Formulation for biocontrol of insect pests in grain stores

Abstract

Disclosed is a liquid bait with high attractant and phagostimulant capacity, allowing insect pests in the store to consume, preferentially, the biological insecticide(s) incorporated within the formulation of the bait, causing their death by ingestion.

Description

FORMULATION FOR THE BIOLOGICAL CONTROL OF INSECT PESTS IN GRAIN STORAGE This patent application is an extension of application AR 2023 01 00449 filed on 24 / 02 / 2023 by the same Applicant.

[0001] The main purpose of this invention patent is to present a new technology developed exclusively for the biological control of insect pests in grain storage.

[0002] The new technology, called “Liquid Biocarrier,” is a liquid bait with a high attractant and phagostimulant capacity, allowing storage pest insects (hereafter referred to as insects) to preferentially consume the biological insecticide(s) incorporated into its formulation, causing their death by ingestion. It is used for the biological control of primary and secondary infestation insects (both larval and adult stages) and mites in stored grains in general, oilseeds, legumes, and organic grains in general (popcorn, flint corn, peanuts, nuts, etc.).

[0003] This new technology and type of formulation for the post-harvest handling of cereals and oilseeds, generically called "Liquid Biocarrier", consists of a substrate rich in proteins, sugars and carbohydrates, which acts as a food attractant (kairomone and phagostimulant), which incorporates The active ingredients of the biological insecticide. These active ingredients are obtained through extraction, fermentation, or biotechnological processes. State of the art prior to the invention:

[0004] Grain storage arises from the randomness and seasonality of agricultural production. The goal of grain storage is to preserve grains for extended periods without compromising their quality or safety. Direct consumption and industrial processing depend on maintaining their quality during transport and storage.

[0005] Quality deterioration is related to properties such as percentage of protein, starch, oil, baking quality, percentage of damaged grains and nutritional value, among others.

[0006] Stored grains have one main enemy: storage insect pests. Consequently, all post-harvest efforts must be clearly focused on preventing their development. Approximately 5-20% of production is lost due to insect damage during the post-harvest stage, depending primarily on three variables: the variety of grain stored, the degree of infestation, and the storage period.

[0007] Insect pests of stored grains cause direct damage through the consumption of dry matter and loss of commercial quality (broken grain, contamination, objectionable odor, etc.), and indirect damage such as grain heating, moisture migration, fungal growth, and mycotoxin formation. Numerous insect and mite species attack cereal and oilseed grains and are divided into primary and secondary pests. Primary pests attack whole, healthy grains. They generally complete most of their life cycle within the grain, and the adult insect leaves the grain damaged upon emergence. Secondary pests attack broken grains or grains previously damaged by primary pests. They also affect products obtained from grain milling.

[0008] Stored grains can become infested with insects through three main routes: • Field infestation: Infestation begins in the field crop, prior to harvest. The insects are transported to storage as adults along with the grain and / or as eggs or larvae within the grain. In the latter case, the infestation is difficult to detect through sampling and receiving analysis at the plant. • Infestation at the storage facility: Grain infestation comes from insect populations that remain in the storage facilities feeding on bulk waste that has not been properly removed through sanitation and cleaning practices. • Migratory infestation: This comes from flying insects, both from primary or secondary infestation, such as light insects and mites, which arrive in bulk transported by the wind, from the vicinity of the storage plant, transport, etc.

[0009] There are currently very few options available to control an infestation of insects and mites during the post-harvest of grains.

[0010] In general terms, there are two types of bulk chemical treatments widely used: • Preventive treatments: These consist of applying residual insecticides to the grain being stored. The aim is to prevent the development of pests during a certain storage period. Synthetic insecticides, in liquid or solid formulations, are used for this purpose. They are sprayed or sprinkled onto the bulk grain upon arrival of the new harvest at the processing plant, or during transshipment. • Curative treatments: fumigant gases are used to eliminate existing pests. They have no residual effect.

[0011] Currently, there is a limited supply of chemical groups of insecticides for the preventive control of insects and mites during the Post-harvest use of organophosphate and pyrethroid insecticides in cereals and oilseeds is subject to strict maximum residue limits in various markets worldwide, and resistance is widespread. Furthermore, major regulatory agencies globally (European Union, Brazil, Canada, United States, and others), and even the Codex Alimentarius Commission, are considering banning their use in post-harvest applications of cereals and oilseeds.

[0012] As an alternative to synthetic insecticides, biological insecticides (biochemical, botanical, or microbiological) have been developed, which are currently approved for use exclusively in extensive and intensive agriculture. With current application methods, their use is not applicable to post-harvest treatment of cereals and oilseeds, as they act systemically.

[0013] The application of the formulation of the present invention, called “Liquid Biocarrier,” represents an innovative technological solution to this problem. Firstly, “Liquid Biocarrier” acts as an attractive and phagostimulant substrate for the insect pest, serving as its preferred food source over the grain in question. Secondly, it contains an active ingredient capable of killing the insect pest through ingestion. It allows the use of active ingredients that are harmless to humans and animals, and is also compatible with organic agriculture.

[0014] These biological insecticides act exclusively through the digestive system, making them ineffective when sprayed directly onto the grain. Therefore, they must be carried in a food bait (kairomone). “Liquid Biocarrier” uses a vegetable oil with added food substrate / attractant bait / phagostimulant as a carrier. This ensures greater adhesion and persistence of the formulation on the treated grain. Objectives of the present invention:

[0015] The objective of this invention is to present a new technology developed from a formulation composed of a liquid vehicle and a food substrate that acts as an attractant / phagostimulant bait, serving as a carrier for the biological active ingredient. The formulation attracts the insect and promotes its consumption (phagostimulant), thereby ingesting the insecticidal active ingredient, which acts via the digestive tract, causing death. Death occurs after a single or repeated ingestions, depending on the type of insect.

[0016] The objective of the present invention is to present a new technology that employs biological insecticides, which act exclusively through the digestive system, and when associated with a food bait, it is ensured that the insect is attracted, goes in search of the food, ingests it (phagostimulant) and dies, this food being the carrier of the biological insecticide.

[0017] The objective of the invention is to present a new technology that can have countless compositions, these compositions having as common points the use of insecticides classified as "biological", being able to use in said formulation one or more of said biological active principles, being those currently available on the market or any other new type of active principle to be developed in the future, this being at least one active principle associated with a food bait with attractive properties (kairomones), carried in vegetable oil. Review of the invention:

[0018] NEW EXCLUSIVE TECHNOLOGY FOR BIOLOGICAL CONTROL OF INSECT PESTS IN GRAIN STORAGE, characterized as a liquid bait that has a high attractive and phagostimulant capacity, which allows the insect pests in storage to preferentially consume the biological insecticide(s) incorporated into its formulation, causing their death by ingestion. Detailed description of the present invention:

[0019] The active ingredients comprising the formulation of the present invention (Liquid Biocarrier) are substances considered to be of slight toxicological concern by FAO / WHO and which do not have residue tolerances for international trade (MRLs), with the exception of spinosyns, which despite Their biological origin has established MRLs in some countries, even though they are authorized for organic agriculture.

[0020] The different active ingredients that could be integrated into the formulation of the present invention (Liquid Biocarrier), depending on the grain and target pest to be controlled, are listed below: • Microbial disruptors of insect digestive membranes. Bacillus thuringiensis, its subspecies and the insecticidal proteins they produce. • Allosteric modulators of the nicotinic acetylcholine receptor - Spinosins • Baculovirus. Host-specific occluded pathogenic viruses. Granuloviruses (GVs), Nucleopolyhedroviruses (NPVs). • Plant extracts and crude oils • Azadirachtins • Entomopathogenic fungi • Peptides derived from natural toxins, whether of animal or plant origin, or derived from molecular biology techniques.

[0021] Food attractants can be of various compounds and their combination with at least one active ingredient gives rise to numerous compositions, the novelty of the invention being the association of a food substrate that includes one or more biological active ingredients, all this delivered in a liquid bait. It is important to note that although there is precedent for the use of pheromone-based sex attractants in post-harvest applications, these are specific to the species being sampled; that is, they are used for pest monitoring, but are not intended to stimulate and enhance insecticide consumption.

[0022] Generic composition of the formulation of the present invention: Vehicle: vegetable oil (sunflower, corn or soybean). Attractant / phagostimulant: wheat germ, proteins and carbohydrates (sugars). Active ingredient (one active ingredient or a combination of active ingredients). Food-grade stabilizers and preservatives.

[0023] In the formulation process of liquid Biocarrier, a mixer is used. The components of the feed substrate / attractant and phagostimulant bait are added first, followed by the biological active ingredient(s). Vegetable oil and the necessary preservatives and stabilizers are then incorporated to maintain the homogenization of the formulation. Finally, a sample is taken from the batch for quality analysis and if it meets the specifications, it is packaged, labeled and packed.

[0024] The new result and industrial application of the formulations of the present invention consists of the effectiveness in controlling storage insect pests, treating the grains with a biological insecticide associated with an attractant food carrier (kairomone). Currently, biological insecticides are not used post-harvest, as their use is ineffective because they act exclusively through the digestive system. If the biological insecticide is sprayed directly onto the grain (this being the only known application technique or methodology used prior to the present invention), the insect would be forced to eat the grain, and in turn, the small portion of the grain it eats would have to provide the required amount of insecticide to cause its death, which is impossible because: • Not all insects can attack whole grain, • With direct spraying, not all grains come into contact with the biological insecticide, • In those grains that do come into contact with the biological insecticide, the concentration of the insecticide on the grain is not sufficient to cause the death of the insect that ingests it.

[0025] Liquid Biocarrier is a new technology that attracts the insect above the mass of grains, the insect being attracted to this formulation, and when it goes in search of it, it preferentially ingests it and its death occurs, this result being not only novel, but also unprecedented in the art.

[0026] It has been proven through multiple tests that both primary and secondary infestation insects consume the substrate Food (feed bait or kairomone), plus the biological active ingredient that makes up the formulations of the invention (Liquid Biocarher). This causes the insect's death by ingestion of the biological insecticide. Agronomic efficacy trials:

[0027] Attached is a report of a series of evaluation trials of different formulations.

[0028] Prototype formulations evaluated in efficacy trials (Non-limiting example of a liquid Biocarrier Composition): a) Prototype formulation 1 b) Prototype formulation 2 c) Prototype formulation 3 d) Prototype formulation 4 e) Prototype formulation 5 e) Prototype formulation 6

[0029] As a result, it is concluded that the five liquid Biocarrier formulations of the present invention evaluated: • Biocarrier 1: 1% Spinosad, equivalent to a dose of 1 ppm / Tn grains; • Biocarrier 2: 1% Spinosad + 2% Azadirachtin, equivalent to a dose of 1 ppm Spinosad + 2 ppm Azadirachtin / Tn grains; • Biocarrier 3, 2% Bacillus thuringiensis subsp. tenebrionis, equivalent to a dose of 2 ppm of Bacillus thuringiensis subsp. tenebrionis / Tn grains; • Biocarrier 4: 2% Bacillus thuringiensis subsp. Kurstaki, equivalent to 2 ppm doses of Bacillus thuringiensis subsp. Kurstaki / Tn grains; • Biocarrier 5: 1% Spinosad + 2% Bacillus thuringiensis subsp. tenebrionis, equivalent to a dose of 1 ppm Spinosad + 2 ppm Bt tenebhonis / Tn grains. • Biocarrier 6: 1% Spinosad + 2% Bacillus thuringiensis subsp. Kurstaki, equivalent to doses of 1 ppm Spinosad + 2 ppm Bt Kurstaki / Tn grains. They achieved excellent control of a wide range of post-harvest cereal pests.

[0030] The trials demonstrated that the formulations of the invention are highly attractive to the eight species evaluated, and that the control strategy is efficient in achieving residual protection of the bulk for at least six months.

[0031] The trials fulfill the objective of demonstrating the versatility of the effectiveness of liquid Biocarrier, by using different active ingredients in its composition, in different concentrations and even in mixtures of them.

[0032] Furthermore, the formulations of the invention proved to be more effective than the most widely used commercial control in the post-harvest of cereals (synthetic insecticide - active ingredient Deltamethna), showing significantly higher levels of control than the practice of control by spraying the mass of grains with a residual synthetic insecticide.

[0033] Liquid Biocarrier also represents an important contribution to food safety.

[0034] Liquid biocarrier represents a sustainable strategy for managing resistance to synthetic insecticides, demonstrated in this trial with the partial control efficacy of deltamethrin.

[0035] The results found suggest that the residual control effectiveness of the formulations of the invention could reach at least a period of 6 months. Evaluation of the effectiveness of eleven (11) liquid Biocarrier formulations, indicated as an example, for the control of insect pests of stored grains, in residual studies. Campaign: 2022 / 23

[0036] The objective pursued in these evaluations, given merely as examples Not limiting the scope of the invention is to compare the effectiveness of control over insect pests of stored grains of said eleven (1 1 ) formulations of the invention, (Liquid Biocarrier) in residual treatments. Materials and methods:

[0037] The specimens of Large Woodworm (Tenebrionis mauritanicus), Flat Woodworm (Cryptolestes ferrugineus), Toothed Woodworm (Oryzaephilus surinamensis), Cereal Weevil (Sitophilus granarlas), Rice Weevil (Sitophilus oryzae), Corn Weevil (Sitophilus zeamais), Grain Borer (Rhizopertha dominica) and Cereal Popcorn (Sitotroga cerealella), which were used in the bioassays corresponded to homogeneous cohorts, from different geographical regions of Argentina, raised in glass containers, under controlled temperature (28°C ± 1 °C) and relative humidity (60%) conditions and fed with whole wheat and oat grains.

[0038] The application of the treatments to the wheat grains was carried out in the laboratory using a micro-doser to simulate the treatment the grains receive upon arrival in bulk at the storage facilities. Each experimental unit (EU) consisted of a 2000 cm³ glass flask 3 Each experimental unit (EU) contained a lid and a 154 µm metal mesh sieve, filled with 1000 g of treated cereal grains, according to the assigned treatment. Fifty individuals from the same cohort of adult Tenebrionis mauritanicus, Cryptolestes ferrugineus, and Oryzaephilus surinamensis were introduced into each unit in jars containing wheat. Sitophilus Granaries, Sitophilus oryzae, Sitophilus zeamais, Rhizopertha dominica and Sitotroga cerealella larvae were incorporated into flasks with corn.

[0039] The experimental units (EUs) were assigned to each of the evaluated treatments using a completely randomized design (CRD) with four replicates per treatment. All the grain used during the trial was treated on the same day, so this moment was considered "time zero." Live, affected, and dead insects of each species were counted on 1, 3, and 7 days after the insects were introduced into each experimental unit. At the end of the first month of observation, 50 more insects were introduced. This methodology was continued until the 6-month period was completed.

[0040] Each unit was stored in a rearing chamber under controlled temperature and humidity conditions (28 °C ± 1 °C and 70% RH) and in the absence of light. To determine whether there was a significant effect between the mortality rate and the number of affected individuals in the populations exposed to the different doses of insecticides evaluated, a variance analysis was performed for each month with a significance level of 5% (α = 0.05). A Tukey test for multiple post-hoc comparisons was performed. Mortality values ​​were corrected using Abbott's formula. Abbott's equation: Effectiveness 100 (100) Where: TDK = Infestation in treated plot after applying the treatment. Cd = Infestation in control plot after applying the treatment.

[0041] Treatments evaluated: • Treatment 1: Absolute control • Treatment 2: Commercial chemical control • Treatment 3: 1 ppm of spinosad SC 48% • Treatment 4: 1 ppm of spinosad in liquid Biocarrier • Treatment 5: 1 ppm spinosad + 2 ppm azadirachtin in liquid Biocarrier • Treatment 6 (a): 2 ppm of Bacillus thuringiensis subsp. Tenebrionis* • Treatment 6 (b): 2 ppm of Bacillus thuringiensis subsp. Kurtaki** • Treatment 7 (a): 2 ppm of Bacillus thuringiensis subsp. tenebrionis in liquid Biocarrier* • Treatment 7 (b): 2 ppm of Bacillus thuringiensis subsp. kurtaki in liquid Bio Biocarrier* • Treatment 8 (a): 2 ppm of Bacillus thuringiensis subsp. tenebríonis + 1 ppm spinosad in liquid Biocarríer* • Treatment 8 (a): 2 ppm of Bacillus thuringiensis subsp. kurtaki + 1 ppm spinosad in liquid Biocarríer** *(a) specific formulation for Coleoptera (SC 10%) **(b) specific formulation for Lepidoptera (EC 3.5%) Variables: Grains - Dosage - Formulation - Pests Experimental design: CRD - 4 repetitions Results

[0042] The analysis of cumulative mortality results after 7 days of insect exposure is shown below for each of the re-infestation periods. Graphs summarizing the cumulative mortality results by observation day and reinfestation month are attached as appendices to this report.

[0043] A) Biocarrier Liquid Spinosad 1% formulation. Biocarrier Liquid Spinosad 1% significantly reduced the survival of the eight pest species evaluated compared to the untreated control and the commercial chemical control – deltamethrin (see Table 1-8), in both corn and wheat. Biocarrier Liquid Spinosad 1% achieved effective residual control of the eight pest species evaluated during the six months of the trial. Biocarrier Liquid Spinosad 1% achieved significantly higher control of weevils, woodworms, and lepidoptera than that obtained with the agricultural formulation of Spinosad SC 48%, at an equivalent dose of active ingredient per ton of corn or wheat grain, during the six re-infestations. Average efficacy values ​​between 91.1% and 94.9% were achieved by the sixth month of re-infestation.

[0044] B) Biocarrier liquid formulation Spinosad 1% + Azadirachtin 2%. The liquid biocarrier Spinosad 1% + Azadirachtin 2% significantly reduced the survival of all eight pest species evaluated compared to the untreated control and the commercial chemical control – Deltamethrin (see Table 1-8), in both corn and wheat. The liquid biocarrier Spinosad 1% + Azadirachtin 2% achieved effective residual control of all eight pest species evaluated throughout the trial, with average efficacy values ​​between 93 and 97.5% at the sixth month of re-infestation.

[0045] C) Liquid Biocarrier formulation Bacillus thuringiensis subsp. tenebrionis 2%. Liquid Biocarrier Bt tenebrionis 2% significantly reduced the survival of the seven evaluated beetle species compared to the absolute control and the commercial chemical control - Deltamethrin (see Table 1-8), in both corn and wheat. Liquid Biocarrier Bt tenebrionis 2% achieved effective residual control of the seven evaluated beetle species during the six months of the trial. Liquid Biocarrier Bt tenebrionis 2% achieved significantly higher control of weevils and grain beetles than that obtained by the agricultural formulation of Bt tenebrionis SC 10%, at an equivalent dose of active ingredient per ton of corn or wheat grain, during the six re-infestations performed. Average efficacy values ​​between 84.7% and 93.7% were achieved by the sixth month of re-infestation.

[0046] E) Liquid Biocarrier formulation Bacillus thuringiensis subsp. kurstaki 2%. Liquid Biocarrier Bt Kurstaki 2% significantly reduced the survival of Sitotroga cerealella compared to the absolute control and the commercial chemical control - Deltamethrin (see Table 1-8), in both maize and wheat. Liquid Biocarrier Bt kurstaki 2% achieved effective residual control of Sitotroga cerealella during the six months of the trial. Liquid Biocarrier Bt kurstaki 2% achieved significantly better control of the cereal moth than that obtained by the agricultural formulation of Bt t kurstaki EC 3.5%, at an equivalent dose of active substance per ton of maize or wheat grain, during the six re-infestations carried out. Achieving average efficacy values ​​between 91.4 and 93% by the sixth month of re-infestation.

[0047] F) Biocarrier Liquid formulation Spinosad 1% + Bacillus thuringiensis subsp. tenebrionis 2%. Biocarrier Liquid Spinosad 1% + Bt tenebrionis 2% significantly reduced the survival of the 7 beetle species evaluated, compared to the absolute control and the commercial chemical control - Deltamethrin (see Table 1-8), in both corn and wheat. Biocarrier Liquid Spinosad 1% + Bt tenebrionis 2% achieved effective residual control of the 7 beetle species evaluated throughout the trial, with average efficacy values ​​between 95 and 98.2% at the sixth month of re-infestation.

[0048] G) Biocarrier Liquid formulation Spinosad 1% + Bacillus thuringiensis subsp. Kurstaki 2%. Biocarrier Liquid Spinosad 1% + Bt Kurstaki 2% significantly reduced the survival of Sitotroga cerealella compared to the absolute control and the commercial chemical control - Deltamethrin (see Table 1-8), in both maize and wheat. Biocarrier Liquid Spinosad 1% + Bt Kurstaki 2% achieved effective residual control of Sitotroga cerealella throughout the trial, with average efficacy values ​​between 96.3 and 97.3% at the sixth month of re-infestation.

[0049] Table 1: Cumulative mortality for Sitophilus granarius (Grain Weevil) at 7 days for each month of observation in corn and wheat grains. Month 1 Month 3 month 6 day a:0.05 average a:0.05 average a:0.05 Deltamethrin 10% EC 74.5 B 72.4 B 69.3 C Spinosad SC 48% 68.0 C 71.9 B 63.2 CD Bt tenebrionis SC 10% 63.0 C 68.2 AB 58.6 D Biocarrier Spinosad 100.0 A 98.6 A 93.9 AB Biocarrier Spinosad+ Azadiractin 100.0 A 100.0 A 94.8 AB Biocarrier Bt tenebrionis 98.0 A 93.6 A 91.1 B Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 97.6 A month 1 month 3 month 6 day a:0.05 mean a:0.05 mean a:0.05 Deltamethrin 10% EC 84.5 B 72.4 B 64.8 C Spinosad SC 48% 77.0 C 71.9 B 71.6 BC Bt tenebrionis SC 10% 69.0 C 68.2 B 64.2 C Biocarrier Spinosad 100.0 A 100.0 A 93.8 A Biocarrier Spinosad+ Azadiractin 100.0 A 100.0 A 94.6 A Biocarrier Bt tenebrionis 97.9 A 98.6 A 91.1 A Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 96.7 A Means with a common letter are not significantly different (p > 0.05)

[0050] Table 2: Accumulated mortality for Sitophilus oryzae (Arroz Gorgojo), within 7 days for each month of observation in corn and weed grains. Gorgojo del arroz e mes 1 month 3 month 6 Products dia a:0.05 average a:0.05 average a:0.05 Deltamethrin 10% EC 84.1 B 76.1 C 78.3 B Spinosad SC 48% 81.0 BC 78.6 C 75.3 B Bt tenebrionis SC 10% 75.0 C 77.3 C 69.8 BC Biocarrier Spinosad 100.0 A 99.5 A 93.6 A Biocarrier Spinosad+ Azadirachtin 100.0 A 100.0 A 94.4 A Biocarrier Bt tenebrionis 100.0 A 91.3 AB 93.7 A Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 97.3 A Gorgojo del arroz e mes 1 month 3 month 6 Products day a:0.05 mean a:0.05 mean a:0.05 Deltamethrin 10% EC 84.1 B 78.1 B 67.1 B Spinosad SC 48% 61.0 C 66.3 C 56.7 C Bt tenebrionis SC 10% 55.0 C 69.5 C 51.2 C Biocarrier Spinosad 100.0 A 99.0 A 93.5 A Biocarrier Spinosad+ Azadiractin 100.0 A 99.5 A 95.1 A Biocarrier Bt tenebrionis 100.0 A 82.5 B 93.0 A Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 96.3 A Means with a common letter are not significantly different (p > 0.05)

[0051] Table 3: Cumulative mortality for Sitophilus zeamais (Maize weevil), at 7 days for each month of observation in maize and wheat grains. Deltamethrin 10% EC 82.0 B 71.4 C 76.3 C Spinosad SC 48% 80.0 BC 68.4 C 74.4 C Bt tenebrionis SC 10% 75.0 C 61.3 C 69.8 CD Biocarrier Spinosad 99.0 A 98.1 A 92.1 A Biocarrier Spinosad+ Azadiractin 100.0 A 100.0 A 93.0 A Biocarrier Bt tenebrionis 97.0 A 93.3 AB 90.2 AB Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 95.5 A month 1 month 3 month 6 where ot:O,O5 medium a:0.05 medium ot:O,O5 Deltamethrin 10% EC 72.0 B 74.6 B 53.8 C Spinosad SC 48% 66.4 C 64.7 C 61.8 BC Bt tenebrionis SC 10% 55.0 C 65.3 C 51.2 B Biocarrier Spinosad 98.0 A 99.0 A 94.1 AB Biocarrier Spinosad+ Azadiractin 100.0 A 100.0 A 97.5 A Biocarrier Bt tenebrionis 97.0 A 98.7 A 90.2 AB Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 98.2 A Means with a common letter are not significantly different (p > 0.05)

[0052] Table 4: Cumulative mortality for Carcoma flattened (C. errugineus), at 7 days for each month of observation in maize and wheat grains. month 1 month 3 month 6 day a:0.05 mean a:0.05 mean a:0.05 Deltamethrin 10% EC 76.0 B 72.3 B 70.7 C Spinosad SC 48% 61.0 C 75.5 B 56.7 D Bt tenebrionis SC 10% 59.0 C 64.5 BC 54.9 D Biocarrier Spinosad 100.0 A 99.0 A 93.4 AB Biocarrier Spinosad+ Azadiractin 100.0 A 99.0 A 97.2 A Biocarrier Bt tenebrionis 100.0 A 97.5 A 93.0 AB Biocarrier Spinosad + Bt tenebrionis 100.0 A 98.8 A 98.1 A month month 3 month 6 where ot:O,O5 medium a:0.05 medium ot:O,O5 Deltamethrin 10% EC 77.5 B 68.5 B 66.9 BC Spinosad SC 48% 51.8 C 60.0 B 48.2 C Bt tenebrionis SC 10% 53.6 C 62.3 B 49.8 C Biocarrier Spinosad 96.0 AB 98.0 A 93.3 A Biocarrier Spinosad+ Azadiractin 100.0 A 100.0 A 94.0 A Biocarrier Bt tenebrionis 100.0 A 95.0 A 91.0 A Biocarrier Spinosad + Bt tenebrionis 99.0 A 100.0 A 96.5 A Means with a common letter are not significantly different (p > 0.05)

[0053] Table 5: Cumulative mortality for Carcoma grande (T. mauritanicus), at 7 days for each month of observation in maize and wheat grains. month 1 month 3 month 6 day a:0.05 mean a:0.05 mean a:0.05 Deltamethrin 10% EC 71.4 B 75.6 B 66.4 C Spinosad SC 48% 69.0 B 70.6 B 64.2 C Bt tenebrionis SC 10% 61.0 BC 69.7 B 56.7 CD Biocarrier Spinosad 100.0 A 98.7 A 93.0 AB Biocarrier Spinosad+ Azadiractin 100.0 A 100.0 A 96.0 A Biocarrier Bt tenebrionis 100.0 A 96.8 A 91.0 AB Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 97.0 A month 1 month 3 month 6 day a:0.05 mean a:0.05 mean a:0.05 Deltamethrin 10% EC 78.3 C 78.6 B 66.3 C Spinosad SC 48% 72.9 C 69.7 C 67.8 C Bt tenebrionis SC 10% 55.8 D 67.5 C 51.9 D Biocarrier Spinosad 100.0 A 100.0 A 94.9 A Biocarrier Spinosad+ Azadiractin 100.0 A 100.0 A 95.3 A Biocarrier Bt tenebrionis 97.0 A 100.0 A 91.4 AB Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 97.2 A Means with a common letter are not significantly different (p > 0.05)

[0054] Table 6: Cumulative mortality in Toothed Carcoma (O. surinamensis), at 7 days for each month of observation in maize and wheat grains. month 1 month 3 month 6 day a:0.05 mean a:0.05 mean a:0.05 Deltamethrin 10% EC 67.9 B 73.9 B 71.7 B Spinosad SC 48% 77.0 C 70.1 B 70.6 B Bt tenebrionis SC 10% 69.0 D 66.2 BC 64.2 BC Biocarrier Spinosad 100.0 A 98.9 A 93.3 A Biocarrier Spinosad+ Azadiractin 100.0 A 100.0 A 94.2 A Biocarrier Bt tenebrionis 100.0 A 97.0 A 93.0 A Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 95.6 A month 1 month 3 month 6 day a:0.05 mean a:0.05 mean a:0.05 Deltamethrin 10% EC 77.9 C 67.6 B 72.4 C Spinosad SC 48% 65.7 D 71.6 B 61.1 D Bt tenebrionis SC 10% 57.1 D 73.3 B 53.1 E Biocarrier Spinosad 94.2 AB 100.0 A 93.4 AB Biocarrier Spinosad+ Azadiractin 98.4 A 100.0 A 95.0 A Biocarrier Bt tenebrionis 95.1 A 100.0 A 90.5 AB Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 98.0 A Means with a common letter are not significantly different (p > 0.05)

[0055] Table 7: Cumulative mortality in cereal popcorn (S. cerealella), at 7 days for each month of observation in maize and wheat grains. Deltamethrin 10% EC 77.1 B 71.5 B 70.3 B Spinosad SC 48% 64.5 C 75.7 B 60.0 AB Bt tenebrionis SC 10% 62.8 0 71.5 B 58.4 A Biocarrier Spinosad 100.0 A 98.7 A 93.0 A Biocarrier Spinosad+ Azadiractin 100.0 A 99.2 A 95.1 AB Biocarrier Bt tenebrionis 100.0 A 94.8 AB 93.0 A Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 96.3 A month 1 month 3 month 6 there is to: there is to: there is a:0.05 Deltamethrin 10% EC 77.1 C 67.2 BC 71.7 B Spinosad SC 48% 65.7 D 70.5 BC 61.1 C Bt tenebrionis SC 10% 53.4 D 73.2 BC 49.7 D Biocarrier Spinosad 96.4 A 100.0 A 92.7 A Biocarrier Spinosad+ Azadiractin 99.2 A 100.0 A 94.3 A Biocarrier Bt tenebrionis 92.9 B 100.0 A 91.4 A Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 97.3 A Means with a common letter are not significantly different (p > 0.05)

[0056] Table 8: Cumulative mortality Cereal drill (R dominica), at 7 days for each month of observation in maize and wheat grains. month 1 month 3 month 6 there is a:0.05 there is a:0.05 there is a:0.05 Deltamethrin 10% EC 78.3 B 64.5 B 61.4 C Spinosad SC 48% 63.1 C 69.0 B 58.7 C Bt tenebrionis SC 10% 58.3 C 66.9 B 54.2 CD Biocarrier Spinosad 98.0 A 96.8 A 91.1 AB Biocarrier Spinosad+ Azadiractin 100.0 A 100.0 A 93.0 A Biocarrier Bt tenebrionis 96.0 A 96.1 A 89.3 AB Biocarrier Spinosad + Bt tenebrionis 100.0 A 100.0 A 95.0 A month 1 month 3 months 6 there is to: there is to: there is a:0.05 Deltamethrin 10% EC 78.3 B 67.0 B 71.4 C Spinosad SC 48% 61.2 C 68.6 B 56.9 D Bt tenebrionis SC 10% 57.4 C 70.3 B 53.4 D Biocarrier Spinosad 95.6 A 100.0 A 91.9 AB Biocarrier Spinosad+ Azadiractin 97.1 A 100.0 A 94.9 A Biocarrier Bt tenebrionis 91.0 AB 100.0 A 84.7 B Biocarrier Spinosad + Bt tenebrionis 99.8 A 100.0 A 97.6 A Means with a common letter are not significantly different (p > 0.05) ANNEXES OF SUPPLEMENTARY INFORMATION

[0057] Figure 1 : Cumulative mortality for Sitophilus granarias (Cereal Weevil), by time of observation, in the first artificial infestation. Corn

[0058] Figure 2: Cumulative mortality for Sitophilus granaria (Cereal Weevil), by observation time, in the third artificial infestation. Maize

[0059] Figure 3: Cumulative mortality for Sitophilus granaria (Cereal Weevil), by observation time, in the sixth artificial infestation. Maize

[0060] Figure 4: Cumulative mortality for Sitophilus granaria (Cereal Weevil), by observation time, in the first artificial infestation. Wheat

[0061] Figure 5: Cumulative mortality for Sitophilus granaria (Cereal Weevil), by observation time, in the third artificial infestation. Wheat

[0062] Figure 6: Cumulative mortality for Sitophilus granaria (Cereal Weevil), by observation time, in the sixth artificial infestation. Wheat

[0063] Figure 7: Cumulative mortality for Sitophilus oryzae (Rice weevil), by observation time, in the first artificial infestation. Maize

[0064] Figure 8: Cumulative mortality for Sitophilus oryzae (Rice weevil), by observation time, in the third artificial infestation. Maize

[0065] Figure 9: Cumulative mortality for Sitophilus oryzae (Rice weevil), by observation time, in the sixth artificial infestation. Maize

[0066] Figure 10: Cumulative mortality for Sitophilus oryzae (Rice weevil), by observation time, in the first artificial infestation. Wheat

[0067] Figure 11: Cumulative mortality for Sitophilus oryzae (Weevil of the

[0068] Figure 12: Cumulative mortality for Sitophilus oryzae (Weevil of the

[0069] Figure 13: Cumulative mortality for Sitophilus zeamais (Corn weevil), by observation time, in the first artificial infestation. Corn

[0070] Figure 14: Cumulative mortality for Sitophilus zeamais (Corn weevil), by observation time, in the third artificial infestation. Corn

[0071] Figure 15: Cumulative mortality for Sitophilus zeamais (Cellworm)

[0072] Figure 16: Cumulative mortality for Sitophilus zeamais (Corn weevil), by observation time, in the first artificial infestation. Wheat

[0073] Figure 17: Cumulative mortality for Sitophilus zeamais (Corn weevil), by observation time, in the third artificial infestation. Wheat

[0074] Figure 18: Cumulative mortality for Sitophilus zeamais (Corn weevil), by observation time, in the sixth artificial infestation. Wheat

[0075] Figure 19: Cumulative mortality of C. ferrugineus (Flatcorn Beetle), by observation time, in the first artificial infestation. Maize

[0076] Figure 20: Cumulative mortality of C. ferrugineus (Flatcorn Beetle), by observation time, in the third artificial infestation. Maize 5

[0078] Figure 21: Cumulative mortality Cryptolestes ferrugineus (Woodworm)

[0079] Figure 22: Cumulative mortality of C. ferrugineus (Flatcorn Beetle), by observation time, in the first artificial infestation. Wheat

[0080] Figure 23: Cumulative mortality of C. ferrugineus (Flatcorn Beetle), by observation time, in the third artificial infestation. Wheat

[0081] Figure 24: Cumulative mortality of Cryptolestes ferrugineus (Flatcorn Beetle), by observation time, in the sixth artificial infestation. Wheat

[0082] Figure 25: Cumulative mortality for T. mauritanicus (Large Corn Earworm), by observation time, in the first artificial infestation. Maize

[0083] Figure 26: Cumulative mortality for T. mauritanicus (Large Corn Earworm), by observation time, in the third artificial infestation. Maize

[0084] Figure 27: Cumulative mortality for T. mauritanicus (Large Corn Earworm), by observation time, in the sixth artificial infestation. Maize

[0085] Figure 28: Cumulative mortality for T. mauritanicus (Large Wheat Earworm), by observation time, in the first artificial infestation. Wheat

[0086] Figure 29: Cumulative mortality for T. mauritanicus (Large Wheat Earworm), by observation time, in the third artificial infestation. Wheat

[0087] Figure 30: Cumulative mortality for Tenebrionis mauritanicus (Large woodworm), by observation time, in the sixth artificial infestation. Wheat

[0088] Figure 31: Cumulative mortality for O. surinamensis L. (Woodworm)

[0089] Figure 32: Cumulative mortality for O. surinamensis L. (Toothed corn earworm), by observation time, in the third artificial infestation. Maize 5

[0090] Figure 33: Cumulative mortality for O. surinamensis L. (Woodworm)

[0091] Figure 34: Cumulative mortality for O. surinamensis L. (Woodworm) 5

[0100] Figure 35: Cumulative mortality for O. surinamensis L. (Toothed grain beetle), by observation time, in the third artificial infestation. Wheat

[0101] Figure 36: Cumulative mortality for O. surinamensis L. (Toothed grain beetle), by observation time, in the sixth artificial infestation. Wheat

[0102] Figure 37: Cumulative mortality of S. cerealella (Grain Moth), by observation time, first artificial infestation. Corn

[0103] Figure 39: Cumulative mortality for S. cerealella (Grain Moth), by observation time, third artificial infestation. Corn

[0104] Figure 42: Cumulative mortality of S. cerealella (Grain Moth), by observation time, sixth artificial infestation. Corn

[0105] Figure 37: Cumulative mortality of S. cerealella (Grain Moth), by observation time, first artificial infestation. Wheat

[0106] Figure 39: Cumulative mortality of S. cerealella (Grain Moth), by observation time, in the third artificial infestation. Wheat

[0107] Figure 42: Cumulative mortality of S. cerealella (Grain Moth) by observation time, sixth artificial infestation. Wheat

[0108] Figure 43: Cumulative mortality of Rhizopertha dominica (Grain borer), by observation time, in the first artificial infestation. Maize

[0109] Figure 45: Cumulative mortality of Rhizopertha dominica (Grain borer), by observation time, in the third artificial infestation. Maize

[0110] Figure 48: Cumulative mortality of Rhizopertha dominica (Corn borer), by observation time, in the sixth artificial infestation. Maize

[0111] Figure 43: Cumulative mortality of Rhizopertha dominica (Grain borer), by observation time, in the first artificial infestation. Wheat

[0112] Figure 45: Cumulative mortality of Rhizopertha dominica (Grain borer), by observation time, in the third artificial infestation. Wheat

[0113] Figure 48: Cumulative mortality of Rhizopertha dominica (Grain borer), by observation time, in the sixth artificial infestation. Wheat DIFFERENCES WITH SOLID BIOCARRIER: • Given the spray application method, Biocarher Liquid allows for better distribution and greater uniformity of the treatment. • Liquid Biocarrier does not generate foreign matter in the treated merchandise. • Biocarrier Liquid refers to both a ready-to-use product and a concentrated product that requires dilution before application. In the latter case, the logistics associated with its marketing are considerably simplified. Note: The tests performed with Biocarrier Liquid were carried out with an active ingredient concentration in the range of 0.2 to 3%. This concentration may correspond to the formulation as a ready-to-use product or may be the result of dilution in its concentrated form. Types of formulations / commercial presentations of liquid Biocarrier: • LPU Liquid Bait (Ready to Use) • Concentrated bait • Dispersible concentrate • Oil-in-water emulsion • Water-in-oil emulsion • Concentrated gel or paste • Contact liquid or gel • Miscible liquid • Microemulsion • Microencapsulated suspension • Spreading-oil • Concentrated suspension • Ultra-low volume suspension • Suspo-emulsion formulation, causing death by ingestion.

Claims

1. CLAIMS 1- NEW TECHNOLOGY AND FORMULATION TYPE FOR BIOLOGICAL CONTROL OF INSECT PESTS OF GRAIN STORAGE, the present invention patent has as its main objective the presentation of a new technology, in liquid formulation, developed exclusively for biological control of insect pests of grain storage, characterized by incorporating one or more food attractants and 0.2% to 3% of at least one biological active principle. 2- NEW TECHNOLOGY AND FORMULATION TYPE FOR BIOLOGICAL CONTROL OF INSECT PESTS OF GRAIN STORAGE, according to what is claimed in 1, characterized in that the active ingredient is chosen from at least one of the compounds with the following characteristics or mixtures thereof: • Bacillus thuringiensis, its subspecies and the insecticidal proteins they produce. • Allosteric modulators of the nicotinic acetylcholine receptor - Spinosins • Host-specific occluded pathogenic viruses. Granuloviruses (GVs), Nucleopolyhedroviruses (NPVs). • Plant extracts and crude oils • Azadirachtins Entomopathogenic fungi • Peptides derived from natural toxins, whether of animal or plant origin, or derived from molecular biology techniques. 3- NEW TECHNOLOGY AND FORMULATION TYPE FOR BIOLOGICAL CONTROL OF INSECT PESTS OF GRAIN STORAGE, according to what is claimed in 1, characterized in that the bait is chosen for consumption over cereals and oilseeds, in its formulation it contains two or more of the following components, in order to meet the objectives of attraction and preferential consumption over the mass of grains: • crude protein; • fatty acids; • glucose; sucrose • p-methoxyethanol • Pectins

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