METHOD FOR CALCULATING MYCOTOXIN RISK IN POULTRY AND SWINE USING CONTAMINATION DATA AND RISK FACTORS AND ADJUSTMENTS

Description

METHOD FOR CALCULATING MYCOTOXIN RISK IN POULTRY AND SWINE USING CONTAMINATION DATA AND RISK FACTORS AND ADJUSTMENTS. FIELD OF THE INVENTION

[001] The invention shown in this patent application demonstrates a means of calculating and verifying the risk of mycotoxins in poultry and / or swine, providing information to livestock farmers for risk monitoring and mitigation. The method is carried out by creating an evaluation period for feed and feed raw materials for said poultry and / or swine, collecting data on mycotoxin contamination, and then calculating a risk index for contamination by these mycotoxins using this raw contamination data and its prevalence, in addition to using an epidemiological factor and risk adjustment according to the species, age and animal category analyzed, and to create a standardized numerical base among all the mycotoxins analyzed.The method shown here is specifically used for the analysis and calculation of mycotoxin contamination risk in feed and feed raw materials for different animal species, and more specifically for said risk calculation for poultry and / or swine populations. FUNDAMENTALS OF THE INVENTION

[002] In the specific area of ​​livestock farming, mycotoxins, toxic substances produced by fungi, cause significant damage to the digestive tract, immune and reproductive systems of poultry and / or swine. This presents a challenge for livestock farmers to minimize losses related to the zootechnical performance of these animals and, consequently, financial losses for producers. Prolonged exposure, even to low levels of mycotoxins, can cause chronic intestinal inflammation and immunosuppression, increasing susceptibility to secondary infections, such as E. coli and Salmonella, in these animals. This justifies amplifying the impact of prevalence in risk calculation, as constant contamination, even at low concentrations, can have significant cumulative harmful effects on these animals.

[003] Therefore, veterinary epidemiological models often use parameters adjusted to reflect the long-term and cumulative impact of exposures in birds and / or Petition 870250005593, dated 01 / 23 / 2025, page 5 / 22 2 / 9 swine. The literature on epidemiological modeling, as described by Chubb and Jacobsen, highlights that mathematical models can be used to identify sensitive variables and predict health outcomes in animal populations. Sensitivity analysis is crucial to determine how different components of the model influence the final result. Parameters with high sensitivity, such as prevalence, must be measured accurately and may justify the use of higher powers to reflect their disproportionate impact.

[004] Regarding behavioral models in epidemiology, a study on behavioral epidemic models highlights the importance of adjusting parameters to reflect the population's behavioral response to perceived risk. In models where risk perception alters behaviors (e.g., increased adherence to non-pharmacological interventions), adjustments are made to represent how these changes affect disease dynamics. Analogously, in the context of mycotoxins, prolonged prevalence and continuous risk perception justify the use of higher potencies to capture the cumulative impact of exposure.

[005] As for the impact of mycotoxins on the health of poultry and swine, several studies show that continuous exposure and high prevalence of mycotoxins, such as aflatoxins and fumonisins, have severe and cumulative effects on the health of said poultry and swine. These effects include alterations in weight gain, feed conversion, and biochemical parameters, as well as histopathological damage to vital organs such as the liver, intestines, and kidneys.

[006] Thus, this invention application aims to show a method for calculating and verifying the risk of mycotoxins in poultry and / or swine, in order to allow individuals interested in such data, such as livestock farmers who specifically raise such poultry and / or swine, to have a simple and effective way of analyzing the risk so that they can, if necessary, take the appropriate health measures given the data obtained regarding mycotoxin contamination. STATE OF THE ART

[007] Some of the most relevant prior art documents were cited in order to outline the prior art relevant to the invention. One of these Petition 870250005593, dated 01 / 23 / 2025, p. 6 / 22 3 / 9 documents is patent US2022074900, entitled “Monitoring mycotoxins and its metabolites in the blood of pigs or broiler chickens” and published on 10 / 03 / 2022, with the abstract reproduced below: “The present invention relates to a multiple screening method for the detection of a large number of mycotoxins and metabolites in broiler chickens and pigs, the method comprising collecting blood from broiler chickens and pigs as a dried blood sample, preparing the dried blood sample for analysis and analyzing the prepared dried blood sample by a two-stage liquid chromatography-tandem mass spectrometry (LC-MS / MS) process. In a first LC-MS / MS step, for a given mobile phase of the LC process, the mass spectrometer operates in negative electrospray ionization mode and, for another mobile phase of the LC process, the mass spectrometer operates in ionization mode by positive electrospray.This method can be used advantageously for screening and evaluating the exposure of pigs or broilers to feed contaminated with mycotoxins. Furthermore, this method can be used to assess the impact of adding mycotoxin detoxifying agents to animal feed.

[008] Another document found is patent KR102152310, entitled “Multi-residue and multi-class analytical method for agricultural chemical veterinary drug and mycotoxin in feed” and published on 04 / 09 / 2020, with the following summary: “The present invention relates to a method for the simultaneous analysis of pesticide residues in feed, veterinary drugs and mycotoxins and provides a method for the simultaneous analysis of each feed sample by methods such as GC-MS / MS and LC-MS / MS with only two pretreatments using the polarity difference of each remaining material in the feed. Through the present invention, it is possible to construct an effective analytical method that can save time, cost, etc. to monitor harmful substances in feed, providing an easier analytical method compared to existing technology that uses about 40 methods.”

[009] Another document found is patent US2011306508, entitled “Method of Mycotoxin Detection” and published on 12 / 15 / 2011, with the following summary: “The presence of mycotoxins in agricultural products requires large-scale testing of a wide variety of sample materials to ensure food safety and Petition 870250005593, dated 01 / 23 / 2025, p. 7 / 22 4 / 9 rations. The mycotoxin ochratoxin A represents a breakthrough for all mycotoxins, as the level of sensitivity required for regulatory requirements for this compound at the parts per billion level is as low as or lower than any other mycotoxin. This invention describes the identification of a set of DNA ligands with sufficiently high binding affinity and specificity for ochratoxin A to allow an improvement over existing methods for the separation, concentration, and quantitative determination of ochratoxin A in sample material. OBJECTIVES AND ADVANTAGES OF THE INVENTION

[010] The invention shown in this patent application displays the following specific advantages when compared with the prior art: • The possibility of using different risk assessment periods in the calculation shown in the method allows for the evaluation of this risk in both the short and long term, both of which are important for livestock farmers in different breeding operations or time periods to be analyzed; • The use of the epidemiological factor and risk adjustment for each species, age, and category of poultry or swine allows the calculation to be adapted for different livestock farmers, thus possessing a flexibility not anticipated in the state of the art; and • In normal analyses, verification of information by specialists is necessary since different mycotoxins have different concentrations in samples, while in the invention of this patent application, the risk index adjusts to a normalized numerical base among all the mycotoxins analyzed, allowing easy verification of the risk of mycotoxin contamination even by a normal livestock farmer.

[011] The invention thus demonstrates usability that is not demonstrated in the state of the art, bringing advantages of use to a wide range of livestock farmers who raise poultry and swine for their proper breeding and supply to consumers of the products obtained from these animals, which is something not demonstrated in the state of the art. GENERAL DESCRIPTION OF THE INVENTION Petition 870250005593, dated 01 / 23 / 2025, page 8 / 22 5 / 9

[012] The innovative invention demonstrated in this patent application shows a method for evaluating and analyzing the risk of mycotoxins present in feed and feed raw materials for production animals, and more specifically in poultry and swine farming, said risk detection method being adaptable for livestock farmers of different scales for making managerial decisions in their respective businesses.

[013] The method demonstrated here establishes a period for assessing the risk of animals to mycotoxins, collecting data on contamination of said mycotoxins in feed and feed raw materials for relevant and analyzed animals, and then performing the appropriate mycotoxin risk calculation using the contamination data obtained and factors analyzed by the livestock farmer regarding contamination prevalence, in addition to an epidemiological factor and a risk adjustment, thus adapting this calculation to various possible poultry and swine farming environments, including, but not limited to, breeders, replacement animals, egg-producing birds for consumption and animals destined for slaughter, given the high adaptability of this method.

[014] This method, therefore, is suitable for a wide range of livestock farmers in their calculations for assessing the risk of mycotoxins in poultry and swine, which is something that is not anticipated in the prior art. DESCRIPTION OF THE FIGURES

[015] The invention will be described in detail below, and for better understanding, reference will be made to the attached drawings, in which are represented: Figure 1: Flowchart showing all the main and relevant steps for executing the method. Figure 2: Graphs showing concentrations in micrograms per kilogram, or parts per billion, of different mycotoxins in different samples; Figure 3: Graphs showing adjusted risk indices obtained after analysis of different mycotoxins as shown in this method, and showing different toxins with risk indices having the same normalized numerical basis. DETAILED DESCRIPTION OF THE INVENTION Petition 870250005593, dated 01 / 23 / 2025, page 9 / 22 6 / 9

[016] The METHOD FOR CALCULATING MYCOTOXIN RISK IN POULTRY AND SWINE USING CONTAMINATION DATA AND RISK FACTORS AND ADJUSTMENTS shows a method that collects data on mycotoxin contamination, i.e., various toxic substances produced by different species of fungi, in feed and feed raw materials for production animals, and more specifically, poultry and swine used for human consumption of their meat or secondary products, such as eggs or milk. Risk is a numerical indicator, or index, that demonstrates the intensity of the risk related to the mycotoxin challenge for animals in the herd of a given company, producer or region.

[017] This invention shown here is implemented by means of a method, which is carried out and demonstrated in a series of sequential steps, said steps being demonstrated below in this detailed description.

[018] In an initial Step 1 (1), a data collection time period is established for contamination risk assessment, such as shift, daily, weekly, bi-weekly, or monthly, the number of samples collected in this selected time period, and also which mycotoxins will be analyzed and monitored, these chosen according to the needs of the user or livestock farmer using this method demonstrated here. In an implementation of the method, for example, in a weekly period with fourteen samples, two sample collections are thus carried out each day. This choice of time period and sample quantity is up to the user of this method and depends on their livestock situation, thus adapting to various livestock farmers.Both the choice of time period and the choice of the number of samples are already made by companies and producers, who only provide information about the collection of inputs for the analysis and the calculations themselves. The choice of the method of sample collection, such as whether it is done manually or automatically, is also up to the user of the method according to their needs. We note that the shorter the time period chosen, the greater the accuracy of the risk calculation performed, this also depending on the number of samples analyzed in this period. Petition 870250005593, dated 01 / 23 / 2025, page 10 / 22 7 / 9

[019] In a Step 2 (2), samples are collected for analysis of each selected mycotoxin during the chosen time period, and then the analysis is performed. This collection method uses conventional methods for collecting mycotoxin data existing in the state of the art, such as, for example, through the use of mass spectrometry, immunoenzymatic assays, infrared or any other method available on the market for mycotoxin analysis in feed and feed raw materials for production animals. Some specific methods for collecting samples in the periods specified in Step 1 (1) and the analysis of the mycotoxins themselves are demonstrated in the articles “Comparison of the Efficiency between Two Sampling Plans for Aflatoxins Analysis in Maize” [1] and “Two Sampling Plans for Fumonisin Analysis in Maize” [2],

[020] In Step 3 (3), the average mycotoxin contamination in micrograms per kilogram, i.e., pg / kg, also known as parts per billion, is calculated for each of the collected samples. The prevalence is also calculated, meaning a percentage of positive samples in the selected time period. These contamination and prevalence calculations are performed for each selected mycotoxin to be analyzed. The prevalence calculation is then performed to obtain the prevalence percentage: Prevalence = positive samples / total samples x 100

[021] In Step 4 (4), an epidemiological factor, or FE, between 1.1 and 1.9 is chosen, which is to be applied as a calculation factor in the prevalence calculated in Step 3 (3), and this factor varies according to different use cases. The choice of the epidemiological factor is made by a specialist in the subject of mycotoxins in production animals according to the needs of a specific livestock situation, as it involves knowledge of the mechanisms of action of these toxins in the animal organism, as well as the impact of continuous or punctual exposures.

[022] In Step 5 (5), a mycotoxin risk index calculation is performed, which is then applied to each mycotoxin analyzed by the method, with the main embodiment being made using the following equation: RM = Contamination x PrevalenceFE Petition 870250005593, dated 01 / 23 / 2025, page 11 / 22 8 / 9

[023] In Step 6 (6), the risk index adjustment calculation is performed according to the species, age and animal category with a specific factor, or FA, this having a value between 10° and 10-10, that is, from 1 to 0.0000000001, this factor chosen by the method user according to their needs given their specific livestock situation and also in such a way that all adjusted risk indices have a normalized numerical basis among all mycotoxins analyzed. Examples of this normalized numerical basis can be seen more clearly in Figure 3 of the patent application, which shows graphs of mycotoxin risk indices and with all these graphs having a single index ranging from 0 to 300 for various mycotoxins, independent of the mycotoxin itself.Figure 2 shows standard graphs for verifying mycotoxin contamination, which is normal in the state of the art. As we can see on the y-axis of parts per billion, different mycotoxins can have extremely different concentrations, with concentrations ranging from 0 to 35 in the case of aflatoxins and from 0 to 12000 in the case of zearalenone. Thus, the analysis of data such as those seen in the graphs in Figure 2 requires a specialist to interpret the data obtained on mycotoxin contamination and concentration, given that a concentration in parts per billion may be normal for certain mycotoxins and extremely harmful for others.Thus, the normalized numerical base provided by the use of the specific factor shows a normalized index, as seen in Figure 3, which gives the livestock farmer information that is much easier to interpret than the mere concentration in parts per billion itself, something not anticipated by the state of the art.

[024] In primary realization, the Species, Age and Category Adjusted Mycotoxin Risk, or MRA, is calculated as follows: RMA = RM x FA

[025] Thus, the method shown in this patent application demonstrates an efficient and robust method for creating a mycotoxin risk index, applicable to different breeding situations of different livestock farmers according to their specific needs, which is something that is not demonstrated in the state of the art. Petition 870250005593, dated 01 / 23 / 2025, page 12 / 22 9 / 9

[026] This index thus reflects the reality of different breeding sites or fields, being used to indicate the trend of mycotoxin evolution, and the normalized index is easy to understand even when referring to several different mycotoxins, thus enabling a basis for decision-making, such as the implementation of specific mitigation measures for different mycotoxins. BIBLIOGRAPHIC REFERENCES [1] Mallmann, Adriano Olnei, Alexandro Marchioro, Maurício Schneider Oliveira, Ricardo Hummes Rauber, Paulo Dilkin, and Carlos Augusto Mallmann. 2014. “Comparison of the Efficiency between Two Sampling Plans for Aflatoxins Analysis in Maize”. Brazilian Journal of Microbiology. FapUNIFESP (SciELO). https: / / doi.org / 10.1590 / s151783822014000100006. [2] Mallmann, Adriano Olnei, Alexandro Marchioro, Mauricio Schneider Oliveira, Luciane Minetto, Liziane Rachel da Silva Wovst, Ricardo Hummes Rauber, Paulo Dilkin, and Carlos Augusto Mallmann. 2013. “Two Sampling Plans for Fumonisin Analysis in Corn”. Ciência Rural. FapUNIFESP (SciELO). https: / / doi.org / 10.1590 / s010384782013000300029.

Claims

CLAIMS 1) METHOD FOR CALCULATING MYCOTOXIN RISK IN POULTRY AND SWINE USING CONTAMINATION DATA AND RISK FACTORS AND ADJUSTMENTS, which shows a method that performs a mycotoxin risk index calculation, this method characterized by being carried out in a series of sequential steps, namely: • In Step 1 (1), a data collection time period is established for assessing the risk of contamination, the number of samples collected in this selected time period, and the selection of which mycotoxins will be analyzed, these chosen according to the needs of the method user; • In Step 2 (2), samples of each selected mycotoxin are collected during the chosen time period for analysis;• In Step 3 (3), the average mycotoxin contamination in micrograms per kilogram of each of the collected samples is calculated, and the prevalence as a percentage of positive samples in the selected time period, these calculations performed for each selected mycotoxin; • In Step 4 (4), an epidemiological factor is chosen, this to be applied as a calculation factor in the prevalence calculated in Step 3 (3) and chosen according to a specialist in the subject of mycotoxins in production animals according to the needs of a specific livestock situation; • In Step 5 (5), a mycotoxin risk index calculation is performed, said calculation to be applied to each mycotoxin analyzed;• In Step 6 (6), the risk index adjustment calculation is performed according to the species, age and animal category with a specific factor, this having a value between 10° and 10-10, this factor chosen by the method user according to their specific needs and in such a way that all adjusted risk indices have a normalized numerical basis among all mycotoxins analyzed.; 2) METHOD FOR CALCULATING MYCOTOXIN RISK IN POULTRY AND SWINE USING CONTAMINATION DATA AND RISK FACTORS AND ADJUSTMENTS, according to claim 1, characterized by, in Step 1 (1), using a collection time period chosen from a set of shift periods, daily, weekly, bi-weekly or monthly. 3) METHOD FOR CALCULATING MYCOTOXIN RISK IN POULTRY AND SWINE USING CONTAMINATION DATA AND RISK FACTORS AND ADJUSTMENTS, according to claim 1, characterized in that, in Step 5 (5), the epidemiological factor has a value between 1.1 and 1.9, and the mycotoxin risk calculation is performed using the following method, where FE is the epidemiological factor chosen in Step 4 (4): RM = Contamination x PrevalenceFE. 4) METHOD FOR CALCULATING MYCOTOXIN RISK IN POULTRY AND SWINE USING CONTAMINATION DATA AND RISK FACTORS AND ADJUSTMENTS, according to claim 3, characterized in that, in Step 6 (6), the risk calculation is performed as follows, where FA is the specific factor chosen in this Step 6 (6): RMA = RM x FA.

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