WO2023105361A1

WO2023105361A1 - Composition for controlling gastrointestinal parasites in ruminants and method for producing same

Info

Publication number: WO2023105361A1
Application number: PCT/IB2022/061658
Authority: WO
Inventors: Carlos Rafael Castillo Saldarriaga; Elizabeth Céspedes Gutiérrez; Diego Francisco CORTÉS ROJAS; Fredy Mauricio Cruz Barrera; Jaime Andrés Cubides Cárdenas; Martha GÓMEZ
Original assignee: Corporación Colombiana De Investigación Agropecuaria - Agrosavia
Priority date: 2021-12-10
Filing date: 2022-12-01
Publication date: 2023-06-15
Also published as: CO2021016747A1

Abstract

The present invention relates to compositions for controlling gastrointestinal nematodes in ruminants and to methods for producing said compositions. In particular, the compositions comprise fungal spores that are able to capture nematodes, and a coating emulsion that protects them from the chemical and biological conditions present in the animal's gastrointestinal tract, enhancing their effectiveness and allowing the long-term storage thereof even at high temperatures.

Description

COMPOSITION FOR THE CONTROL OF GASTROINTESTINAL PARASITES IN RUMINANTS AND THEIR PRODUCTION METHOD

FIELD OF THE INVENTION

The present invention relates to products for the biological control of parasites in ruminants. In particular, the present invention refers to parasiticide compositions based on the reproductive structures of fungal microorganisms with the capacity to capture nematodes, and production methods of said compositions.

BACKGROUND OF THE INVENTION

In livestock, the use of intensive dual-purpose production systems has been magnified worldwide. This increasingly recurrent practice has increased the risk of contagion of diseases by pathogenic microorganisms, which has led to the proliferation of reproductive problems, the reduction in daily milk production and the loss of weight of the animals, even being able to on some occasions develop diseases that cause their death.

The control of gastrointestinal nematodes has usually been carried out through the administration of chemical anthelmintics. However, the excessive use of this type of product has caused parasites to develop resistance, which has decreased the effectiveness of its application and has led to the need to administer high doses more frequently.

Likewise, the proliferation of the use of chemicals to deal with this problem has led to a greater incidence of other drawbacks, such as the bioaccumulation of the active principle in the meat and milk of the animal, which harms the safety of food and increases health risks. Additionally, the constant use of these compounds negatively affects the soil ecosystem, as well as it promotes unwanted changes in the beneficial microbial flora present in manure and in animals.

In this way, some biological control solutions have been developed in the art as an alternative to the use of anthelmintics to guarantee the sustainability of these systems, ensuring the obtaining of innocuous food. These products are obtained from reproductive structures, such as conidia and chlamydospores, of fungi with the ability to capture nematodes, with Duddingtonia and Arthrobotrys being the most studied genera in terms of efficacy tests.

Thus, according to this type of development, the spores of the fungus are administered to the animal so that they pass through its digestive tract and are expelled in the feces. In this way, these spores germinate and form organs that trap the nematode larvae, interrupting their life cycle and controlling the spread of parasites in the pasture. However, the delicate nature of the biological agents used can negatively affect the result, since in this process these agents are subjected to different adverse environments.

Indeed, the different cavities of the animal's digestive system, such as the ruminal cavity, the abomasal cavity, and the small intestine, subject the fungus to different chemical and biological environments where certain conditions, such as pH, can change radically. Thus, there is a need in the art to properly protect the spores of the fungus during their passage through the gastrointestinal tract of the animal, without interfering with their germination at the exact moment in which it is required; guaranteeing that the effectiveness of the fungus is the highest possible.

Additionally, it has been shown in the art that the nematophagous activity of the spores gradually decreases over time and is highly dependent on environmental factors, which not only makes production, distribution and storage difficult, but also reduces the number of times in many cases. parasiticide effect that the fungus may have when used in the field. Therefore, protection means are also required to guarantee the stability of the microorganism and mitigate the negative impact that the passage of time and the incidence of the environment can have on its final nematophagous effectiveness.

Similarly, it is known in the art that the presence of a high load of nematodes in animals in the perinatal period has a substantial negative impact on the health of newborn animals, preventing their weight gain and contributing to a great extent to mortality. of the animals at the beginning of grazing. In this way, there is also a need for a safe composition that can be administered to lactating animals and that allows reducing the risk of infection in the offspring, promoting additional weight gain in the first months that improves their survival expectations. .

On the other hand, despite the fact that this type of fungus has been studied in the art and its control has been demonstrated in in vitro tests, its use on a large scale has been limited due to the low concentration obtained in the substrates and fermentation systems used for its production, thus making its implementation difficult.

It is known in the art that liquid fermentation systems for the production of microorganisms have the ease of scaling production as a great advantage over solid fermentation systems. However, the morphology of filamentous fungi is highly variable, ranging from freely dispersed mycelium to pellets of aggregated biomass. This complexity in morphology is a typical problem of fermentation in submerged cultures due to issues of transport phenomena, viscosity of the culture medium and productivity.

In solid cultures, it has been reported that in nutrient-deficient media (such as cornmeal agar) the production of chlamydospores is scarce or, failing that, immature structures are obtained. On the other hand, it is known in the art that the production of chlamydospores is a response to unfavorable growth conditions, such as nutrient deficiency. In the case of predatory fungi, it has been shown that biological activity against nematodes increases under prolonged starvation conditions. However, there is no consensus on the influence of the type and concentration of carbon and nitrogen sources, on the production of reproductive structures of filamentous fungi during submerged cultures.

In this way, there is also a need in the art to provide an appropriate production method for obtaining said nematophagous fungi, which not only allows the production of nematicidal compositions to be viable at higher volumes, but also guarantees that the structures of the spores obtained are the most appropriate to achieve effective nematicidal control.

Thus, based on the technical problems presented in the art related to the loss of nematophagous activity of the microorganism as it progresses through the gastrointestinal tract of the animal, the limited stability of the spores during storage for long periods of time at environmental conditions adverse conditions, and the effectiveness that is required in animals in the gestation period to reduce the mortality of the pups, there is a need for a composition that provides these desired characteristics. Likewise, a production process is required to achieve the best properties of the microorganism and produce the nematicidal composition on a large scale.

In such a way that, although efforts have been made to develop biological alternatives other than chemical anthelmintics, there are still no compositions or processes such as those described here that solve the previously mentioned problems of the prior art.

BRIEF DESCRIPTION OF THE INVENTION

The present invention refers to a composition based on fungi with the capacity to capture nematodes, for the control of gastrointestinal parasites in ruminants. This composition comprises a coating emulsion that enhances the ability of the fungus to capture nematodes in the fecal matter of animals by giving it appropriate protection that guarantees its integrity during its transit through the cavities of the animal's digestive tract, guaranteeing that the fungus germinates easy way and opportunely when required and allowing it to carry out its biocontrol activity in the most efficient way possible.

Thus, in one aspect of the invention, the parasitic composition comprises an effective amount of the fungal microorganism together with a coating emulsion. Said emulsion comprises, in established proportions, excipients such as a coating agent, a surfactant, a solvent and an oily vehicle. In this way, it is also achieved that the composition maintains a greater nematicidal capacity during its storage, even at high temperatures.

In another aspect, the present invention relates to a method for producing the composition based on nematophagous fungi. This method comprises a liquid fermentation process through which it is possible to obtain a higher concentration of chlamydospores in a short period of time, significantly and favorably impacting productivity and subsequent commercial application.

In one aspect of the invention, as part of said method, culture media are used that comprise ingredients such as glucose, yeast extract, sodium chloride, magnesium sulfate, talc, among others. Said culture medium, with its components and specific proportions, also allows obtaining chlamydospores with a robust structure and thick walls in seven days.

BRIEF DESCRIPTION OF THE FIGURES

Figure 1 shows the nematophagous activity obtained for parasiticide compositions according to some embodiments of the present invention, in an in vitro simulation of the cavities of the ruminal digestive tract.

Figure 2 shows the nematophagous capacity obtained for parasiticide compositions according to one embodiment of the present invention, stored for 6 months at a temperature of 20°C, in three types of packaging. Figure 3 shows the nematophagous capacity obtained for parasiticidal compositions according to an embodiment of the present invention, stored for 6 months at a temperature of 30°C, in three types of packaging.

Figure 4 shows the nematophagous capacity obtained for parasiticidal compositions according to an embodiment of the present invention in comparison with the fungus without coating emulsion, during the first 210 days, at a temperature of 20°C.

Figure 5 shows the nematophagous capacity obtained for parasiticidal compositions according to an embodiment of the present invention in comparison with the fungus without coating emulsion, during the first 210 days, at a temperature of 29°C.

Figure 6 shows the number of parasite eggs per gram of fecal material obtained in sheep after administering parasitic compositions to the animals according to some embodiments of the present invention.

Figure 7 shows the average number of larvae in the meadow (L3/Kg) in a paddock plot before grazing and during 2 rotation cycles, after administering parasiticide compositions according to an embodiment of the present invention.

Figure 8 shows the fortnightly weight gain of suckling lambs after dams were administered parasiticidal compositions in accordance with some embodiments of the present invention.

Figure 9 shows the effect of increasing glucose on chlamydospore production in culture media according to some embodiments of the present invention. Figure 10 shows the effect of the presence of talc on chlamydospore production in culture media according to some embodiments of the present invention.

Figure 11 shows the effect of combining glucose and talc conditions for the production of chlamydospores in culture media according to some embodiments of the present invention.

Figure 12 shows the effect of the medium and the scale of fermentation (Incubator and Bioreactor) on the production of chlamydospores in culture media according to some embodiments of the present invention.

Figure 13 shows the type B (a) and type C (b) chlamydospores observed in the bioreactor culture medium, and pellet-like structures made up of mycelium and chlamydospores (c), obtained by some modalities of the production method according to with the present invention.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to compositions for the control of gastrointestinal nematodes in ruminants, and methods for producing said compositions.

A first aspect of the invention corresponds to compositions that comprise an active ingredient together with a coating emulsion, wherein said coating emulsion allows the active ingredient to survive, under the best conditions, its passage through the gastrointestinal tract of an animal. In addition, the coating emulsion according to the present invention gives a good stability to the composition, allowing the activity of the active ingredient to be maintained for several months, even when the storage temperature is considerable and/or different types of containers. According to the present invention, the active ingredient corresponds to any fungal microorganism with nematicidal capacity. Indeed, any genus of fungi that has the ability to capture nematodes can be used in the composition according to the present invention.

In this sense, although the species belonging to these genera may present genetic differences among themselves, they share physiological, structural and morphological characteristics related to the ability to capture nematodes. Therefore, the composition of the present invention, which provides a convenient coating emulsion, can serve any species belonging to genera that have this faculty, since its functionality depends on the ability to form capture nets; capacity that is common to these microorganisms. Therefore, any species belonging to such genera can be used in the composition of the present invention. However, in one embodiment of the invention, the fungal microorganism is Duddingtonia sp or Arthrobotrys sp.

However, the coating emulsion developed allows the nematophagous fungus to cross in the best conditions the different environments to which it is subjected in its passage through the gastrointestinal tract (such as the rumen, abomasum and small intestine) until it is expelled. in the stool. Once there, the coating fades without impeding the effectiveness of the active ingredient, allowing the microorganism to develop and create the three-dimensional structures that capture the nematodes; thus avoiding its proliferation in the prairie.

In this way, the application of the composition according to the present invention substantially reduces infective larvae, presenting an effect against various gastrointestinal parasites, including Haemonchus contortus, Ostertagia spp., Haemonchus spp., Trichostrongylus spp., Ostertagia spp. , Oesophagostomum spp., Nematodirus spp., Bunostomum spp., Coopería spp., Parascaris eguorum, Strongyloides westeri and Oxyuris egui. According to an embodiment of the present invention, the composition for the control of gastrointestinal parasites in ruminants comprises at least 1 x 10 ³ reproductive structures/g of the fungus with the ability to capture nematodes. In a preferred embodiment of the invention, the composition has at least 1 x 10 ⁵ reproductive structures/g of said fungus. Within the reproduction structures that can be used in the composition of the present invention are spores, mycelium and especially chlamysdospores.

In another embodiment of the invention, the composition includes a coating emulsion comprising at least one coating agent, a surfactant, a solvent and/or an oily vehicle.

In a preferred embodiment of the invention, the coating agent, which is responsible for providing protection to the reproductive structures of the fungus against the acidic pH of the abomasum, is between 1 and 15% w/w, more preferably between 2 and 11% w/w, and even more preferably between 4 and 6% w/w, of the coating emulsion. In another preferred embodiment, said coating agent is selected from the group consisting of gum arabic, poly(methacylic acid-co-methyl methacrylate) 1:2, methyl methacrylate copolymers, bovine bone gelatin (130 bloom), cellulose acetate-phthalate , hydroxypropyl methyl cellulose phthalate, hydroxypropyl methyl cellulose acetate succinate, polyvinyl acetate phthalate, or combinations thereof.

In a preferred embodiment of the invention, the surfactant, which reduces the surface tension between the aqueous phase and the oily phase and allows them to mix and form an emulsion, is between 1 and 20% w/w, more preferably between 2 and 5% w/w, of the coating emulsion. In another preferred embodiment said surfactant is selected from the group consisting of polysorbate 20, polysorbate 60, polysorbate 80, sorbitan 20 monooleate, sorbitan 40 monooleate, sorbitan 60 monooleate, sorbitan 80 monooleate, polyoxyethylene 30 lauryl ether, polyoxyethylene lauryl ether 35, polyoxyethylene 52 lauryl ether, polyoxyethylene 56 lauryl ether, polyoxyethylene 58 lauryl ether, polyoxyethylene 72 lauryl ether, polyoxyethylene 76 lauryl ether, ether polyoxyethylene 99 lauryl acid, polyoxyethylene 52 acid, polyoxyethylene 45 acid, polyoxyethylene 53 acid, polyoxyethylene 59 acid, or combinations thereof.

In a preferred embodiment of the invention, the solvent, which fulfills the function of allowing the homogeneous dispersion of the coating agent, is between 30 and 90% w/w, more preferably between 35 and 85% w/w, and even more preferably between 60 and 80% w/w, of the coating emulsion. In another preferred embodiment, said solvent is selected from the group consisting of a solution based on phosphorus or potassium salts such as sodium acid phosphate, potassium acid phosphate, sodium phosphate, potassium phosphate, HEPES solutions (N-( acid 2-Hydroxyethyl) piperazine-N-2- ethane sulfonic acid), water, or combinations thereof

In another preferred embodiment of the invention, the oily vehicle, which is responsible for protecting the reproductive structures of the fungus from enzymatic processes and the action of microorganisms found in the ruminal digestive tract, is between 5 and 30% w/ p, more preferably between 6 and 26% w/w, and even more preferably between 8 and 12% w/w, of the coating emulsion. In another preferred embodiment, said oily vehicle is selected from the group consisting of palm oil, sunflower oil, olive oil, avocado oil, almond oil, peanut oil, sesame oil, soybean oil, beeswax, carnauba wax, mineral oil, or combinations thereof.

In a preferred embodiment of the invention, the coating emulsion can also comprise a pH regulator, this component contributes to the timely dissolution of the coating agent, keeping the pH in the appropriate range. The pH regulator is between 1 and 5% w/w, more preferably between 1 and 3% w/w, of the coating emulsion. In another preferred embodiment said pH regulator is selected from the group consisting of 6N sodium hydroxide, potassium hydroxide, calcium hydroxide, or combinations thereof.

In another preferred embodiment of the invention, the coating emulsion may further comprise a cosolvent. Said cosolvent decreases the viscosity of the dispersion of the coating agent, facilitating its application, and is between 0.1 and 40% w/w, more preferably between 0.3 and 10% w/w, and even more preferably between 0.5 and 3 % w/w, of the coating emulsion. In another preferred embodiment, said cosolvent is selected from the group consisting of 96% ethanol, isopropyl alcohol, butanol, methanol, acetone, toluene, chloroform, or combinations thereof.

In another preferred embodiment of the invention, the coating emulsion can also comprise a plasticizer. This excipient prevents the coating from breaking, improving its elongation and flexibility, and favors the adhesion of the coating agent. Said plasticizer is between 0.1 and 10% w/w, more preferably between 0.2 and 5% w/w, and even more preferably between 0.5 and 1.5% w/w, of the coating emulsion. In another preferred embodiment, said plasticizer is selected from the group consisting of triethyl citrate, oleic acid, glycerin, triacetin, polyethylene glycol (PEG) 300, polyethylene glycol (PEG) 400, polyethylene glycol (PEG) 600, polyethylene glycol (PEG) 1450, polyethylene glycol (PEG) ) 3350, polyethylene glycol (PEG) 8000, or combinations thereof.

In another preferred embodiment of the invention, the coating emulsion may further comprise a solid diluent. Said solid diluent increases the concentration of solids in the coating formulation, allowing a thicker layer to be created. This excipient is between 3 and 25% w/w, more preferably between 5 and 20% w/w, and even more preferably between 7 and 13% w/w, of the coating emulsion. In another preferred embodiment, said solid diluent is selected from the group consisting of kaolin, isolated soy protein, milk whey protein, casein, talc, corn starch, cassava starch, banana starch, soy flour, or combinations. thereof.

In another preferred embodiment of the invention, the coating emulsion can also comprise an enzymatic and microbiological protector, which prevents the action of digestive enzymes or microorganisms on the reproductive structures of the fungus. Said enzymatic and microbiological protector is between 8 to 22% w/w, plus preferably between 10 to 20% w/w, of the coating emulsion. In another preferred embodiment, said enzymatic and microbiological protector is selected from the group consisting of calcium carbonate, calcium hydroxide, or combinations thereof.

Additionally, the composition according to the present invention may be in the form of granules, dispersible granules, powder, dispersible powder, capsules, tablets or pellets. In a preferred embodiment of the invention, the composition is in the form of granules with a particle size between 100 and 6000 pm, and a specific gravity between 0.1 and 2, which can be easily mixed with the animal's food in such a way that it does not perceive changes in its diet, and that its passage through the digestive tract is as fast as possible. In another embodiment of the invention, the composition may be a liquid formulation such as a suspension or an emulsion.

In this way, from the modalities described above, a composition is obtained that comprises a coating emulsion that provides superior nematophagous capacity, by allowing the active ingredient to better withstand its passage through the gastrointestinal tract. Indeed, the compositions according to the present invention reach, on average, a percentage of nematophagy of 89.2%.

Thus, this nematophagous capacity means that, by administering the composition according to the present invention, the number of larvae found in the pasture is reduced by up to 94%. This allows its administration, in conjunction with other strategies such as rotational grazing and good nutrition, to eliminate the presence of nematodes in the pasture. This also favors the weight gain of lactating animals, for which the composition achieves an increase of up to 83%.

Additionally, the composition modalities according to the present invention obtain a better stability, which allows the composition to maintain its effectiveness in prolonged storage conditions, even at temperatures adverse. In this way, compositions according to the present invention can maintain a nematophagous activity greater than 85% even when stored for 6 months at a temperature of 30°C.

Likewise, the composition according to the present invention allows the controlling effect of the fungus to persist in the field even one month after suspending its administration. Likewise, due to the nature of the excipients used and being a biological product, unlike anthelmintics, it does not have a withdrawal time and does not generate residues in the animal's meat or milk, favoring the entire production chain.

On the other hand, a second aspect of the invention corresponds to methods for producing the composition for the control of gastrointestinal nematodes, described above. In particular, a process has been developed that uses a liquid fermentation system that allows reaching a good concentration of chlamydospores in less time, also obtaining chlamydospores with a robust structure and thick walls.

Thus, the method for producing the described composition comprises the steps of: a) producing a fungus with the ability to capture nematodes by liquid fermentation using a culture medium comprising glucose and talc, and b) adding a coating emulsion comprising at least one coating agent, one surfactant, one solvent and one oily vehicle.

In one embodiment of the invention, the biomass obtained in step a) is separated and concentrated to form a granulate with a moisture content between 3% and 30%, and the coating emulsion from step b) is added by coating said granulate. In another embodiment of the invention, the addition of the coating emulsion from step b) is carried out by mixing said emulsion directly on the biomass obtained in step a), and said mixture is subsequently allowed to dry until obtaining a humidity between 3 and 30 %.

In a preferred embodiment of the invention, the glucose concentration in the culture medium is 10 to 70 g/L, more preferably between 20 and 40 g/L.

In another preferred embodiment of the invention, the concentration of talc in the culture medium is 4 to 30 g/L, more preferably 15 to 25 g/L.

In another embodiment of the invention, the culture medium used in the production of the fungus by liquid fermentation may also comprise yeast extract, magnesium sulfate, and calcium chloride.

In a preferred embodiment of the invention, the concentration of the yeast extract in the culture medium is from 1 to 5 g/L, more preferably between 1.5 and 3 g/L.

In another preferred embodiment of the invention, the concentration of magnesium sulfate in the culture medium is 0.1 to 2 g/L, more preferably between 0.1 and 0.9 g/L.

In another preferred embodiment of the invention, the concentration of calcium chloride in the culture medium is 0.1 to 2 g/L, more preferably between 0.1 and 0.9 g/L.

In another embodiment of the invention, the culture medium used in the production of the fungus by liquid fermentation may also comprise potassium acid phosphate in a concentration of 0.5 to 3 g/L, ammonium sulfate in a concentration of 0.5 at 3 g/L, sodium chloride at a concentration of 5 to 40 g/L, potassium chloride at a concentration of 5 to 40 g/L, sucrose at a concentration of 0.1 to 2 g/L, and/or fructose in a concentration of 0.2 to 2 g/L. Additionally, in one embodiment of the method, a pH of 5 to 7, a temperature of 28°C, stirring at 150 rpm and an air flow of 1 vvm (3L/min) are maintained during the process.

The addition of several of the components described in the culture medium favors osmotic stress conditions, which forces microorganisms to adopt adaptive responses that allow them to survive and proliferate. Responses to these conditions include the production of osmoprotective compounds such as glycerol, cytoskeletal reorganization, and cell wall biogenesis, which allows fungi to regulate intracellular osmotic pressure. On the other hand, the addition of substances such as salts and sugars decreases the water activity of the culture medium, causing hyperosmolarity and affecting the morphology of the fungus, mycelial growth, sporulation, and the production of metabolites. The addition of talc allows the formation of Biopellets (spherical agglomerates of mycelium) during liquid fermentation, which provides operational advantages of scale in terms of aeration/agitation, rheology, and subsequent biomass separations.

In this way, although the response to different stress and salinity conditions is highly dependent between genera and even species of fungi, according to the culture medium of the present invention it was found that, by adding the aforementioned components and in the indicated proportions, a better production of chlamydospores and a higher viability for nematophagous fungi, especially Duddingtonia sp., are achieved.

Therefore, using the method for producing the composition for the control of gastrointestinal parasites according to the present invention, a good concentration of chlamydospores is achieved in a shorter time, furthermore obtaining chlamydospores of robust structure with thick walls. Indeed, this method makes it possible to reach a minimum value of around 3 xlO ⁵ Clam mL ^-1 , even in periods of only 4 to 7 days, maintaining the nematophagous activity of the fungus above 90%. The foregoing, together with the steps of adding the coating emulsion comprising the excipients described above, in the specified proportions, makes it possible to obtain the composition with the advantages that have been exposed, and to produce it on a large scale.

EXAMPLES

Example 1. Concentration, viability and nematophagous activity in vitro Eight types of composition were tested using the fungus Duddingtonia sp. These compositions exhibited variations in emulsion coating as shown in Table 1.

Tab to 1

The developed prototypes were characterized in relation to concentration (chlamydospores/g), viability (UFC/g) and in vitro nematophagous activity (% nematophagy), which were determined according to the following procedures:

Concentration (chlamydospores/ ): The count was made from a 5g dilution of each modality in 50 mL of water, an aliquot of 10 pL was taken and the chlamydospores were counted in a Neubauer chamber using an optical microscope. with the 40X objective. This reading was carried out in the 4 squares divided into 16 quadrants of the chamber and finally the concentration of chlamydospores was determined using the following formula. a * 10,000 * b Concentration = - c

Where: a, is the number of cells (Chlamydospores), 10,000 is the Neubauer chamber correction factor, b, is the reciprocal of the dilution, and c, is the number of frames (4) counted in the chamber.

Viability determined as colony-forming units (UFC/a): from a dilution of 5g of each modality in 50 mL of water, serial dilutions were made in a 0.1% (v/v) Polysorbate 80 solution of According to the concentration, 100 pL of the selected dilutions were subsequently inoculated in triplicate in Petri dishes with YMA Agar plus Triton X-100 and the sample was distributed with a sterile Drigalsky spatula. After 6 days of incubation at 28 °C ± 2 °C, the colony-forming units (CFU) were counted for each sown dilution and the viability of the chlamydospores was determined, reporting the average count made in the Petri dishes. .

In vitro nematophakic activity: Initially, the concentration of chlamydospores of the sample to be evaluated was determined in order to determine the volume necessary to reach a concentration of 1.0xl0 ⁶ chlamydospores per box, inoculated by dripping in different areas of the Petri dish. with agar water. 8 boxes were inoculated per treatment or sample to be evaluated. After 72 hours of incubation at room temperature and darkness, the growth of the fungus was verified by observation under an inverted microscope and the necessary volume of a suspension of Panagrellus redivivus nematodes was added to put 200 larvae on each plate containing the fungus, additionally Larvae were placed in Petri dishes with water agar without fungus, this being the test control. After 48 hours of the fungus-larva interaction at room temperature and darkness, the free larvae were separated, that is, those that did not are captured by the fungus on agar using the Baermman funnel technique. For this, the agar was placed in the funnel and covered with water at 37 °C, after 12 hours the contents of the receiving tubes were collected, approximately 100 pL of lugol were added and the larvae count recovered from each replicate of the sample evaluated and the control. The percentage of nematophagy was calculated using the following equation.

Where X L3 Control is the average number of larvae recovered from the control and X L3 Treatment is the average number of larvae recovered in the treatment.

The biomass necessary for the elaboration of the prototypes was obtained from the growth of the fungus in solid fermentation, using broken rice as a substrate. The results for the evaluated compositions are shown below:

Table 2 Table 2 presents the results of the characterization of the composition modalities according to the present invention. Regarding the concentration, it is observed that all the modalities presented results higher than 1x10 ⁷ chlamydospores/g and no significant differences were observed between them. This order of concentration obtained facilitates the administration of the composition, since it makes it possible to administer a low amount of it to achieve the required dosage. Likewise, these concentrations will allow the dilution of the prototype with some nutritional supplement to facilitate the administration of the fungus to the animals.

With respect to viability, it is observed that the data are similar between the modalities and that they are in an exponential unit lower than the concentration result, that is, the concentration is of the order of lxlO ⁷ chlamydospores/g and the viability lxlO ⁶ CFU/g . In this way, no significant differences were observed in the viability of the chlamydospores of the different modalities, which allows us to infer that the formulation process and the components of the coating emulsion do not affect this response, obtaining satisfactory results for all the modalities of treatment. the present invention.

Regarding the nematophagous activity, it is observed that all the modalities had a percentage of around 80% or higher. In fact, the compositions according to the present invention obtained, on average, a percentage of nematophagy of 89.2%. In this way, it is evident that the added excipients allow the composition to achieve an important nematophagous capacity, which is essential to achieve the expected effect of the microorganism.

In accordance with the foregoing, it is concluded that the composition according to the present invention obtains favorable results in concentration, viability and nematophagous activity in vitro. Example 2. Nematophagous activity under digestive tract conditions

The composition according to the present invention allows to achieve a good nematophagous activity by protecting the fungus in an appropriate way during its passage through the gastrointestinal tract of the animal.

In order to evaluate the protection provided by the coating emulsion according to the present invention, an in vitro evaluation was carried out simulating the three most representative cavities of the ruminant gastro-intestinal tract: ruminal, abomasal and small intestine cavities.

The types of coating emulsion according to the present invention that were evaluated in this test correspond to compositions B, E and H, described in Table 1. Additionally, the performance of the chlamydospores of the fungus without any type of coating was evaluated. (control) in order to comparatively analyze the results. The procedure performed is described below:

Rumen cavity: The test began by preparing artificial saliva, which was brought to a temperature of 39 °C ± 2 °C and was subsequently gassed with CO2 until reaching a pH of 6.5 to 7.0. Simultaneously, in a container previously tempered at 90 °C, 100 mL of ruminal fluid were collected from a healthy fasting fistulated sheep. A 4:1 mixture of Saliva-ruminal fluid was made, adding 80 mL of filtered ruminal fluid to 320 mL of constantly gassed saliva.

Next, 0.8 g of kikuyu grass and 5 g of the evaluated composition modality were added to a container to adjust the effective working volume to an approximate concentration of 1x10 ⁶ chlamydospores/mL, later 80 chlamydospores were added to this container. mL of the mixture of ruminal fluid and saliva while each container was gassed. The test was carried out in an incubator with constant orbital shaking at 150 rpm and 39 °C ± 2 °C. The sampling was carried out at 16 hours, in each sampling it was guaranteed that the containers kept the temperature when being sampled on a heating plate (39 °C ± 2 °C), the containers were gassed with CO2 with the mixture of saliva and ruminal fluid, the pH was adjusted in each sampling in the two treatments with solutions of 3N NaOH or 0.5% HCI as required, and the pH was adjusted to that established for the test (6.5-7.0 rumen). .

A volume of 2 mL was sampled for each container, and it was replaced by 2 mL of the saliva/ruminal fluid mixture, or only saliva, kept at 39 °C ± 2 °C. The samples were centrifuged at 5000 rpm for 3 minutes, the supernatant was decanted and later washed with sterile water, centrifuging with the aforementioned conditions, in order to stop the effect that the ruminal fluid could have on the nematophagous fungus. The pellet was then resuspended in 2 mL of sterile water. Finally, in each sample, the nematophagous activity was determined.

Abomasal cavity: Once the ruminal cavity procedure was completed, the samples were treated to simulate the abomasal cavity in which the pH decreases to 2 and pepsin is added. For this, a 50% (v/v) HCI solution was prepared with which the pH of the samples was adjusted to 2.5. Subsequently, a 50 mg/mL pepsin stock solution was prepared and the required amount was added to the samples to obtain a final pepsin concentration of 4.75 mg/mL. Each Erlenmeyer was closed with a rubber stopper fitted with a valve to regulate the gas outlet. The sampling was carried out after 2 hours, following the same procedure described for the rumen cavity, for the determination of the nematophagous capacity.

Small intestine: Once the exposure time to the abomasal cavity ended, the samples passed through the simulation of the small intestine. For this, the pH was adjusted to 8.0 with 6N NaOH, then powdered pancreatin (Sigma-Aldrich, P1750) was added to obtain a concentration of 3.38 mg/mL. Sampling was done after of 5 hours of exposure, following the same procedure described for the rumen cavity for the determination of the nematophagous capacity.

Figure 1 shows the results of the nematophagous activity of the composition modalities according to the present invention, after being subjected to the physicochemical conditions found in the ruminal, abomasal and small intestine cavities, successively.

As can be seen in this figure, the modalities according to the present invention reach at the end of the run a substantially higher nematophagous capacity compared to the fungus without coating emulsion (control). Indeed, compositions H, B and E obtained an increase in the final nematophagous capacity of 23%, 32% and 25%, respectively, compared to the nematophagous capacity reached by the control.

In this way, at the end of the passage through the three cavities, satisfactory results were obtained for all the modalities evaluated. It is observed that the compositions H, B and E presented variations throughout the test, so that the nematophagous capacity increased with successive exposure to the cavities, reaching the values of 84%, 89% and 85% at the end of the test. , respectively. This behavior demonstrates the ability of the coating emulsion developed to protect the fungus in a pH-dependent manner.

From the above, it is further evidenced that the inclusion of an oily vehicle in the coating emulsion, which is present in all the compositions according to the present invention, has a low digestibility due to the metabolic characteristics of the digestive tract of ruminants, which contributes to obtain the effect of protecting the reproductive structures of the fungus from enzymatic processes and microorganisms that are found in the digestive tract and that affect the performance of the fungus, allowing it to obtain better nematophagy results at the end of its journey. In this way, it is observed that the compositions according to the present invention provide adequate protection to the fungus, so that the nematophagous activity obtained when passing through the gastrointestinal tract is much greater than that achieved only by the fungus without coating.

Example 3. Stability of the composition during storage

3.1 Evaluation of different containers

In order to evaluate the ability of the compositions according to the present invention to maintain their nematophagous capacity under different storage conditions, composition H was evaluated, and 3 types of packaging were compared: i) rigid containers with screw caps of terephthalate polyethylene (PET bottles) ii) high density polyethylene bags (Ziploc bags), and iii) vacuum-sealed trilaminate bags (metalized bags).

Each of these containers was selected based on the characteristics of the materials in terms of permeability to water vapor and oxygen. Indeed, the permeability to oxygen and humidity are characteristics of the packaging material used and can alter the physicochemical and microbiological properties of the product they contain.

Each experimental unit consisted of a container with 12 g of the composition. The containers were stored at two temperatures: (1) 20 °C and (2) 30 °C and sampling was carried out up to 6 months after the composition was packaged, evaluating the nematophagous activity in vitro and the concentration of chlamydospores under these conditions.

Figures 2 and 3 show the results achieved by the composition according to the present invention after 2 and 6 months of storage at the two temperatures (20°C and 30°C) and in the three preselected types of packaging. . It is observed that, for the composition according to the present invention, there are no significant differences between the packages evaluated at this sampling time. Additionally, although a decrease in nematophagous activity is evidenced with increasing storage temperature, considering that the percentage of nematophagy of the composition at time zero was 94.63%, it is observed that the composition according to the The present invention achieves that the decrease in nematophagy after two months is not, in most cases, greater than 5%. Even more, after 6 months of storage, the composition manages to maintain the percentage of nematophagy in any case above 85% (89% on average), even at temperature conditions of 30°C.

Additionally, the coating emulsion also allows the concentration of chlamydospores to not change over time. In this particular aspect, the composition according to the present invention obtained values in the order of lxlO ⁷ chlamydospores/g in each situation, similar to the concentration registered at the beginning of the study.

In this way, the composition according to the present invention achieves that, after 2 months of storage of the product, the nematophagous capacity of the microorganism is greater than 88% at temperatures of 20°C and 30°C, regardless of the type of container used.

Additionally, the composition according to the present invention ensures that the nematophagous capacity of the microorganism is maintained above 85%, even when it is stored for as long as 6 months and at a temperature as high as 30°C, at any time. container type.

3.2. Nematophagous capacity over time of the composition stored at different temperatures

Continuing with the evaluation of the stability at different temperatures of the composition of the present invention, the nematophagous capacity of the composition H modality was recorded in comparison with the fungus without coating (control), for a period of 7 months at 20 C and 29 C, in order to evaluate the contribution of the coating emulsion to the performance of the active ingredient.

The composition was packaged in 10 mL glass vials with screw caps, which were stored in climatic chambers with temperature control. For the analyzes at each sampling time (in vitro nematophagous capacity), 3 vials were arranged for each storage temperature.

According to the results obtained, shown in figures 4 and 5, it is observed that the composition according to the present invention maintains a better percentage of nematophagy compared to the fungus without coating, even during a period of 210 days. Indeed, during this period of time, the composition came to register a nematophagous capacity up to 20% higher than the nematophagous capacity granted by the uncoated fungus, this is due to the nature of the excipients used, as well as the specific proportions used.

From these results it is evident that the coating emulsion of the composition according to the present invention not only guarantees a forceful effect in the control of nematodes, by offering a better performance in the percentage of nematophagy, but also contributes to reducing the dose. of administration necessary to achieve the desired objective. This also allows the composition to be stored for a considerable time at high temperatures without the risk of losing its effectiveness.

Example 4. Reduction of the number of parasite eggs in fecal matter by action of the composition

As part of an in vivo test carried out to evaluate the effectiveness of the composition according to the present invention, 24 animals of the Hampshire, Corriedale and Creole breeds were selected, forming 4 experimental groups of 6 animals distributed according to gestation period (ultrasound transabdominal), type of delivery (only one multiple lambing ewes per group), and breed group (2 ewes of each breed in each group). The animals were not treated with any anthelmintic 90 days before gestation and were placed in four different batches, each one with three plots, in which they rotated independently. The pasture where the animals were kept was previously homogeneously contaminated by a group of 12 rearing lambs with an average HGP (approximately 1600).

For this experimental development, composition modalities B, E and H, described in table 1, were evaluated together with a control group to which neither the nematophagous fungus nor any other equivalent treatment was administered. The administration of the formulations was carried out during 5 days of the week in the morning hours.

All the compositions evaluated were administered during the last 30 to 45 days prior to parturition and up to 3 months after parturition, there were no adverse effects on the final development of the lambs or on the animals themselves during gestation. The females consumed the dose of the microorganism without presenting rejection of the ration or adverse gastrointestinal effects.

In order to evaluate the effectiveness of the formulations, fecal matter samples were taken periodically from the animals. The results of the excretion of eggs per gram of faeces (HPG) are shown in figure 6.

It is important to highlight that in the control group there were HPG counts higher than 6000 and Famacha values of 4-5, therefore, during the 75 to 90 days two animals in this group were dewormed, which explains their reduction from the day 75.

However, the results clearly show that the compositions according to the present invention show a significant reduction in the number of nematode eggs compared to the control. Indeed, the composition according to the present invention achieved a reduction of up to 78%. Additionally, during the trial two grazing cycles were performed. The grazing scheme consisted of an occupation period of 21 days and 42 days of pasture rest.

In this way, the quantity of larvae in the meadow (L3 Kg /DM) after the departure of the animals from the rotation was evaluated, taking two samples (approximately 400 g) from the plot with a previously defined zigzag type sampling, which were processed with washings, subsequent larval migration and drying to calculate the dry matter in the samples.

According to the results, shown in figure 7 for the plot, the contamination of the pasture decreased considerably after the second cycle of rotation when administering the compositions according to the present invention. Indeed, the composition modalities H and E evaluated showed a significant reduction, reaching a reduction in the number of larvae of up to 94%.

In this way, it is evident that the periodic administration of the compositions according to the present invention, together with rotational grazing and good nutrition, can eliminate the presence of nematodes in the meadow. The results obtained show a drastic decrease in eggs in periods as short as 120 days of administration.

Example 5. Fortnightly weight gain of lactating lambs

For this experimental development, the administration of the compositions was carried out during 5 days of the week in the morning hours. Ewes began consuming the compositions approximately 30 to 45 days prior to calving, ensuring that at birth the pasture had a low load of gastrointestinal nematodes. Additionally, a group of sheep to which the composition was not administered (control) was evaluated. As shown in Figure 8, with respect to the weight gain of the lactating lambs, when the mothers continued to be supplemented with the nematophagous fungi, the lambs of the compositions according to the present invention gained more weight during lactation with respect to to the control group.

For example, the weighing of the lambs of the group to which the modality of composition H was administered had 9.66%, 71.66% and 83.93% greater weight gain than the control group, reaching an average weight gain 41% older. This may be related to the fact that females, being less parasitized thanks to the controlling effect of the composition according to the present invention, have a better condition for lactation, favoring weight gain in lambs.

The results obtained demonstrate that the administration of the composition according to the present invention is safe during the last third of pregnancy and during lactation. In other words, the composition according to the present invention managed to reduce peripartum egg excretion and improve the weight gain of lambs during lactation.

In this way, the suckling lambs of mothers to which the composition according to the present invention was administered had a gain up to 40% higher than lambs of mothers that were not fed with this composition. Demonstrating its differential effect.

Example 6. Design of culture medium for the production of chlamydospores: effect of increased glucose

Within the design strategy of the liquid culture medium for the production of chlamydospores of the nematophagous fungus, the M culture medium was evaluated, whose components are shown in table 3. This particular medium obtained a yield of 1.23 ± 0.69 xlO ⁴ Clam mL ^-1 , 1.07±0.9 xlO ⁷ Clam g ¹ dry biomass.

Table 3

From this means, the possibility of having a limitation in the carbon source was considered. In this sense, two treatments with higher glucose concentration (coded as M-3% and M-7%) were evaluated, in order to increase carbon availability and maintain osmotic stress on the fungus. In these trials, medium M was used as a control and samples were taken at 7 and 14 days.

As part of the experimental development, fermentations were carried out with an effective working volume of 100 mL of medium, inoculated with 4 agar discs of the fungus with 8 days of growth and kept at a constant agitation of 150 rpm at 28±2°C. in a refrigerated incubator for 14 days. Samples of the fermentative medium were taken at 7 and 14 days of incubation to determine the concentration of chlamydospores (Clam mL-1), viability (UFC mL-1) and biological activity (% nematophagy).

As can be seen in figure 9, the results obtained after 7 days of sampling for the treatments M-3% (1.86±1.01 xlO ⁴ Clam mL ¹ ) and M-7% (2.41±1.35 xlO ⁴ Clam mL ^-1 ) presented concentrations of chlamydospores higher than the control (1.23±0.69 xlO ⁴ Clam mL ^-1 )

Similarly, after 14 days of sampling, the concentrations of chlamydospores for the treatments M-3% (1.09±0.43 x 10 ⁶ Clam mL ^-1 ) and M-7% (5.04±8.74 xlO ⁴ Clam mL ^-1 ) were equally superior to the control (1.95±1.25 xlO ⁴ Clam mL ^-1 ). However, the Chlamydospore concentration for the M-3% treatment presented a substantial change with respect to that obtained after 7 days of fermentation, indicating a high growth rate of the fungus during the second week of incubation.

Indeed, as shown in Figure 9, the M-3% treatment reached a much higher concentration at 14 days than the control and the M-7% treatment.

Additionally, the increase in the concentration of glucose in the medium increased the viability of the fungus at 7 days in the treatments M-3% (3.37±0.49 xlO ⁶ CFU mL ^-1 ) and M-7% (3.45±0.54 xlO ⁶ UFC mL ¹ ), with respect to the control (2.82±0.30 xlO ⁶ UFC mL ¹ ). Similarly, at 14 days, the viabilities reached by the M-3% (9.33±4.00 xlO ⁶ CFU mL ^-1 ) and M-7% (9.22±2.91 xlO ⁶ CFU mL ^-1 ) treatments were significantly higher. compared to the control (4.00±1.76 xlO ⁶ CFU mL ^-1 ).

Regarding the nematophagous capacity, an increase was also registered for the M-3% (98.69%) and M-7% (91.34%) treatments, compared to the M control (83.48%).

The results found allow us to conclude that the addition of around 3% (30 g/L) of glucose to the medium produces a significant increase in the concentration of chlamydospores, viability and nematophagous activity of the nematophagous fungus D.flagrans with respect to the control.

In this sense, very high concentrations of glucose in the medium seem to have a negative effect on the growth and activity of the fungus due to extreme conditions of osmotic stress.

Example 7. Design of culture medium for the production of chlamydospores: Effect of osmotic stress conditions on the medium

In order to favor the growth of the fungus and improve the production of chlamydospores, osmotic stress conditions were evaluated by adding two concentrations of talc microparticles: 5 g L ¹ and 20 g L . In this test, the Sabouraud liquid medium and the M culture medium, described in Table 3, were implemented as control. The experimental development carried out corresponds to that described in the previous example, using a refrigerated incubator.

As shown in figure 10, the results obtained after 7 days of sampling with the treatments with 5 g L ¹ of talc (3.38±0.96 xlO ⁴ Clam mL ¹ ) and 20 g L ¹ of talc (5.44±1.45 xlO ⁴ Clam mL ^-1 ) were significantly higher than those obtained with the Sabouraud controls (2.25±2.09 xlO ⁴ Clam mL ¹ ) and the culture medium M (1.38±0.67 xlO ⁴ Clam mL ^-1 )

A similar behavior is observed after 14 days of sampling, where the results obtained with 5 g L ¹ of talc (6.19±2.19 xlO ⁴ Clam mL ^-1 ) and 20 g L ¹ of talc (2.31±0.70 xlO ⁵ Clam mL ¹ ) were notably higher than those registered for the Sabouraud controls and the M culture medium, for which 1.84±0.86 xlO ⁴ Clam mL ¹ and 2.58±1.22 xlO ⁴ Clam mL ^-1 were registered, respectively.

In this way, as observed in figure 10, the highest concentration of chlamydospores was obtained for the culture media with the addition of talc, being significantly higher when 20 g L ¹ were added.

On the other hand, the biological activity was not affected by osmotic stress or by talc, in all the evaluated treatments the percentage of nematophagy was higher than 93%. The treatments with 5 g L ¹ of talc (93.25±4.12%) and 20 g L ¹ of talc (93.83±4.12%) showed a higher biological activity than the Sabouraud control treatments (92.99±4.21%) and the M culture medium. (90.65±4.74%).

The results found allow us to conclude that the addition of talc contributes to achieve better results of concentration and viability of the fungus, without affecting its nematophagous activity. Example 8. Design of culture medium for the production of chlamydospores: Combined effect of stress conditions

In this last phase of experimentation on a laboratory scale, the best environmental conditions found for the production of chlamydospores were evaluated: 20 g L ¹ of talc and 3% glucose. This culture medium, coded as O, is made up as shown in Table 4.

As can be seen in figure 11, no statistically significant differences were found between 7 and 14 days of sampling for the culture medium O (3.18±1.20 xlO ⁵ clam mL ¹ and 3.63±1.72 xlO ⁵ clam mL ^-1 , respectively). . This result indicates that the combined effect of adding 20 g L ¹ of D¡05 and 3% glucose to the culture medium allowed the fermentation time to be reduced from 14 days to a maximum of 7 days.

Additionally, no loss in the biological activity of the fungus was observed. In fact, this remained above 90%, with nematophagy obtained at 7 and 14 days being 93.1 ± 2.3% and 96.2 ± 2.4%, respectively.

Based on these results, it is concluded that the medium that incorporated talc together with glucose (Culture medium O) improved the yield during growth and the biological characteristics of the fungus in submerged culture. Additionally, the behavior of this medium was evaluated by bioreactor tests. As part of this experimental development, the fermentations were carried out in a 5 L bioreactor with an effective working volume of 3 L, adapted with a dissolved oxygen sensor. The medium was inoculated with 10% v/v of a chlamydospore suspension with a concentration of 10 ⁶ clam mL-1, prepared by scraping 5 boxes of wheat flour agar from a second passage of the fungus D. flagrans 8 days old. growth. The bioreactor was kept under constant agitation at 150 rpm and 28 ± 2 °C with an aeration of 1 vvm (3 L/min of sterile air) for 7 days. At the end of the fermentation, the biomass produced was recovered by centrifugation (4500 rpm, 20 minutes) discarding the supernatant.

The average concentration of chlamydospores obtained in this trial was 3.41±0.70 xlO ⁵ Clam mL ¹ , which was 3 times higher than at the beginning of fermentation.

Figure 12 shows the significant effect of the medium and the fermentation scale. Both at the incubator and bioreactor level, the significant difference of the O culture medium on the concentration of chlamydospores with respect to the M culture medium is clearly demonstrated.

In the same way, it is evident that the culture in a bioreactor with the culture medium O allowed to improve the production of chlamydospores (5.13±1.46 xlO ⁵ Clam mL ¹ ) in comparison with the result obtained in an incubator (3.18±1.20 xlO ⁵ Clam mL ^{- 1} ).

In this way, sufficient statistical evidence was found to affirm that culture in a bioreactor with culture medium O significantly improves the production of D. flagrans chlamydospores compared to culture medium M under the same fermentation conditions. This result is an indicator that culture medium O, unlike medium M, favors sporulation and not germination of the chlamydospores produced, favoring the production of the composition on a larger scale. Finally, the fermentation tests in the bioreactor with culture medium O allowed to obtain chlamydospores with a robust structure as shown in figure 13, with thick walls classified as type B and C (Figure 13-a and Figure 13-b). This type of wall offers greater resistance to the fungus against the physicochemical and microbiological conditions found in the ruminal digestive tract, allowing it to maintain its integrity until it reaches the fecal matter. In addition, the formation of pellet-like structures (Figure 13-c) made up of mycelium and chlamydospores was observed.

Claims

CLAIMS A composition for the control of gastrointestinal parasites in ruminants comprising: a. at least lxlO ³ reproductive structures/g of a fungus with the ability to capture nematodes; and b. a coating emulsion comprising a coating agent, a surfactant, a solvent and an oily vehicle, wherein the coating agent is between 1 and 15% w/w, the surfactant is between 1 and 20 % w/w, the solvent is between 30% and 90% w/w and the oily vehicle is between 5% and 30% w/w, of the coating emulsion. The composition according to Claim 1, wherein the coating emulsion also comprises a pH regulator in a proportion of 1 to 5% w/w. The composition according to Claim 1, wherein the coating emulsion further comprises a cosolvent in a proportion of 0.1 to 40% w/w/w/or a plasticizer in a proportion of 0.1 to 10% w/w. The composition according to Claim 1, wherein the coating emulsion also comprises a solid diluent in a proportion of 3 to 25% w/w/w/or an enzymatic and microbiological protector in a proportion of 8 to 22% w/w. The composition according to Claim 1, wherein the coating agent is selected from the group consisting of gum arabic, poly (methacylic acid -co-methyl methacrylate) 1:2, methyl methacrylate copolymers, gelatin

Four. Five bovine bone (130 bloom), cellulose acetate phthalate, hydroxypropyl methyl cellulose phthalate, hydroxypropyl methyl cellulose acetate succinate, polyvinyl acetate phthalate, or combinations thereof. The composition according to Claim 1, wherein the surfactant is selected from the group consisting of polysorbate 20, polysorbate 60, polysorbate 80, sorbitan monooleate 20, sorbitan monooleate 40, sorbitan monooleate 60, sorbitan monooleate 80, ether polyoxyethylene 30 lauryl ether, polyoxyethylene 35 lauryl ether, polyoxyethylene 52 lauryl ether, polyoxyethylene 56 lauryl ether, polyoxyethylene 58 lauryl ether, polyoxyethylene 72 lauryl ether, polyoxyethylene 76 lauryl ether, polyoxyethylene 99 lauryl ether, polyoxyethylene 52 acid, polyoxyethylene 45 acid, acid polyoxyethylene 53, polyoxyethylene 59 acid, or combinations thereof. The composition according to Claim 1, wherein the solvent is selected from the group consisting of a solution based on phosphorus or potassium salts such as sodium hydrogen phosphate, potassium hydrogen phosphate, sodium phosphate, potassium phosphate, HEPES (N-(2-Hydroxyethyl) piperazine- N-2-ethane sulfonic acid) solutions, water, or combinations thereof. The composition according to Claim 1, wherein the oily vehicle is selected from the group consisting of palm oil, sunflower oil, olive oil, avocado oil, almond oil, peanut oil, sesame oil, oil soy, beeswax, carnauba wax, mineral oil, or combinations thereof. The composition according to Claim 2, wherein the pH regulator is selected from the group consisting of 6N sodium hydroxide, potassium hydroxide, calcium hydroxide, or combinations thereof.

46 The composition according to Claim 3 wherein the cosolvent is selected from the group consisting of 96% ethanol, isopropyl alcohol, butanol, methanol, acetone, toluene, chloroform, or combinations thereof. The composition according to Claim 3 wherein the plasticizer is selected from the group consisting of triethyl citrate, oleic acid, glycerin, triacetin, polyethylene glycol (PEG) 300, polyethylene glycol (PEG) 400, polyethylene glycol (PEG) 600, polyethylene glycol (PEG) ) 1450, polyethylene glycol (PEG) 3350, polyethylene glycol (PEG) 8000, or combinations thereof. The composition according to Claim 4 wherein the solid diluent is selected from the group consisting of kaolin, isolated soy protein, whey protein, casein, talc, corn starch, cassava starch, banana starch, soybean meal, or combinations thereof. The composition according to Claim 4 wherein the enzymatic and microbiological protector is selected from the group consisting of calcium carbonate, calcium hydroxide, or combinations thereof. The composition according to Claim 1 wherein the fungus is Duddingtonia sp or Arthrobotrys sp. The composition according to Claim 1 that is in the form of granules, dispersible granules, powder, dispersible powder, capsules, tablets, pellets, emulsion or suspension. The composition according to Claim 15, wherein the composition in the form of granules has a particle size between 100 and 6000 pm.

47 The composition according to Claim 1 that presents a nematophagous activity of at least 80%. A method for producing the composition for the control of gastrointestinal parasites in ruminants, said method comprising the steps of: a) producing a fungus with the ability to capture nematodes by liquid fermentation using a culture medium comprising glucose and talc, and b) adding a coating emulsion comprising at least one coating agent, a surfactant, a solvent and an oily vehicle, wherein in the culture medium of step a. glucose is found at a concentration of 10 to 70 g/L, and talc is found at a concentration of 4 to 30 g/L. The method according to Claim 18, wherein the biomass obtained in step a) is separated and concentrated to form a granule with a humidity between 3% and 30%, and the addition of the coating emulsion from step b ) is performed by coating said granules. The method according to Claim 18, wherein the addition of the coating emulsion from step b) is carried out by mixing said emulsion directly on the biomass obtained in step a), and said mixture is subsequently allowed to dry until obtaining a humidity between 3 and 30%. The method for producing the composition for the control of gastrointestinal parasites in ruminants according to claim 18, wherein the culture medium from step a. it also comprises yeast extract in a concentration of 1 to 5 g/L, magnesium sulfate in a concentration of 0.1 to 2 g/L and/or calcium chloride in a concentration of 0.1 to 2 g/L. The method for producing the composition for the control of gastrointestinal parasites in ruminants according to claim 18, wherein the culture medium from step a. It also includes potassium hydrogen phosphate in a concentration of 0.5 to 3 g/L, ammonium sulfate in a concentration of 0.5 to 3 g/L, sodium chloride in a concentration of 5 to 40 g/L, chloride potassium in a concentration of 5 to 40 g/L, sucrose in a concentration of 0.1 to 2 g/L and/or fructose in a concentration of 0.2 to 2 g/L.