-19 depositada con el número de depósito CM-CNRG TB175. También cabe la posibilidad de que pudiera ser útil cualquier otra cepa aislada de Bacillus subtilis . In one embodiment of the mixture in question, it is when the isolated strain of Bacillus subtilis is the ARS BS-19 strain deposited with the deposit number CM-CNRG TB175. It is also possible that any other isolated strain of Bacillus subtilis could be useful.
Otra realización preferente de dicha mezcla, es cuando las cepas aisladas están en una relación 1:2:2. Another preferred embodiment of said mixture is when the isolated strains are in a 1:2:2 ratio.
Un segundo objeto de esta invención es un biofungicida para el control de hongos micotoxigénicos , dicho biofugicida comprende, una mezcla probiótica de conformidad con la presente invención. A second object of this invention is a biofungicide for the control of mycotoxigenic fungi, said biofungicide comprising a probiotic mixture in accordance with the present invention.
Una realización del biofungicida en cuestión, es cuando controla los hongos micotoxigénicos: Aspergillus spp . , Fusarium spp., Penicillium spp., Alternarla spp., Claviceps spp., por citar algunos ejemplos. Un tercer objeto de la presente invención es un método para el control de hongos micotoxigénicos, el cual comprende, aplicar una cantidad efectiva del biofungicida propuesto por la presente invención, a tejidos vegetales infectados con hongos micotoxigénicos; preferentemente tejidos vegetales útiles en la preparación de mezclas alimenticias; por lo que la mezcla alimenticia puede ser un forraje para ganado vacuno, caprino, porcino, equino, etc. One embodiment of the biofungicide in question is when it controls mycotoxigenic fungi: Aspergillus spp. , Fusarium spp., Penicillium spp., Alternarla spp., Claviceps spp., to name a few examples. A third object of the present invention is a method for the control of mycotoxigenic fungi, which comprises applying an effective amount of the biofungicide proposed by the present invention to plant tissues infected with mycotoxigenic fungi; preferably plant tissues useful in the preparation of food mixtures; Therefore, the food mixture can be forage for cattle, goats, pigs, horses, etc.
Una realización del método de acuerdo con la presente invención es cuando la cantidad efectiva es de 0.3 mL por cada 1 Kg de tejido vegetal. An embodiment of the method according to the present invention is when the effective amount is 0.3 mL per 1 Kg of plant tissue.
EJEMPLOS DE LA INVENCIÓN EXAMPLES OF THE INVENTION
La razón de ser de los siguientes ejemplos es la de ilustrar la concepción de la presente invención y algunas de sus realizaciones preferentes. Por lo tanto, tales ejemplos no deben ser considerados como una limitante para el alcance de protección de dicha invención. The purpose of the following examples is to illustrate the conception of the present invention and some of its preferred embodiments. Therefore, such examples should not be considered as a limitation for the scope of protection of said invention.
Ejemplo 1. Evaluación del efecto de control del biofungicida que contiene a la mezcla probiótica en el control de hongos micotoxigénicos en maiz (Zea mays L.) Example 1. Evaluation of the control effect of the biofungicide containing the probiotic mixture in the control of mycotoxigenic fungi in corn (Zea mays L.)
Ob etivo Objective
Evaluación del biofungicida que contiene una mezcla probiótica de: las cepas RVP BTI-20 (Bacillus thuringiensis var israelensis) r ARS TH-19 (Metarhizium anisopliae) y ARS BS-19 (Bacillus subtilis) en el control de hongos micotoxigénicos en granos de maiz (Zea mays L.) forrajero. Metodología Evaluation of the biofungicide containing a probiotic mixture of: RVP BTI-20 (Bacillus thuringiensis var israelensis) r ARS TH-19 (Metarhizium anisopliae) and ARS BS-19 (Bacillus subtilis) strains in the control of mycotoxigenic fungi in corn grains (Zea mays L.) forager. Methodology
Se utilizaron granos de maíz (Zea mays L.) forrajero, a los cuales se les cuantificó la cantidad de las micotoxinas: Aflatoxina, Ocratoxina, DON, T2 y Fumonisinas presentes en los granos; para después inocularlos con el fitopatógeno Fusarium spp. y Aspergillus spp., para asegurar la presencia de micotoxinas. Forage corn grains (Zea mays L.) were used, to which the amount of mycotoxins was quantified: Aflatoxin, Ochratoxin, DON, T2 and Fumonisins present in the grains; and then inoculate them with the phytopathogen Fusarium spp. and Aspergillus spp., to ensure the presence of mycotoxins.
Los granos de maíz inoculados fueron separados en dos partes, donde una parte fue tratada con el biofungicida que comprende a la mezcla probiótica que a su vez contiene: las cepas RVP BTI-20 (Bacillus thuringiensis var israelensis) r ARS TH-19 (Metarhizium anisopliae) y ARS BS-19 (Bacillus subtilis) . Se realizaron dos mediciones de las micotoxinas, a los 3 y 6 meses . The inoculated corn grains were separated into two parts, where one part was treated with the biofungicide that includes the probiotic mixture that in turn contains: the RVP BTI-20 (Bacillus thuringiensis var israelensis) r ARS TH-19 (Metarhizium) strains. anisopliae) and ARS BS-19 (Bacillus subtilis). Two measurements of mycotoxins were carried out, at 3 and 6 months.
Resultados Results
De acuerdo con los resultados que se observan en la figura 1, a manera de conclusión podemos decir que el biofungicida que comprende la mezcla probiótica: cepas RVP BTI-20 (Bacillus thuringiensis var israelensis) , ARS TH-19 (Trichoderma harzianum) y ARS BS-19 (Bacillus subtilis) , sí mostró una sinergia para el control de hongos micotoxigénicos , tales como, Aspergillus spp. , Fusarium spp., Penicillium spp., Alternaría spp. y Claviceps spp.; los cuales tienen la capacidad de producir diferentes micotoxinas como aflatoxina, ocratoxina, zearalenona, DON, T2, fumonisina entre otras; las cuales son dañinas para los animales de ganado como vacuno, ovinos, etc., causándoles daños que van desde vómito y diarrea causando deshidratación hasta daño hepático o cáncer; por lo cual debe de ser controlada su concentración en los granos utilizados para f orra j e . According to the results seen in Figure 1, in conclusion we can say that the biofungicide that comprises the probiotic mixture: RVP BTI-20 (Bacillus thuringiensis var israelensis), ARS TH-19 (Trichoderma harzianum) and ARS strains BS-19 (Bacillus subtilis) did show synergy for the control of mycotoxigenic fungi, such as Aspergillus spp. , Fusarium spp., Penicillium spp., Alternaria spp. and Claviceps spp.; which have the capacity to produce different mycotoxins such as aflatoxin, ochratoxin, zearalenone, DON, T2, fumonisin among others; which are harmful to livestock animals such as cattle, sheep, etc., causing damage ranging from vomiting and diarrhea causing dehydration to liver damage or cancer; Therefore, its concentration in the grains used for forage must be controlled.

Claims

REIVINDICACIONES
1. Una mezcla probiótica, caracterizada porque comprende: i) una cepa aislada de Bacillus thuringiensis var israelensis ; ii) una cepa aislada de Trichoderma harzianum; y iii) una cepa aislada de Bacillus subtilis . 1. A probiotic mixture, characterized in that it comprises: i) an isolated strain of Bacillus thuringiensis var israelensis; ii) an isolated strain of Trichoderma harzianum; and iii) an isolated strain of Bacillus subtilis.
2. La mezcla de la reivindicación anterior, donde la cepa aislada de Bacillus thuringiensis var israelensis es la cepa RVP BTI-20 depositada con el número de depósito CM- CNRG TB193. 2. The mixture of the preceding claim, wherein the isolated strain of Bacillus thuringiensis var israelensis is the RVP BTI-20 strain deposited with the deposit number CM-CNRG TB193.
3. La mezcla de la reivindicación 1, donde la cepa aislada de Trichoderma harzianum es la cepa ARS TH-19 depositada con el número de depósito CM-CNRG TB174. 3. The mixture of claim 1, wherein the isolated strain of Trichoderma harzianum is the ARS TH-19 strain deposited with the deposit number CM-CNRG TB174.
4. La mezcla según la reivindicación 1, donde la cepa aislada de Bacillus subtilis es la cepa ARS BS-19 depositada con el número de depósito CM-CNRG TB175. 4. The mixture according to claim 1, wherein the isolated strain of Bacillus subtilis is the ARS BS-19 strain deposited with the deposit number CM-CNRG TB175.
5. La mezcla de acuerdo con la reivindicación 1, donde las cepas están en una relación 1:2:2. 5. The mixture according to claim 1, wherein the strains are in a 1:2:2 ratio.
6. Un biofungicida para el control de hongos micotoxigénicos , caracterizado porque comprende, una mezcla probiótica de conformidad con las reivindicaciones anteriores. 6. A biofungicide for the control of mycotoxigenic fungi, characterized in that it comprises a probiotic mixture in accordance with the preceding claims.
7. El biofungicida de la reivindicación anterior, donde los hongos micotoxigénicos son: Aspergillus spp . , Fusarium spp . , Penícíllíum spp . , Alternaría spp . , y/o Claviceps spp . 7. The biofungicide of the previous claim, where the mycotoxigenic fungi are: Aspergillus spp. , Fusarium spp. , Penícíllíum spp . , Alternaría spp . , and/or Claviceps spp.
8. Un método para el control de hongos micotoxigénicos , caracterizado porque comprende, aplicar una cantidad efectiva del biofungicida de conformidad con las reivindicaciones 6 y 7, a tejidos vegetales útiles en la preparación de mezclas alimenticias. 8. A method for the control of mycotoxigenic fungi, characterized in that it comprises applying an effective amount of the biofungicide in accordance with claims 6 and 7, to plant tissues useful in the preparation of food mixtures.
9. El método de la reivindicación anterior, donde la cantidad efectiva es de 0.3 mL por cada 1 Kg de tejido vegetal . 9. The method of the previous claim, where the effective amount is 0.3 mL per 1 kg of plant tissue.
10. El método según la reivindicación anterior, la mezcla alimenticia es un forraje para ganado. 10. The method according to the previous claim, the feed mixture is a livestock forage.
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