WO2021087890A1 - Method for building personalized probiotics database, and use thereof in screening of probiotics - Google Patents

Abstract

A method for building a personalized probiotics database, and the use thereof in the screening of probiotics. The method comprises co-culturing probiotics and a biological sample, then analyzing a secretion volume of IL-10, a secretion volume ratio of IL-4/IFN-γ, and a secretion volume ratio of IFN-γ/IL-4, and ranking, according to numerical values from high to low, the corresponding probiotics for screening.

Description

The establishment method of individualized probiotics database and its application in the screening of probiotics Technical field

The invention relates to a method for establishing an individualized probiotic database and its application in the screening of probiotics. In particular, the aforementioned individualized probiotics database contains at least two or more probiotics screening indicators, so as to perform probiotics screening that meets individualized differences.

Background technique

The current health industry is a health care product designed for the general public. Because of ignoring individual differences, its health care effect is very limited. The concept of "precision medicine" is to formulate prevention and auxiliary strategies for individual differences in physique. The concept is applied to the development of health care products, and product design can be made due to individual differences to increase the effectiveness of health care products.

In summary, in the current health industry related to probiotics, how to develop and screen individual differentiated probiotics on a scientifically based platform is still a topic that urgently needs breakthroughs and developments based on individual conditions.

Summary of the invention

In view of the above-mentioned background of the invention, in order to meet the requirements of the industry, the present invention provides a method for establishing an individualized probiotic database and its application in the screening of probiotics, thereby achieving the purpose of accurate matching of probiotics to suitable individuals.

Explanations of some commonly used terms in the contents of the present invention are as follows.

The individual described in the content of the present invention includes all mammals such as humans, pets, livestock, or other organisms that can be used with probiotics to regulate their physiological functions.

The screening of probiotic strains or different strains of the same species as described in the content of the present invention and their applications belong to the scope of individual health care, and do not involve disease treatment and diagnosis behaviors.

In the content of the present invention, IL-4 is interleukin 4; IL-10 is interleukin 10; IFN-γ is gamma interferon.

The Th1 helper cells described in the content of the present invention are type 1 helper T cells (T helper 1 cells); Th2 helper cells are type 2 helper T cells (T helper 2 cells).

The sorting method described in the summary of the present invention is to arrange the numerical values in order of priority from high to low, with the highest numerical value representing the first rank, the second highest numerical value representing the second rank, and again representing the third rank.

The first objective of the present invention is to provide a method for establishing an individualized probiotic database. The establishment method includes but is not limited to the five steps described below.

Step 1: Provide multiple probiotic strains or different strains of the same species; the multiple probiotic strains refer to species belonging to different genus, different strains of the same genus or different strains of the same strain, and their content ratio Bifidobacterium bifidum, Bifidobacterium longum, Bifidobacterium infantis, Lactobacillus gasseri, Lactobacillus lactis, Lactococcus lactis, Bacillus coagulans, Lactobacillus delbrueckii subsp.bulgaricus, Lactobacillus johnsonii, Lactobacillus casei, Lactobacillus casei, Lactobacillus casei Lactobacillus paracasei), Bifidobacterium adolescentis, Lactobacillus salivarius, Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus brevis, Bifidobacterium brevis ( Bifidobacterium breve, Lactobacillus acidophilus, Streptococcus thermophilus, Lactobacillus helveticus, Clostridium butyricum, Bifidobacterium lactis, Lactobacillus rhamnosus (Lactobacillus rhamnosus), Lactobacillus reuteri (Lactobacillus reuteri), Streptococcus thermophilus (Faecalibacterium prausnitzii) or Akkermansia (Muciniphila).

Step 2: The multiple probiotic strains or different strains of the same species are separately co-cultured with biological samples derived from the same individual in vitro. The biological sample contains immune cells.

Step 3: Perform an analysis step to obtain cytokine information secreted by biological samples co-cultured with the multiple probiotic strains or the same different strains, the cytokine information including the secretion of IL-4 The ratio of the secretion of IFN-γ (IL-4/IFN-γ), the ratio of the secretion of IFN-γ and the secretion of IL-4 (IFN-γ/IL-4) and the secretion of IL-10 .

Step 4: Perform a sorting step, which is based on the value of the cytokine information to create sorting information of the value from high to low, and the sorting information includes the multiple probiotics corresponding to IL-4/IFN-γ. The ranking of bacteria, the ranking of the plurality of probiotics corresponding to IFN-γ/IL-4, the ranking of the plurality of probiotics corresponding to the secretion of IL-10, or any combination of the above rankings.

Step 5: Perform a pairing step to complete the establishment of the individualized probiotic database; the pairing step includes making the IL-4/IFN-γ and the corresponding probiotic strains or the same species respectively The ranked first three probiotics of different strains and the secretion amount of the IL-10 and the corresponding multiple probiotic strains or the first three ranked probiotics of the same species of different strains are paired to a Th1 Individuals with overexpression of helper cells; and the IFN-γ/IL-4 and the corresponding probiotic strains or the top three probiotics of the same species and different strains and the secretion of the IL-10 The amount and the corresponding probiotic strains of the plurality of probiotic strains or the top three ranked probiotics of the same species and different strains are matched to an individual with overexpression of Th2 helper cells.

Innovatively, the personalized probiotic database established according to this method contains the aforementioned cytokine information (IL-10 secretion amount, IL-4/IFN-γ and IFN-γ/IL-4) personalized probiotics established The selection index platform contains at least two or more probiotic strains or selection indexes of different strains of the same species, so that it can screen probiotics that conform to individualization or individual immune differentiation.

Specifically, the probiotic strains or the screening indexes and information of different strains of the same species provided by the selection index platform for individualized probiotics include the sorting, ranking, and information of the multiple probiotics corresponding to IL-4/IFN-γ. The ranking of the plurality of probiotics corresponding to IFN-γ/IL-4, the ranking of the plurality of probiotics corresponding to the secretion of IL-10, and the pairing information of the individualized probiotics.

Specifically, the pairing information of the personalized probiotics includes a probiotic aptitude table for individuals with overexpression of Th1 helper cells or a probiotic aptitude table for individuals with overexpression of Th2 helper cells.

Among them, the above-mentioned probiotic adaptation table of individuals with overexpression of Th1 helper cells is based on the secretion of IL-4/IFN-γ and IL-10 as indicators, and the corresponding multiple probiotic strains or the same species are different The probiotics in the top three rankings of the strains are matched to the individuals with overexpression of Th1 helper cells.

Among them, the above-mentioned probiotic adaptation table of individuals with excessive expression of Th2 helper cells is based on the secretion of IFN-γ/IL-4 and the IL-10 as indicators, and the corresponding multiple probiotic strains or the same species The probiotics in the top three rankings of different strains are matched to the individuals with over-expression of Th2 helper cells.

The second object of the present invention is to provide a method for screening probiotics that regulate individualized immunity. The screening method is to screen the probiotics that regulate the individualized immunity based on the information of the individualized probiotics database established by the first objective. The process is not subject to human mental judgment, and the screening results are directly produced by the information processing system. In other words, this screening method is an innovative application of big data analysis of probiotics, and then screening and matching probiotics that meet individualized immunity.

Specifically, the screening method includes screening IL-4/IFN-γ and corresponding multiple probiotic strains or first-ranked probiotics of the same species and different strains to give an individual with overexpression of Th1 helper cells Or screening IFN-γ/IL-4 and the corresponding multiple probiotic species or probiotics in the first order of the same species and different strains to give an individual with overexpression of Th2 helper cells.

The main role of Th1 helper cells is to resist the immune response of bacteria and protozoa in the cells. When Th1 helper cells are overexpressed and produce relatively too much IFN-γ, it can cause autoimmune diseases such as multiple sclerosis, psoriasis, rheumatoid arthritis, type 1 diabetes, organ transplant rejection, etc.

The main role of Th2 helper cells is to resist the immune response of extracellular multicellular parasites. When Th2 helper cells are overexpressed and produce relatively too much IL-4, it will cause allergy-related diseases, such as allergic rhinitis, asthma, and atopic dermatitis.

Preferably, the screening method further comprises screening the secretion amount of IL-10 and the corresponding probiotics of the plurality of probiotic strains or the sorted first-order probiotics of the same species and different strains to give the Th1 helper cell over-expression An individual, an individual with Th2 helper cell overexpression, or an individual with both Th1 helper cell overexpression and Th2 helper cell overexpression.

Accordingly, the method for screening probiotics that regulate individualized immunity according to the second objective of the present invention is based on the performance of the individualized probiotics selection index platform in the aforementioned individualized probiotics database for cells with different T helper types. Individuals are matched for precise probiotic screening. That is to say, the probiotic screening method provided by the second objective of the present invention is to screen out suitable probiotics according to individual differences, so as to achieve precise individual health care or regulation of individual immunity according to individual differences. purpose.

The third object of the present invention is to provide a method for screening probiotics that regulate the overexpression of Th1 helper cells, which is based on the individualized probiotic database obtained by the method for establishing an individualized probiotic database according to the first object. The screening of probiotics that can regulate the overexpression of Th1 helper cells; among them, IL-4/IFN-γ and the corresponding probiotic strains or the top three probiotics of the same species and different strains are screened as The probiotics can regulate the overexpression of Th1 helper cells.

Preferably, the above-mentioned method for screening probiotics that can regulate the overexpression of Th1 helper cells further comprises screening the secretion amount of IL-10 and the corresponding first order of the multiple probiotic strains or different strains of the same species. The probiotics are given to individuals with overexpression of Th1 helper cells. By increasing the secretion of IL-10 of the individual, the number of Th1 helper cells and the number of other immune cells in the individual are adjusted to balance, such as Th2 helper cells, while suppressing The individual’s inflammation reduces allergies or autoimmune reactions.

The fourth object of the present invention is to provide a method for screening probiotics that can regulate Th2 helper cell overexpression, which is based on the individualized probiotic database obtained by the method for establishing an individualized probiotic database according to the first object. The screening of probiotics that can regulate the overexpression of Th2 helper cells; where IFN-γ/IL-4 and the corresponding multiple probiotic strains or the top three probiotics of the same species and different strains are screened as The probiotics can regulate the overexpression of Th2 helper cells.

Preferably, the above-mentioned method for screening probiotics that can regulate the overexpression of Th2 helper cells further comprises screening the secretion amount of IL-10 and the corresponding first order of the multiple probiotic strains or different strains of the same species. The probiotics are given to individuals with overexpression of Th2 helper cells. By increasing the secretion of IL-10 of the individual, the number of Th2 helper cells and the number of other immune cells in the individual can be adjusted to balance, such as Th1 helper cells, while suppressing The individual’s inflammation reduces allergies or autoimmune reactions.

In summary, the content of the present invention includes (1). Provide a method for establishing an individualized probiotic database, which establishes the selection of individualized probiotic strains or different strains of the same species using the aforementioned cytokine information The personalized probiotic database of the index platform, which contains the ranking of the multiple probiotics corresponding to IL-4/IFN-γ, the ranking of the multiple probiotics corresponding to IFN-γ/IL-4, The ranking of the multiple probiotics and the pairing information of the individualized probiotics corresponding to the secretion of IL-10 are used as the basis for the selection of individualized probiotics. (2) Provide a method for screening probiotics that regulate individualized immunity. This method uses a dual-index system to screen out probiotics suitable for different individuals based on different individual conditions, so as to achieve precise selection of appropriate probiotics based on individualized differences Probiotics are for the purpose of individual health care or regulation of individual immunity. (3) Provide a method for screening probiotics that can regulate the overexpression of Th1 helper cells. This method is to screen IL-4/IFN-γ and the corresponding multiple probiotic strains or the same kind of different strains before sorting. The probiotics in the third sequence are used as the probiotics that can regulate the overexpression of Th1 helper cells, and at the same time or further screen the secretion of IL-10 and the corresponding multiple probiotic strains or different strains of the same species. The probiotics ranked first in the ranking serve as the probiotics capable of regulating the overexpression of Th1 helper cells. (4) Provide a method for screening probiotics that can regulate the overexpression of Th2 helper cells. The method is to screen IFN-γ/IL-4 and the corresponding multiple probiotic strains or the predecessor of the sorting of different strains of the same species. The probiotics in the third sequence are used as the probiotics that can regulate the overexpression of Th2 helper cells, and at the same time or further screen the secretion of IL-10 and the corresponding multiple probiotic strains or different strains of the same species. The probiotics in the first rank of the ranking serve as the probiotics that can regulate the overexpression of Th2 helper cells. Each probiotic screening method described in the present invention uses the personalized probiotic database described in the present invention as a screening and measurement system, applying big data analysis, and directly selecting and matching individualized differences by the information processing system without human mental judgment. Of probiotic strains.

Brief description of the drawings

no

The best way to realize the invention

The foregoing and other technical content, features, and effects of the present invention will be clearly presented in the following detailed description of a preferred embodiment with reference to the drawings. In order to thoroughly understand the present invention, detailed steps and their composition will be proposed in the following description. Obviously, the implementation of the present invention is not limited to the specific details familiar to those skilled in the field. On the other hand, well-known components or steps are not described in details to avoid unnecessary limitation of the present invention. The preferred embodiments of the present invention will be described in detail as follows. However, in addition to these detailed descriptions, the present invention can also be widely implemented in other embodiments, and the scope of the present invention is not limited, which is subject to the following patent scope .

The first embodiment of the present invention is to provide a method for establishing an individualized probiotic database. The establishment method includes but is not limited to the five steps described below.

Step 1: Provide multiple probiotic strains or different strains of the same species; the multiple probiotic strains refer to species belonging to different genus, different strains of the same genus or different strains of the same strain, and their content ratio Bifidobacterium bifidum, Bifidobacterium longum, Bifidobacterium infantis, Lactobacillus gasseri, Lactobacillus lactis, Lactococcus lactis, Bacillus coagulans, Lactobacillus delbrueckii subsp.bulgaricus, Lactobacillus johnsonii, Lactobacillus casei, Lactobacillus casei, Lactobacillus casei Lactobacillus paracasei), Bifidobacterium adolescentis, Lactobacillus salivarius, Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus brevis, Bifidobacterium brevis ( Bifidobacterium breve, Lactobacillus acidophilus, Streptococcus thermophilus, Lactobacillus helveticus, Clostridium butyricum, Bifidobacterium lactis, Lactobacillus rhamnosus (Lactobacillus rhamnosus), Lactobacillus reuteri (Lactobacillus reuteri), Streptococcus thermophilus (Faecalibacterium prausnitzii) or Akkermansia (Muciniphila).

Step 2: The multiple probiotic strains or different strains of the same species are separately co-cultured with biological samples derived from the same individual in vitro. The biological sample contains immune cells.

Step 3: Perform an analysis step to obtain cytokine information secreted by biological samples co-cultured with the multiple probiotic strains or the same different strains, the cytokine information including the secretion of IL-4 The ratio of the secretion of IFN-γ (IL-4/IFN-γ), the ratio of the secretion of IFN-γ and the secretion of IL-4 (IFN-γ/IL-4) and the secretion of IL-10 .

Step 4: Perform a sorting step, which is based on the value of the cytokine information to create sorting information of the value from high to low, and the sorting information includes the multiple probiotics corresponding to IL-4/IFN-γ. The ranking of bacteria, the ranking of the plurality of probiotics corresponding to IFN-γ/IL-4, the ranking of the plurality of probiotics corresponding to the secretion of IL-10, or any combination of the above rankings.

Step 5: Perform a pairing step to complete the establishment of the individualized probiotics database. The pairing step includes making the IL-4/IFN-γ and the corresponding probiotic strains or the same species. The ranked first three probiotics of different strains and the secretion amount of the IL-10 and the corresponding multiple probiotic strains or the first three ranked probiotics of the same species of different strains are paired to a Th1 Individuals with overexpression of helper cells; and the IFN-γ/IL-4 and the corresponding probiotic strains in the top three ranks of the probiotics and the secretion of IL-10 and the corresponding Multiple probiotic strains or the top three ranked probiotics of the same species and different strains are paired to an individual with overexpression of Th2 helper cells.

In a specific embodiment, the biological sample contains immune cells.

In a specific embodiment, the immune cells are isolated from a blood sample or an oral mucosal sample.

In a specific embodiment, the analysis method used in the analysis step includes enzyme immunoassay/enzyme-linked immunosorbent assay (ELISA), flow cytometry, chemiluminescence immunoassay (CLIA) or flow cytometry bead array multiple analysis system . Preferably, the analysis method is enzyme immunoassay/enzyme-linked immunosorbent assay (ELISA).

In a specific embodiment, the ratio of the secretion of IL-4 to the secretion of IFN-γ (IL-4/IFN-γ) and the secretion of IL-10 is an individual whose Th1 helper cell overexpression Selection index of probiotics.

In a specific embodiment, the ratio of the secretion of IFN-γ to the secretion of IL-4 (IFN-γ/IL-4) and the secretion of IL-10 is an individual whose Th2 helper cell overexpression Selection index of probiotics.

In a specific embodiment, the sorting step and the pairing step are performed by an information processing system.

In a representative embodiment, the process steps include: (1) confirming individual differences, which can be learned by allergen testing or individual investigation; (2) taking blood or oral mucosa samples of the above individuals; (3) by Immune cells are isolated from the blood or oral mucosa sample; (4) The immune cells are co-cultured with multiple probiotic strains or different strains of the same species; (5) The immune cells and the multiple probiotic bacteria are analyzed by ELISA technology The content of cytokine secreted by different strains of the same species or the same species after co-cultivation, the cytokine includes IFN-γ, IL-4 and IL-10; (6) The multiple probiotics are output by a computer equipped with a bioinformatics statistical computing system Bacteria or different strains of the same species and the ranking of the above-mentioned cytokine-related information. The order of the ranking is the numerical value of the above-mentioned cytokine-related information in descending order. The ranking includes IL-4/IFN-γ respectively. The corresponding ranking of the plurality of probiotics, the ranking of the plurality of probiotics corresponding to IFN-γ/IL-4, and the ranking of the plurality of probiotics corresponding to the secretion of IL-10 respectively; and (7 ) Produce an individualized and differentiated probiotic adaptation table, wherein the IL-4/IFN-γ and the corresponding probiotic strains or the top three probiotics in the order of the same species and different strains and The secretion of IL-10 and the corresponding probiotic strains or the top three probiotics of the same species and different strains are matched to an individual with overexpression of Th1 helper cells; and the IFN-γ/ IL-4 and the corresponding probiotic strains or the top three probiotics of the same species and different strains and the secretion amount of the IL-10 and the corresponding probiotic strains or The top three probiotics of the same species and different strains are matched to an individual with overexpression of Th2 helper cells; thereby completing the establishment of the individualized probiotic database.

The second embodiment of the present invention is to provide a method for screening probiotics that regulate individualized immunity, which is an individualized probiotic database obtained according to the method for establishing an individualized probiotic database according to the first embodiment of the present invention Perform the screening of the probiotics that modulate the individualized immunity; it includes screening IL-4/IFN-γ and the corresponding multiple probiotic strains or the first-ranked probiotics of the same species and different strains. An individual with overexpression of Th1 helper cells; or screening of IFN-γ/IL-4 and corresponding multiple probiotic strains or the first order probiotics of the same species and different strains to give a Th2 helper cell excessive Individuals of performance.

Preferably, the method for screening probiotics that regulate individualized immunity further comprises screening the secretion amount of IL-10 and the corresponding first sequence of the multiple probiotic strains or the sequence of different strains of the same species. The probiotics can be administered to individuals with overexpression of Th1 helper cells, individuals with overexpression of Th2 helper cells, or an individual with overexpression of Th1 helper cells and Th2 helper cells at the same time.

The third embodiment of the present invention is to provide a method for screening probiotics that can regulate Th1 helper cell overexpression, which is an individualized probiotic obtained according to the method for establishing an individualized probiotic database as described in the first embodiment The database is used to screen the probiotics that can regulate the excessive performance of Th1 helper cells; among them, IL-4/IFN-γ and the corresponding multiple probiotic strains or the top three probiotics of the same species and different strains are screened. Bacteria are the probiotics that can regulate the overexpression of Th1 helper cells.

Preferably, the method for screening probiotics that can regulate the overexpression of Th1 helper cells further comprises screening the secretion of IL-10 and the corresponding sequence of multiple probiotic strains or different strains of the same species. A sequence of probiotics is given to individuals with overexpression of Th1 helper cells.

The fourth embodiment of the present invention is to provide a method for screening probiotics that can regulate Th2 helper cell overexpression, which is an individualized probiotic obtained according to the method for establishing an individualized probiotic database as described in the first embodiment The database is used to screen the probiotics that can regulate the excessive performance of Th2 helper cells; among them, IFN-γ/IL-4 and the corresponding multiple probiotic strains or the top three probiotics of the same species and different strains are screened. Bacteria are the probiotics that can regulate the overexpression of Th2 helper cells.

Preferably, the method for screening probiotics that regulate the overexpression of Th2 helper cells further comprises screening the secretion amount of IL-10 and the corresponding probiotics of the first order of the plurality of probiotic strains Strains are administered to individuals who overexpress the Th2 helper cells.

The following example is an experiment conducted based on the content described in the above-mentioned embodiment, and is used as a detailed description of the present invention accordingly.

The pre-experimental tasks include: preparation of balanced salt solution (or sterile PBS), preparation of wash buffer (wash buffer (PBS+0.05% Tween-20)), preparation of stop solution (2N H ₂ SO ₄ )) and preparation Medium (medium).

Probiotics re-dissolution and sterilization: re-dissolve the powder of the capsule with sterile PBS 5ml; take 1ml of bacterial solution at 80°C for 30min (sterilization), and then dilute it to the required concentration for use in the co-cultivation step with biological samples.

Individual biological samples and co-cultivation: Taking individual white blood cells as biological samples co-cultured with probiotic strains, the separation and extraction steps and co-cultivation steps are as follows: Individual blood samples and balanced salt solution are mixed 1:1 to form a diluted Blood sample; shake Ficoll-Paque PLUS evenly; add Ficoll-Paque PLUS to centrifuge tube; carefully add diluted blood; centrifuge 400×g for 30-40 minutes (18-20°C); remove the uppermost plasma (plasma) ; Obtain the second layer of Lymphocytes Monocytes Platelets; add at least three times the volume of the balanced salt solution to the obtained cells and gently disperse the cells; centrifuge at 60-100×g for 10 minutes (18-20°C); remove the supernatant, Add the same volume of balanced salt solution as in step 8 and gently break up the cells; centrifuge at 60-100×g for 10 minutes (18-20°C); remove the supernatant, break up the cells with cell culture solution, and count the cells; Plant the cells in a 24-well plate, 4x10 ⁵ /well; co-culture with probiotics of different counts for 40 hours; aspirate the supernatant, centrifuge at 4°C to separate the suspended bacteria, the supernatant can be used for ELISA experiments Perform analysis of IFN-γ, IL-10 and IL-4.

General procedure for ELISA analysis of cytokines

In this ELISA analysis experiment, the BioLegend's ELISA MAX ^TM Deluxe Set kit was used to analyze the secretion amounts of IFN-γ, IL-4 and IL-10, respectively.

The steps involved in the analysis process are as follows: immobilize 100ul of the diluted capture antibody on a 96-well plate (if IFN-γ is to be analyzed, the capture antibody is IFN-γ capture antibody), and let it stand overnight at 2～8℃; use Wash 4 times with PBS buffer, then add 200ul content diluent (Assay diluent), let stand for 1 hour at room temperature; wash 4 times with PBS buffer, then add 100ul diluted standard and biological sample to be tested, Let stand for 2 hours at room temperature; wash 4 times with PBS buffer, then add 100ul of diluted detection antibody (if IFN-γ is to be analyzed, the detection antibody is IFN-γ detection antibody), and stand at room temperature for 1 hour ; Wash 4 times with PBS buffer, then add 100ul of diluted avidin (Avidin-HRP), let stand at room temperature for 0.5 hours; wash 5 times with PBS buffer, and then add 100ul color developing solution in the dark ( TMB substrate solution), let stand at room temperature for 20 minutes; add 100ul stop solution (stop solution) to stop the reaction, and then read the 450nm and 570nm instrument analysis values. In this way, the secretion of IFN-γ, IL-4 and IL-10 can be obtained.

According to the cytokine information obtained from the above ELISA analysis, input the information processing system to complete the establishment of a database of individualized probiotic bacteria.

Individual A: After the investigation of the physiological condition, the individual A has symptoms related to autoimmune disorders and type 1 diabetes; the individual A is defined as an individual with overexpression of Th1 cells.

The sorting and adaptation table of individual A’s individualized probiotics database prepared according to the present invention is shown in Table 1. The sample used for detection and analysis is individual A’s blood sample.

Table I

Blood sample

Individual B: After the investigation of the physiological condition, the individual B has symptoms related to autoimmune disorders and rheumatoid arthritis; the individual B is defined and classified as an individual with overexpression of Th1 cells.

The sorting and adaptation table of individual B’s individualized probiotic database prepared according to the present invention is shown in Table 2, and the specimen used for detection and analysis is the oral specimen of individual B.

Table II

Oral specimen

According to Table 1 or Table 2, the probiotics with IL-4/IFN-γ ranking 1 or 2 or the probiotics with IL-10 (pg/mL) ranking 1 or 2 are given priority to individual A or individual B.

Individual C: After a physical investigation, the individual C has allergic symptoms such as nose allergy and atopic dermatitis; the individual C is defined as an individual with overexpression of Th2 cells.

The sorting and adaptation table of the individualized probiotic database of individual C prepared according to the present invention is shown in Table 3, and the specimen for detection and analysis is the blood specimen of individual C.

Table Three

Blood sample

Individual D: After the investigation of the physiological condition, the individual D has symptoms of allergy to pollen; the individual D is defined and classified as an individual with overexpression of Th2 cells.

The sorting and adaptation table of individual D’s individualized probiotic database prepared according to the present invention is shown in Table 4, and the specimen used for detection and analysis is the oral specimen of individual D.

Table Four

Oral specimen

According to Table 3 or Table 4, the probiotics with IFN-γ/IL-4 ranking 1 or 2 or the probiotics with IL-10 (pg/mL) ranking 1 or 2 are preferentially screened and given to individual C or individual D.

Although specific examples are used to describe the present invention above, the scope of the present invention is not limited thereby. As long as the spirit of the present invention is not deviated, those skilled in the art understand that various modifications or changes can be made without departing from the intent and scope of the present invention. In addition, the abstract part and title are only used to assist in searching for patent documents, and are not used to limit the scope of rights of the present invention.

Claims (15)

A method for establishing an individualized probiotic database, which is characterized in that the steps include: (1) Provide multiple probiotic strains or different strains of the same species; (2) The multiple probiotic strains or different strains of the same species are separately co-cultured with biological samples derived from the same individual in vitro; (3) Perform an analysis step to obtain the cytokine information secreted by the biological samples co-cultured with the multiple probiotic strains or the same different strains, and the cytokine information includes the secretion amount of IL-4 The ratio of the secretion of IFN-γ (IL-4/IFN-γ), the ratio of the secretion of IFN-γ and the secretion of IL-4 (IFN-γ/IL-4) and the secretion of IL-10 ； (4) Perform a sorting step. The sorting step creates sorting information from high to low based on the value of the cytokine information, and the sorting information includes the plurality of probiotics corresponding to IL-4/IFN-γ. The sorting of strains or different strains of the same species, the multiple probiotic strains corresponding to IFN-γ/IL-4, or the sorting of different strains of the same species, and the multiple probiotics corresponding to the secretion of IL-10. Bacterial species or a sequence of different strains of the same species or a combination of the above-mentioned sequences; and (5) Perform a pairing step to complete the establishment of the individualized probiotic database. The pairing step is to make the IL-4/IFN-γ and the corresponding probiotic strains or the same species The ranked first three probiotics of different strains and the secretion amount of the IL-10 and the corresponding multiple probiotic strains or the first three ranked probiotics of the same species of different strains are paired to a Th1 Individuals with overexpression of helper cells; and the IFN-γ/IL-4 and the corresponding probiotic strains or the top three probiotics of the same species and different strains and the secretion of the IL-10 The amount and the corresponding probiotic strains of the plurality of probiotic strains or the top three ranked probiotics of the same species and different strains are matched to an individual with overexpression of Th2 helper cells. The method for establishing an individualized probiotic database according to claim 1, wherein the plurality of probiotic strains refer to strains belonging to different genus, or the same strain but different strains, which include Bifid Bacillus vulgaris, Rhizopus bifidus, Lactobacillus gasseri, Lactobacillus gasseri, Lactobacillus lactis, Lactococcus lactis, Bacillus coagulans, Lactobacillus bulgaricus, Lactobacillus johnsonii, Lactobacillus kjerii, Lactobacillus gasseri, Lactobacillus gasseri, Lactobacillus lactis, Lactococcus lactis, Bacillus coagulans, Lactobacillus bulgaricus, Lactobacillus johnsonii, Lactobacillus kjerii, Lactobacillus paracasei, Bifidobacterium adolescentis, Lactobacillus salivarius, Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus brevis, Bifidobacterium breve, Lactobacillus acidophilus, Streptococcus thermophilus, Lactobacillus helveticus, Clostridium butyricum, Bifidobacterium reiters, Lactobacillus rhamnosus, Lactobacillus reuteri, Streptococcus thermophilus, Faeculus prasutii or Akkermansia. The method for establishing an individualized probiotic database according to claim 1, wherein the biological sample contains immune cells. The method for establishing an individualized probiotic database according to claim 3, wherein the immune cells are isolated from a blood sample or an oral mucosal sample. The method for establishing an individualized probiotic database according to claim 1, wherein the analysis method used in the analysis step includes enzyme immunoassay/enzyme-linked immunosorbent assay (ELISA), flow cytometry, chemical Luminescence immunoassay (CLIA) or flow microbead array multiple analysis system. The method for establishing an individualized probiotic database according to claim 1, wherein the ratio of the secretion of IL-4 to the secretion of IFN-γ (IL-4/IFN-γ) and the IL-10 The amount of secretion is used as an indicator of probiotic selection for individuals with overexpression of Th1 helper cells. The method for establishing an individualized probiotic database according to claim 1, wherein the ratio of the secretion of IFN-γ to the secretion of IL-4 (IFN-γ/IL-4) and the IL-10 The amount of secretion is used as an indicator of probiotic selection for individuals with overexpression of Th2 helper cells. The method for establishing an individualized probiotic database according to claim 1, wherein the sorting step and the matching step are executed by an information processing system. A method for screening probiotics that regulate individualized immunity, characterized in that it is an individualized probiotics database obtained according to the method for establishing an individualized probiotics database according to claim 1 to adjust individualized immunity Screening of probiotics; it includes screening IL-4/IFN-γ and the corresponding multiple probiotic strains or the first-ranked probiotics of the same species and different strains to give a Th1 helper cell overexpression Individuals; or screen IFN-γ/IL-4 and the corresponding multiple probiotic strains or the first-ranked probiotics of the same species and different strains to give an individual with overexpression of Th2 helper cells. The method for screening probiotics that regulate individualized immunity according to claim 9, characterized in that it further comprises screening the secretion of IL-10 and the corresponding ranking of the multiple probiotic strains or different strains of the same species The probiotics in the first order are given to individuals with overexpression of Th1 helper cells, individuals with overexpression of Th2 helper cells, or individuals with both overexpression of Th1 helper cells and overexpression of Th2 helper cells. The method for screening probiotics that modulate individualized immunity according to claim 9, characterized in that: a plurality of probiotic strains or different strains of the same species are provided; the plurality of probiotic strains refer to the belonging of different bacterial genera Bacteria species, different species of the same bacterium or different species of the same species but different strains, which include Bifidia, Rhizopus Bifidus, Infant Bifidus, Lactobacillus gasseri, Lactobacillus lactis, Lactococcus lactis, Bacillus coagulans, Lactobacillus bulgaricus, Lactobacillus johnsonii, Lactobacillus Kjeldahl, Lactobacillus paracasei, Bifidobacterium adolescentis, Lactobacillus salivarius, Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus brevis, Lactobacillus brevis, Short double Fidobacteria, Lactobacillus acidophilus, Streptococcus thermophilus, Lactobacillus helveticus, Clostridium butyricum, Bifidobacterium reiters, Lactobacillus rhamnosus, Lactobacillus reuteri, Streptococcus thermophilus, Prussian feces Bacillus or Akkermansia. A method for screening probiotics capable of regulating the overexpression of Th1 helper cells, characterized in that it is an individualized probiotic database obtained according to the method for establishing an individualized probiotic database according to claim 1 to perform the regulation of Th1 Screening of probiotics with over-expression of helper cells; among them, IL-4/IFN-γ and the corresponding multiple probiotic strains or the top three probiotics of the same species and different strains are screened as the probiotics with Probiotics that regulate the overexpression of Th1 helper cells. The method for screening probiotics capable of regulating the overexpression of Th1 helper cells according to claim 12, characterized in that it further comprises screening the secretion amount of IL-10 and the corresponding multiple probiotic strains or different strains of the same species The probiotics in the first order of the ranking are given to individuals with overexpression of Th1 helper cells. A method for screening probiotics capable of regulating the overexpression of Th2 helper cells, characterized in that: it is an individualized probiotic database obtained according to the method for establishing an individualized probiotic database according to claim 1 to perform the regulation of Th2 Screening of probiotics with overexpression of helper cells; among them, IFN-γ/IL-4 and the corresponding multiple probiotic strains or the first three sequenced probiotics of the same species and different strains are screened as the probiotics with Probiotics that regulate the overexpression of Th2 helper cells. The method for screening probiotics that can regulate the overexpression of Th2 helper cells according to claim 14, characterized in that it further comprises screening the secretion amount of IL-10 and the corresponding multiple probiotic strains or different strains of the same species The probiotics in the first order of the ranking are given to individuals with over-expression of Th2 helper cells.