Diagnostic support method for enteritis based on biogas analysis

Abstract

To provide a diagnostic support method for enteritis in a simple and non-invasive manner.SOLUTION: Provided is a diagnostic support method for enteritis in a subject, comprising: measuring the concentration of hydrogen in a gas mixture discharged from the subject; and comparing the measured value or the standard deviation of the measured value with a reference value.SELECTED DRAWING: Figure 4

Description

[Technical field]

[0001] The present invention relates to a method for assisting in the diagnosis of enteritis based on the analysis of biological gases. [Background technology]

[0002] Inflammatory bowel disease (IBD) is a chronic inflammatory disease that is believed to develop due to a complex interaction of genetic and environmental factors. The number of patients with IBD is steadily increasing worldwide, which is a major socio-economic burden. IBD pathology often repeats relapse and remission, and early intervention before the disease progresses to severe or chronic status is important for the treatment of IBD. However, an effective method for predicting disease relapse has not yet been established.

[0003] Currently, endoscopic examination is the most common method for diagnosing IBD, but endoscopic examination has many issues, such as the length of time required, high cost, and high invasiveness. Therefore, there is a need to establish a non-invasive method for predicting IBD disease. Among these, breath analysis has attracted attention as a new biomarker for IBD.

[0004] In fact, previous studies have reported that there are differences in breath components between healthy subjects and IBD patients, suggesting a relationship between IBD and breath. Non-Patent Document 1 analyzes volatile organic compounds (VOCs) in breath components of ulcerative colitis patients, Crohn's disease patients, and healthy subjects using selective ion flow tube mass spectrometry (SIFT-MS), and reports differences in VOC profiles between groups. Non-Patent Document 2 analyzes volatile organic compounds (VOCs) in breath of pediatric ulcerative colitis patients, Crohn's disease patients, and controls using ion molecule reaction mass spectrometry, and reports that the VOC pattern of IBD patients can be distinguished from that of controls. Non-Patent Document 3 reports on VOCs in the breath and feces of irritable bowel syndrome (IBS) patients, IBD patients, and healthy subjects. Non-Patent Document 4 reports that VOCs in breath can be used to distinguish healthy subjects, active Crohn's disease patients, and Crohn's disease patients in remission.

[0005] Most of the studies to date have focused on volatile compounds with large molecular weights that are easy to quantify, and there has been insufficient research on other molecules, including low molecular weight compounds. Furthermore, most studies have focused on active IBD patients, so there has been no fundamental evaluation of IBD and changes in exhaled breath components. [Prior art documents] [Non-patent literature]

[0006] [Non-Patent Document 1] Journal of Crohn's and Colitis, 2015, 731-737 [Non-Patent Document 2] PLoS ONE 12(8): e0184118 [Non-Patent Document 3] EBioMedicine 54 (2020) 102725 [Non-Patent Document 4] Inflamm Bowel Dis 2015;21:17761785) Summary of the Invention [Problem to be solved by the invention]

[0007] The present inventors analyzed the gas mixture excreted from a subject to evaluate whether components in the gas mixture could serve as biomarkers to aid in the diagnosis of enteritis.

[0008] An object of the present invention is to provide a non-invasive and simple method for assisting in the diagnosis of enteritis. [Means for solving the problem]

[0009] The present invention encompasses the embodiments described below. Section 1. 1. A method for aiding in the diagnosis of enteritis in a subject, comprising: Measuring the concentration of hydrogen in the gas mixture emitted from the subject; and Comparing said measurements or standard deviations of measurements with reference values. A method comprising: Section 2. Item 2. The method according to item 1, wherein the enteritis comprises inflammatory bowel disease. Section 3. Item 10. The method of item 1, wherein the gas mixture exhaled by the subject comprises exhaled breath. Section 4. the reference value being an average value of the concentration of hydrogen in the gas mixture excreted from patients with enteritis, a median value of the concentration of hydrogen in the gas mixture excreted from patients with enteritis, a value lower than the average value or the median value, a cut-off value of the concentration of hydrogen in the gas mixture that distinguishes between patients with enteritis and healthy subjects, or a standard deviation of the concentration of hydrogen in the gas mixture excreted from patients with enteritis, 4. The method according to any one of items 1 to 3, further comprising determining that there is a high possibility of enteritis when a measured value of hydrogen concentration or a standard deviation of the measured value is equal to or less than the reference value. Section 5. The reference value is an average value of the concentration of hydrogen in the gas mixture exhaled from a healthy subject, a median value of the concentration of hydrogen in the gas mixture exhaled from a healthy subject, a value higher than the average value or the median value, or a standard deviation of the concentration of hydrogen in the gas mixture exhaled from a healthy subject; Item 4. The method according to any one of Items 1 to 3, further comprising determining that the possibility of contracting enteritis is low when the measured value of the hydrogen concentration or the standard deviation of the measured value is equal to or greater than the reference value. Section 6. the reference value being a value of the concentration of hydrogen in a gas mixture emitted from the subject, previously measured in the subject; The method according to any one of items 1 to 3, further comprising determining that there is a high possibility that enteritis has progressed when the concentration of hydrogen in the gas mixture discharged from the subject measured at a certain time point in the subject is lower than the standard value, or determining that there is a high possibility that enteritis has not progressed when the concentration of hydrogen in the gas mixture discharged from the subject measured at a certain time point in the subject is equal to or higher than the standard value. Section 7. The subject is a patient with enteritis, the reference value being a measurement of the concentration of hydrogen in the gas mixture excreted from the subject prior to administration of a drug; measuring the concentration of hydrogen includes measuring the concentration of hydrogen in a gas mixture excreted from the subject after administering the drug; The method according to any one of items 1 to 3, further comprising determining that the therapeutic effect of the drug is high when a measured value of the concentration of hydrogen in the gas mixture discharged from the subject after administration of the drug is higher than the reference value, or determining that the therapeutic effect of the drug is low when a measured value of the concentration of hydrogen in the gas mixture discharged from the subject after administration of the drug is equal to or lower than the reference value. Section 8. A method for screening for a substance effective in treating enteritis, comprising the steps of: administering a test substance to a subject suffering from enteritis; comparing the concentration of hydrogen in the gas mixture exhaled from the subject prior to administration of the test substance with the concentration of hydrogen in the gas mixture exhaled from the subject after administration of the test substance; and selecting the test substance as a candidate effective for treating enteritis when the concentration of hydrogen in the gas mixture exhaled from the subject after administration of the test substance is higher than the concentration of hydrogen in the gas mixture exhaled from the subject before administration of the test substance. The method further comprising: Section 9. Item 9. The method according to item 1 or 8, further comprising measuring the concentration of methanethiol in the gas mixture discharged from the subject. Section 10. Item 9. The method of any one of items 1 to 8, further comprising measuring the concentration of hydrogen sulfide in the gas mixture excreted by the subject. Section 11. Item 9. The method of item 1 or 8, further comprising measuring the concentration or amount of lipocalin-2 in the subject's stool. Section 12. Item 9. The method of item 1 or 8, further comprising measuring the relative abundance of at least one species of enterobacteria selected from the group consisting of Akkermansiaceae, Rikenellaceae, and Tannerellaceae in the feces of the subject. [Brief description of the drawings]

[0010] [Figure 1] Drug administration schedule for mice. Mice were administered DSS or dextran for 5 days and observed for 15 days. [Diagram 2] Schematic diagram of a biogas measuring device. [Diagram 3] (A) Body weight change over 15 days in the DSS-administered group, (B) Changes in fecal lipocalin-2 concentration on days 0, 1, 3, 5, 7, 9, 12, and 15 in the DSS-administered group. (C) Body weight change over 15 days in the dextran-administered group, (D) Changes in fecal lipocalin-2 concentration on days 0, 1, 3, 5, 7, 9, 12, and 15 in the dextran-administered group. *p < 0.05, **p < 0.01, ***p < 0.001. (A,C) Friedman's test. [Figure 4] Changes in biogas concentrations. Changes in concentrations of (A) H2, (B) NH3, (C) H2S, (D) CH3SH, and (E) C2H5SH in the DSS-administered group. Changes in concentrations of (F) H2, (G) NH3, (H) H2S, (I) CH3SH, and (J) C2H5SH in the dextran-administered group. (K) Correlation heat map between biogas and average body weight. (L) Correlation heat map between biogas and average fecal Lipocalin-2 concentrations. (K,L) are Pearson correlation coefficients. [Diagram 5] Graph showing H2 concentration and standard deviation (solid line) over 15 days in (A) the DSS-administered group and (B) the dextran-administered group. [Figure 6] ROC curves for each biogas. (A) H2, (B) H2S. [Figure 7]Diversity of gut bacteria. (A) Amplicon sequence variants (ASVs) observed. Observed ASVs (left) and Shannon's index (right), *p < 0.05, **p < 0.01. (B) Principal coordinate analysis using unweighted Unifrac distance. [Figure 8] Changes in gut microbiota over time. (A) Composition of gut microbiota. (B) Changes in the abundance of Akkermansiaceae. (C) Changes in the abundance of Rickenellaceae. (D) Changes in the abundance of Tannerellaceae. *p < 0.05, **p < 0.01, ***p < 0.001. Friedman's test [Figure 9] Heat map of correlation between the proportion of intestinal bacteria and fecal Lipocalin-2 concentration, body weight, and H2 concentration. Pearson correlation coefficient [Figure 10] (D) Schematic diagram of H2, Lipocalin-2, and gut bacteria with high and low correlation coefficients in the correlation heat map. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0011] As used herein, the singular forms (a, an, the) are intended to include both the singular and the plural unless otherwise expressly stated in the specification or clearly contradictory in context.

[0012] In this specification, the term "comprise" is a concept that encompasses "consist essentially of" and "consist only of."

[0013] In the present specification, the subject is preferably a mammal. The mammal includes mammals including humans, such as humans, cows, horses, pigs, monkeys, dogs, cats, mice, rats, guinea pigs, hamsters, rabbits, goats, sheep, etc., preferably humans, mice, and rats, more preferably humans.

[0014] In this specification, "affected" includes "onset."

[0015] As used herein, the term "treatment" refers to the cure or amelioration of a disease or symptom, or the suppression of a symptom, and includes "prevention." "Prevention" refers to preventing the onset of a disease or symptom.

[0016] As used herein, "aid in diagnosis" may be used interchangeably with "detection."

[0017] According to one aspect of the invention there is provided a method for aiding in the diagnosis of enteritis in a subject, comprising the steps of: Measuring the concentration of hydrogen in the gas mixture emitted from the subject; and Comparing said measurements or the standard deviation of said measurements with a reference value. A method is provided, comprising:

[0018] In the following, unless otherwise specified, the units of the measured values ​​and the reference values ​​to be compared are the same, i.e., the measured hydrogen concentration is compared with the reference value whose unit is hydrogen concentration, and the standard deviation of the hydrogen concentration is compared with the reference value whose unit is standard deviation.

[0019] The present inventors found that the hydrogen (H2) concentration in the gas mixture excreted from a subject decreased with the progression of experimental colitis. The hydrogen concentration in the gas mixture excreted from a subject can be used as a biomarker for predicting the onset of colitis.

[0020] In one embodiment, the subject is a subject suffering from enteritis, suspected of having enteritis, or at risk of suffering from enteritis. A "patient suspected of having enteritis" may be a subject who is subjectively suspected by the subject himself / herself, or a subject who has been determined or diagnosed with enteritis cancer based on some objective grounds.

[0021] In one embodiment, the enteritis comprises inflammatory bowel disease, including ulcerative colitis and Crohn's disease.

[0022] In one embodiment, the gas mixture exhaled by the subject comprises exhaled breath.

[0023] Aids in the diagnosis of enteritis include determination of the possibility of suffering from enteritis, determination of the possibility of developing enteritis, determination of the presence or absence of enteritis, determination of the severity of enteritis, prediction of the prognosis of enteritis after treatment, determination of the preventive effect of enteritis, determination of the effectiveness of treatment of enteritis, determination of enteritis patients for whom a therapeutic drug is effective (responders), test methods for diagnosing enteritis (particularly early diagnosis), and test methods for treating enteritis (particularly early treatment).

[0024] In one embodiment, the reference value is a mean value of the concentration of hydrogen in the gas mixture excreted by patients with enteritis, a median value of the concentration of hydrogen in the gas mixture excreted by patients with enteritis, a value lower than the mean value or the median value, a cut-off value of the concentration of hydrogen in the gas mixture that distinguishes between patients with enteritis and healthy subjects, or a standard deviation of the concentration of hydrogen in the gas mixture excreted by patients with enteritis; The method for assisting in the diagnosis of enteritis in a subject may further include determining that there is a high likelihood of enteritis if the measured value of hydrogen concentration or the standard deviation of the measured value is equal to or less than the reference value.

[0025] In one embodiment, the reference value is a mean value of the concentration of hydrogen in the gas mixture excreted by patients with enteritis, a median value of the concentration of hydrogen in the gas mixture excreted by patients with enteritis, a cut-off value of the concentration of hydrogen in the gas mixture that separates patients with enteritis from healthy subjects, or a standard deviation of the concentration of hydrogen in the gas mixture excreted by patients with enteritis; The method for assisting in the diagnosis of enteritis in a subject may further include determining that the subject is unlikely to suffer from enteritis if the measured hydrogen concentration or the standard deviation of the measured hydrogen concentration is higher than the reference value.

[0026] In one embodiment, the reference value is a mean value of the concentration of hydrogen in the gas mixture excreted by patients with enteritis, a median value of the concentration of hydrogen in the gas mixture excreted by patients with enteritis, a value lower than the mean value or the median value, a cut-off value of the concentration of hydrogen in the gas mixture that separates patients with enteritis from healthy subjects, or a standard deviation of the concentration of hydrogen in the gas mixture excreted by healthy subjects; The method for assisting in the diagnosis of enteritis in a subject may further include determining that there is a high likelihood of enteritis when the measured value of the hydrogen concentration or the standard deviation of the measured value is equal to or less than the reference value, or determining that there is a low likelihood of enteritis when the measured value of the hydrogen concentration or the standard deviation of the measured value is higher than the reference value.

[0027] The cutoff value for distinguishing between enteritis patients and healthy individuals is well known in the field of medical statistics, and a person skilled in the art can appropriately calculate the cutoff value for the concentration of hydrogen in a gas mixture using statistical methods based on the data of enteritis patients and healthy individuals.

[0028] In one embodiment, the reference value is the average value of the concentration of hydrogen in the gas mixture exhaled by the healthy individual, the median value of the concentration of hydrogen in the gas mixture exhaled by the healthy individual, a value higher than the average value or the median value, or the standard deviation of the concentration of hydrogen in the gas mixture exhaled by the healthy individual; The method for assisting in the diagnosis of enteritis in a subject may further include determining that the possibility of suffering from enteritis is low when the measured value of the hydrogen concentration or the standard deviation of the measured value is equal to or greater than the above-mentioned reference value.

[0029] In one embodiment, the reference value is the average concentration of hydrogen in the gas mixture exhaled by healthy individuals or the median concentration of hydrogen in the gas mixture exhaled by healthy individuals; The method for assisting in the diagnosis of enteritis in a subject may further include determining that there is a high likelihood of enteritis if the measured hydrogen concentration or the standard deviation of the measured value is lower than the reference value.

[0030] In one embodiment, the reference value is the average value of the concentration of hydrogen in the gas mixture exhaled by the healthy individual, the median value of the concentration of hydrogen in the gas mixture exhaled by the healthy individual, a value higher than the average value or the median value, or the standard deviation of the concentration of hydrogen in the gas mixture exhaled by the healthy individual; The method for assisting in the diagnosis of enteritis in a subject may further include determining that the possibility of enteritis is low if the measured value of the hydrogen concentration or the standard deviation of the measured value is equal to or greater than the reference value, or determining that the possibility of enteritis is high if the measured value of the hydrogen concentration or the standard deviation of the measured value is lower than the reference value.

[0031] In one embodiment, the reference value is a value of the concentration of hydrogen in a gas mixture emitted from the subject, previously measured in the subject; The method for assisting in the diagnosis of enteritis in a subject further includes determining that there is a high likelihood that enteritis is progressing when the concentration of hydrogen in the gas mixture exhaled from the subject measured at a certain time point in the subject is lower than the reference value.

[0032] In one embodiment, the reference value is a value of the concentration of hydrogen in a gas mixture emitted from the subject, previously measured in the subject; The method for assisting in the diagnosis of enteritis in a subject further includes determining that there is a high likelihood that enteritis has not progressed if the concentration of hydrogen in the gas mixture discharged from the subject measured at a certain point in time is equal to or greater than the reference value.

[0033] In one embodiment, the reference value is a value of the concentration of hydrogen in a gas mixture emitted from the subject, previously measured in the subject; The method for assisting in the diagnosis of enteritis in the subject further includes determining that there is a high possibility that enteritis has progressed when the concentration of hydrogen in the gas mixture discharged from the subject measured at a certain time point in the subject is lower than the reference value, or determining that there is a high possibility that enteritis has not progressed when the concentration of hydrogen in the gas mixture discharged from the subject measured at a certain time point in the subject is equal to or higher than the reference value.

[0034] In one embodiment, the subject is a patient with enteritis, the reference value being a measurement of the concentration of hydrogen in the gas mixture excreted from the subject prior to administration of a drug; measuring the concentration of hydrogen includes measuring the concentration of hydrogen in a gas mixture excreted from the subject after administering the drug; The method for assisting in the diagnosis of enteritis in the subject further includes determining that the therapeutic effect of the drug is high when a measured value of the concentration of hydrogen in the gas mixture discharged from the subject after administration of the drug is higher than the reference value.

[0035] In one embodiment, the subject is a patient with enteritis, the reference value being a measurement of the concentration of hydrogen in the gas mixture excreted from the subject prior to administration of a drug; measuring the concentration of hydrogen includes measuring the concentration of hydrogen in a gas mixture excreted from the subject after administering the drug; The method for assisting in the diagnosis of enteritis in the subject further includes determining that the therapeutic effect of the drug is low when a measured value of the concentration of hydrogen in the gas mixture discharged from the subject after administration of the drug is equal to or lower than the reference value.

[0036] In one embodiment, the subject is a patient with enteritis, the reference value being a measurement of the concentration of hydrogen in the gas mixture excreted from the subject prior to administration of a drug; measuring the concentration of hydrogen includes measuring the concentration of hydrogen in a gas mixture excreted from the subject after administering the drug; The method for assisting in the diagnosis of enteritis in a subject further includes determining that the therapeutic effect of the drug is high when a measured value of the hydrogen concentration in the gas mixture discharged from the subject after administration of the drug is higher than the reference value, or determining that the therapeutic effect of the drug is low when a measured value of the hydrogen concentration in the gas mixture discharged from the subject after administration of the drug is equal to or lower than the reference value.

[0037] The method for assisting in the diagnosis of enteritis in a subject according to one aspect of the present invention can be a method for providing an index for assisting in the diagnosis of enteritis in a subject, or a method for analyzing biogas in a subject suffering from enteritis, suspected of having enteritis, or at risk of suffering from enteritis. Furthermore, the method for assisting in the diagnosis of enteritis in a subject according to one aspect of the present invention can be performed in vitro without being a medical procedure.

[0038] According to another aspect of the present invention, there is provided a method for screening for a substance effective in treating enteritis, comprising the steps of: administering a test substance to a subject suffering from enteritis; comparing the concentration of hydrogen in the gas mixture exhaled from the subject prior to administration of the test substance with the concentration of hydrogen in the gas mixture exhaled from the subject after administration of the test substance; and selecting the test substance as a candidate substance effective for treating enteritis when the concentration of hydrogen in the gas mixture excreted from the subject after administration of the test substance is higher than the concentration of hydrogen in the gas mixture excreted from the subject before administration of the test substance. A method is provided that includes:

[0039] In one embodiment, the method of aiding in the diagnosis of enteritis in a subject or the screening method further comprises measuring the concentration of methanethiol (CH3SH) in the gas mixture excreted from the subject.

[0040] The concentration of methanethiol in the gas mixture excreted by the subject decreases as experimental colitis progresses and can therefore be used as a biomarker to improve the prediction of the onset of colitis.

[0041] In one embodiment, the reference value of the hydrogen concentration is the average value of the hydrogen concentration in the gas mixture discharged from the enteritis patient, the median value of the hydrogen concentration in the gas mixture discharged from the enteritis patient, a value lower than the average value or the median value, or a cutoff value of the hydrogen concentration in the gas mixture that distinguishes between the enteritis patient and the healthy subject; The standard value of the methanethiol concentration is the average value of the methanethiol concentration in the gas mixture discharged from the enteritis patient, the median value of the methanethiol concentration in the gas mixture discharged from the enteritis patient, the average value of the enteritis patients or a value higher than the median value, or a cutoff value of the methanethiol concentration in the feces for distinguishing between the enteritis patient and the healthy subject, The measured concentration of hydrogen in the gas mixture emitted from the subject is equal to or less than the reference concentration of said hydrogen, and When the measured value of the methanethiol concentration in the gas mixture discharged from the subject is equal to or lower than the reference value of the methanethiol concentration, it is determined that there is a high possibility of the subject suffering from enteritis.

[0042] In one embodiment, the reference value of the hydrogen concentration is the average value of the hydrogen concentration in the gas mixture discharged from the enteritis patient, the median value of the hydrogen concentration in the gas mixture discharged from the enteritis patient, a value lower than the average value or the median value, or a cutoff value of the hydrogen concentration in the gas mixture that distinguishes between the enteritis patient and the healthy subject; The standard value of the methanethiol concentration is the average value of the methanethiol concentration in the gas mixture discharged from the enteritis patient, the median value of the methanethiol concentration in the gas mixture discharged from the enteritis patient, the average value of the enteritis patients or a value higher than the median value, or a cutoff value of the methanethiol concentration in the feces for distinguishing between the enteritis patient and the healthy subject, A measured concentration of hydrogen in a gas mixture emitted from the subject is greater than a reference concentration of said hydrogen, and When the measured value of the methanethiol concentration in the gas mixture discharged from the subject is higher than the reference value of the methanethiol concentration, the possibility of suffering from enteritis is determined to be low.

[0043] In one embodiment, the standard value of the hydrogen concentration is the average value of the hydrogen concentration in the gas mixture discharged from a healthy subject, the median value of the hydrogen concentration in the gas mixture discharged from a healthy subject, or a value higher than the average value or the median value, and the standard value of the methanethiol concentration is the average value of the methanethiol concentration in the gas mixture discharged from a healthy subject, the median value of the methanethiol concentration in the gas mixture discharged from a subject, or a value lower than the average value or the median value; When the measured hydrogen concentration is equal to or greater than the standard hydrogen concentration value and the measured methanethiol concentration in the gas mixture discharged from the subject is equal to or greater than the standard methanol concentration value, the possibility of contracting enteritis is determined to be low.

[0044] In one embodiment, the standard value of the hydrogen concentration is the average value of the hydrogen concentration in the gas mixture discharged from a healthy subject, the median value of the hydrogen concentration in the gas mixture discharged from a healthy subject, or a value higher than the average value or the median value, and the standard value of the methanethiol concentration is the average value of the methanethiol concentration in the gas mixture discharged from a healthy subject, the median value of the methanethiol concentration in the gas mixture discharged from a subject, or a value lower than the average value or the median value; When the measured hydrogen concentration in the gas mixture discharged from the subject is lower than the standard value for the hydrogen concentration, and the measured methanethiol concentration in the gas mixture discharged from the subject is lower than the standard value for the methanethiol concentration, the possibility of contracting enteritis is determined.

[0045] Like hydrogen concentration, methanethiol concentration decreases with the progression of experimental colitis, and therefore can be used in steps similar to the above-mentioned comparison and determination steps using hydrogen concentration to determine the possibility of progression of colitis, to determine the therapeutic effect of a drug, and to select candidate substances effective in treating colitis in a screening method for substances effective in treating colitis.

[0046] In one embodiment, the method of aiding in the diagnosis of enteritis in a subject or the screening method further comprises measuring the concentration of hydrogen sulfide in the gas mixture excreted from the subject.

[0047] The concentration of hydrogen sulfide in the gas mixture excreted by the subject decreases as experimental colitis progresses and can therefore be used as a biomarker to improve the accuracy of predicting the onset of colitis.

[0048] In one embodiment, the reference value of the hydrogen concentration is the average value of the hydrogen concentration in the gas mixture discharged from the enteritis patient, the median value of the hydrogen concentration in the gas mixture discharged from the enteritis patient, a value lower than the average value or the median value, or a cutoff value of the hydrogen concentration in the gas mixture that distinguishes between the enteritis patient and the healthy subject; the standard value of hydrogen sulfide concentration is the average value of hydrogen sulfide concentration in the gas mixture discharged from enteritis patients, the median value of hydrogen sulfide concentration in the gas mixture discharged from enteritis patients, the average value of enteritis patients or a value higher than the median value, or the cutoff value of hydrogen sulfide concentration in feces for distinguishing between enteritis patients and healthy subjects; The measured concentration of hydrogen in the gas mixture emitted from the subject is equal to or less than the reference concentration of said hydrogen, and When the measured hydrogen sulfide concentration in the gas mixture discharged from the subject is equal to or lower than the reference value for hydrogen sulfide concentration, it is determined that the subject is highly likely to suffer from enteritis.

[0049] In one embodiment, the reference value of the hydrogen concentration is the average value of the hydrogen concentration in the gas mixture discharged from the enteritis patient, the median value of the hydrogen concentration in the gas mixture discharged from the enteritis patient, a value lower than the average value or the median value, or a cutoff value of the hydrogen concentration in the gas mixture that distinguishes between the enteritis patient and the healthy subject; the standard value of hydrogen sulfide concentration is the average value of hydrogen sulfide concentration in the gas mixture discharged from enteritis patients, the median value of hydrogen sulfide concentration in the gas mixture discharged from enteritis patients, the average value of enteritis patients or a value higher than the median value, or the cutoff value of hydrogen sulfide concentration in feces for distinguishing between enteritis patients and healthy subjects; A measured concentration of hydrogen in a gas mixture emitted from the subject is greater than a reference concentration of said hydrogen, and When the measured hydrogen sulfide concentration in the gas mixture discharged from the subject is higher than the reference value for the hydrogen sulfide concentration, the possibility of suffering from enteritis is determined to be low.

[0050] In one embodiment, the standard value of the hydrogen concentration is the average value of the hydrogen concentration in the gas mixture discharged from the healthy subject, the median value of the hydrogen concentration in the gas mixture discharged from the healthy subject, or a value higher than the average value or the median value, and the standard value of the hydrogen sulfide concentration is the average value of the hydrogen sulfide concentration in the gas mixture discharged from the healthy subject, the median value of the hydrogen sulfide concentration in the gas mixture discharged from the subject, or a value lower than the average value or the median value; When the measured hydrogen concentration is equal to or higher than the standard hydrogen concentration value and the measured hydrogen sulfide concentration in the gas mixture discharged from the subject is equal to or higher than the standard methanol concentration value, the possibility of contracting enteritis is determined to be low.

[0051] In one embodiment, the standard value of the hydrogen concentration is the average value of the hydrogen concentration in the gas mixture discharged from the healthy subject, the median value of the hydrogen concentration in the gas mixture discharged from the healthy subject, or a value higher than the average value or the median value, and the standard value of the hydrogen sulfide concentration is the average value of the hydrogen sulfide concentration in the gas mixture discharged from the healthy subject, the median value of the hydrogen sulfide concentration in the gas mixture discharged from the subject, or a value lower than the average value or the median value; When the measured hydrogen concentration in the gas mixture discharged from the subject is lower than the standard value for hydrogen concentration, and the measured hydrogen sulfide concentration in the gas mixture discharged from the subject is lower than the standard value for hydrogen sulfide concentration, the possibility of contracting enteritis is determined.

[0052] Like hydrogen concentration, hydrogen sulfide concentration decreases with the progression of experimental colitis. Therefore, in steps similar to the above-mentioned comparison and determination steps using hydrogen concentration, hydrogen sulfide concentration can also be used to determine the possibility of progression of colitis, to determine the therapeutic effect of a drug, and to select candidate substances effective in treating colitis in a screening method for substances effective in treating colitis.

[0053] In one embodiment, the method of aiding in the diagnosis of enteritis in a subject or the screening method further comprises measuring the concentration or amount of lipocalin-2 in the subject's faeces.

[0054] There is a strong negative correlation between the concentration of lipocalin-2 in the subject's feces and the concentration of hydrogen in the gas mixture excreted by the subject, and there is a strong negative correlation between the concentration of lipocalin-2 in the subject's feces and the concentration of methanethiol in the gas mixture excreted by the subject.

[0055] In one embodiment, the reference value is the average lipocalin-2 concentration in the feces of enteritis patients, the median lipocalin-2 concentration in the feces of enteritis patients, a value higher than the average or median, or a cutoff value of the lipocalin-2 concentration in the feces that distinguishes between enteritis patients and healthy subjects, When the measured concentration of lipocalin-2 in the subject's feces is equal to or higher than the reference value, the subject is judged to have a high possibility of suffering from enteritis.

[0056] In one embodiment, the reference value is the average lipocalin-2 concentration in the feces of enteritis patients, the median lipocalin-2 concentration in the feces of enteritis patients, or a cutoff value of the lipocalin-2 concentration in the feces that distinguishes between enteritis patients and healthy subjects, When the measured concentration of lipocalin-2 in the subject's feces is lower than the reference value, the possibility of the subject suffering from enteritis is determined to be low.

[0057] In one embodiment, the reference value is the average value of the lipocalin-2 concentration in the feces of healthy subjects, the median value of the lipocalin-2 concentration in the feces of patients with enteritis, or a value lower than the average value or the median value, When the measured concentration of lipocalin-2 in the subject's feces is equal to or lower than the reference value, the possibility of the subject suffering from enteritis is determined to be low.

[0058] In one embodiment, the average value of the lipocalin-2 concentration in the feces of healthy subjects or the median value of the lipocalin-2 concentration in the feces of patients with enteritis is used as a reference value, When the measured concentration of lipocalin-2 in the subject's feces is higher than the reference value, the possibility of the subject suffering from enteritis is determined to be high.

[0059] In one embodiment, the method of assisting in the diagnosis of enteritis in a subject or the screening method further comprises measuring the relative abundance of at least one enterobacteria selected from the group consisting of Akkermansiaceae, Rikenellaceae, and Tannerellaceae in the stool of the subject.

[0060] The present inventors have found that the relative abundance of these specific intestinal bacteria in the feces of subjects is correlated with the pathology of colitis and changes in hydrogen concentration. More specifically, the relative abundance of Akkermansiaceae and Rikenellaceae shows a positive correlation with lipocalin-2, and the relative abundance of Tannerellaceae shows a negative correlation with lipocalin-2. In addition, the relative abundance of Akkeransiaceae and Rikenellaceae shows a negative correlation with hydrogen concentration, and the relative abundance of Tannerellaceae shows a positive correlation with hydrogen concentration.

[0061] In one embodiment, the reference value is the average value of the relative amount of Akkermansiaceae in the feces of enteritis patients, the median value of the relative amount of Akkermansiaceae in the feces of enteritis patients, a value higher than the average value or the median value, or a cutoff value of the relative amount of Akkermansiaceae in the feces that distinguishes between enteritis patients and healthy subjects; When the measured relative amount of Akkermansiaceae in the subject's feces is equal to or greater than the reference value, the subject is judged to have a high possibility of suffering from enteritis.

[0062] In one embodiment, the reference value is the average value of the relative amount of Akkermansiaceae in the feces of enteritis patients, the median value of the relative amount of Akkermansiaceae in the feces of enteritis patients, or a cutoff value of the relative amount of Akkermansiaceae in the feces that distinguishes between enteritis patients and healthy people; When the measured relative amount of Akkermansiaceae in the subject's feces is lower than the reference value, the possibility of the subject suffering from enteritis is determined to be low.

[0063] In one embodiment, the reference value is the average value of the relative amount of Akkermansiaceae in the feces of healthy subjects, the median value of the relative amount of Akkermansiaceae in the feces of enteritis patients, or a value lower than the average value or the median value; When the measured relative amount of Akkermansiaceae in the subject's feces is equal to or lower than the reference value, the possibility of the subject suffering from enteritis is determined to be low.

[0064] In one embodiment, the average value of the relative amount of Akkermansiaceae in the feces of healthy subjects or the median value of the relative amount of Akkermansiaceae in the feces of enteritis patients is used as a reference value; When the measured relative amount of Akkermansiaceae in the subject's feces is higher than the reference value, the possibility of the subject suffering from enteritis is determined to be high.

[0065] In one embodiment, the reference value is the average value of the relative amount of Rikenellaceae in the feces of enteritis patients, the median value of the relative amount of Rikenellaceae in the feces of enteritis patients, a value higher than the average value or the median value, or a cutoff value of the relative amount of Rikenellaceae in the feces that distinguishes between enteritis patients and healthy subjects; When the measured relative amount of Rikenellaceae in the subject's feces is equal to or greater than the reference value, the subject is judged to have a high possibility of suffering from enteritis.

[0066] In one embodiment, the reference value is the average value of the relative amount of Rikenellaceae in the feces of enteritis patients, the median value of the relative amount of Rikenellaceae in the feces of enteritis patients, or a cutoff value of the relative amount of Rikenellaceae in the feces that distinguishes between enteritis patients and healthy subjects; When the measured relative amount of Rikenellaceae in the subject's feces is lower than the reference value, the possibility of the subject suffering from enteritis is determined to be low.

[0067] In one embodiment, the reference value is the average value of the relative amount of Rikenellaceae in the feces of healthy subjects, the median value of the relative amount of Rikenellaceae in the feces of enteritis patients, or a value lower than the average value or the median value; When the measured relative amount of Rikenellaceae in the subject's feces is equal to or lower than the reference value, the possibility of the subject suffering from enteritis is determined to be low.

[0068] In one embodiment, the average value of the relative amount of Rikenellaceae in the feces of healthy subjects or the median value of the relative amount of Rikenellaceae in the feces of enteritis patients is used as a reference value; When the measured relative amount of Rikenellaceae in the subject's feces is higher than the reference value, the possibility of the subject suffering from enteritis is determined to be high.

[0069] In one embodiment, the reference value is the average value of the relative amount of Tannerellaceae in the feces of enteritis patients, the median value of the relative amount of Tannerellaceae in the feces of enteritis patients, a value lower than the average value or the median value, or a cutoff value of the relative amount of Tannerellaceae in the feces that distinguishes between enteritis patients and healthy subjects; When the measured relative amount of Tannerellaceae in the subject's feces is equal to or lower than the reference value, the possibility of the subject suffering from enteritis is determined to be high.

[0070] In one embodiment, the reference value is the average value of the relative amount of Tannerellaceae in the feces of enteritis patients, the median value of the relative amount of Tannerellaceae in the feces of enteritis patients, or a cutoff value of the relative amount of Tannerellaceae in the feces that distinguishes between enteritis patients and healthy subjects; When the measured relative amount of Tannerellaceae in the subject's feces is higher than the reference value, the possibility of the subject suffering from enteritis is determined to be low.

[0071] In one embodiment, the reference value is the average value of the relative amount of Tannerellaceae in the feces of healthy subjects, the median value of the relative amount of Tannerellaceae in the feces of enteritis patients, or a value higher than the average value or the median value; When the measured relative amount of Tannerellaceae in the subject's feces is equal to or greater than the reference value, the possibility of the subject suffering from enteritis is determined to be low.

[0072] In one embodiment, the average value of the relative amount of Tannerellaceae in the feces of healthy subjects or the median value of the relative amount of Tannerellaceae in the feces of enteritis patients is used as a reference value; When the measured relative amount of Tannerellaceae in the subject's feces is lower than the reference value, the possibility of the subject suffering from enteritis is determined to be high.

[0073] In addition to measuring the concentration of hydrogen in the subject gas mixture, the accuracy of aiding in the diagnosis of enteritis can be improved by measuring the relative amounts of one, two or three species selected from Akkermansiaceae, Rikenellaceae, and Tannerellaceae in the stool of a patient with enteritis, comparing them with their respective standard values, and making a judgment.

[0074] The disclosures of all patent applications and publications cited herein are hereby incorporated by reference in their entireties.

[0075] The following examples are intended for illustrative purposes only and are not intended to limit the scope of the present invention in any way. Unless otherwise specified, reagents are commercially available or can be obtained or prepared according to conventional techniques or procedures known in the art. EXAMPLES

[0076] 1. Example 1-1.Animal testing All animal experiments were performed according to protocols approved by the Keio University Animal Care and Use Committee (approval number 17014-3). Mice were maintained in accordance with the Keio University Experimental Animal Guidelines and housed in an environment with a 12 / 12-hour light / dark cycle (lights on from 8:00 to 20:00), a temperature of 25 ± 2°C, and a humidity of 50 ± 5%. Six-week-old C57BL / 6J male mice were purchased from Sankyo Labo Service (Inasa Branch). After delivery, the mice were acclimated for 5 days on CE-2 (CLEA Japan) diet before being used in the experiments. Food and water were available ad libitum in all experiments.

[0077] 1-2.DSS / Dextran administration experiment DSS (MW: 36-50 kDa) (MP Biomedicals, Chiba, Japan) and Dextran (TCI, Tokyo, Japan) were each dissolved in RO water to a final concentration of 2% (w / v) and sterilized with a 0.22 μm pore size filter (Merk, Darmstadt, Germany). From the start of the experiment, 2% DSS solution or 2% Dextran solution was administered to the animals for 5 days, and then water was replaced for 10 days (see Figure 1).

[0078] 1-3. Measurement of biogas To analyze biogases, a gas chromatograph was installed in a mouse breeding cage and measurements were performed with the cooperation of Professor Ken Uchida and Assistant Professor Takahisa Tanaka (Uchida Laboratory, Department of Materials Engineering, Graduate School of Engineering, The University of Tokyo). As shown diagrammatically in Figure 2, ventilation of the air flow path connected between the cage 10 and the sensor gas chromatograph 14 was performed by a diaphragm pump DAP-15 (ULVAC KIKO) 12. The sensor gas chromatograph 14 is composed of a gas sampler, a gas column, and a semiconductor gas sensor. Biogases collected by the gas sampler were separated in the gas column, and gas detection with high sensitivity was performed.

[0079] The measurement interval of the gas concentration depended on the update time of the gas column and the gas sensor in the sensor gas chromatograph, and the average measurement interval was about 6 min. The measurement of the gas concentration was obtained from the whole gas in the cage, including the gas from the mouse's breath, the gas from the skin, and the gas from the feces.

[0080] The H2 and NH3 concentrations were measured using a sensor gas analyzer ODNA-P3-C (Nissha FIS), and the detection limits (LODs) were all 1000 ppb. The concentrations of H2S, CH3SH, and C2H5SH were measured using a sensor-type gas chromatograph ODSA-P3-A (Nissha FIS), and the LODs were 2 ppb, 5 ppb, and 5 ppb, respectively.

[0081] 1-4. Measurement of fecal lipocalin-2 Mouse feces stored at -80°C were weighed and suspended in phosphate-buffered saline (PBS) containing 0.1% Tween-20 at a concentration of 100 mg / mL. The suspension was vortexed for 20 minutes and centrifuged at 4°C and 13,000 × g for 10 minutes to separate the supernatant. The supernatant was diluted with PBS containing 1% bovine serum albumin (BSA). ELISA was performed according to the Mouse Lipocalin-2 / NGAL DuoSet ELISA (R&D Systems, USA) product protocol.

[0082] 1-5. Fecal DNA extraction and 16S rRNA sequencing DNA was extracted from mouse feces according to the protocol of the EZNA® Stool DNA kit (OMEGA), and then purified using magLEAD 12gc (Precision system science). The 16S rRNA gene V3-V4 region from the collected DNA sample was amplified.

[0083] The sequences of the primers used are as follows:

[0084] Forward: 5'- TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGCCTACGGGNGGCWGCAG - 3' Reverse: 5'-GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAGGACTACHVGGGTATCTAATCC - 3' The amplified products were subjected to PCR to add a unique index sequence to the DNA from each sample, and the library DNA was then purified and sequenced.

[0085] 1-6. Data analysis of 16S rDNA sequencing QIIME2 (ver. 2020.11) (Nat Biotechnol 37, 852-857 (2019), https: / / doi.org:10.1038 / s41587-019-0209-9) was used for the analysis. After removing primer sequences using the QIIME2 plugin cutadapt, quality control was performed using DADA2 (Nat Methods 13, 581-583 (2016), https: / / doi.org:10.1038 / nmeth.3869), paired-end reads were combined, chimera checks were performed, and an ASV table was created. For each representative sequence of ASV, the species was identified based on the database of SILVA ver.138 (BMC Bioinformatics 10, 421 (2009), https: / / doi.org:10.1186 / 1471-2105-10-421) using the BLAST plug-in (Nucleic Acids Res 35, 7188-7196 (2007), https: / / doi.org:10.1093 / nar / gkm864). After random sampling of 11,948 reads using feature-table (Microbiome 5, 27 (2017), https: / / doi.org:10.1186 / s40168-017-0237-y), the composition data was converted and diversity analysis was performed. Alpha diversity was analyzed using Observed ASVs and Shannon's Index, and beta diversity was analyzed using Unweighted-UniFrac distance (Frontier in Microbiology 2022 Mar 3;12:796025. doi:10.3389 / fmicb.2021.796025). Alpha diversity indicates the richness and evenness of the biological species that make up the ecosystem, while beta diversity indicates the degree of difference between ecosystems.

[0086] 1-7.Statistical analysis Statistical analysis was performed using GraphPad Prism version 9.5.0 (GraphPad Software Inc.). All time course data were examined for normality using the Shapiro-Wilk test, and the results showed that the distribution was non-normal. Intergroup significance was confirmed using the Friedman's test followed by Dunn's multiple comparison test.

[0087] The p-values ​​are indicated as *p < 0.05, **p < 0.01, ***p < 0.001, and the standard error is used for the error bars.

[0088] Correlation analysis was performed using RStudio (2022.07.1+554), and visualization was performed using GraphPad Prism version 9.5.0. Fecal lipocalin-2 was converted to common logarithm before use. H2 concentration was calculated as the average value for a total of 24 hours, 12 hours before and after the time of body weight measurement and fecal sample collection. Correlation analysis between the average body weight and H2 concentration, and the average lipocalin-2 and H2 concentration was performed on 6 samples at 6 points excluding days 0 and 1. Correlation analysis between body weight and lipocalin-2 values ​​was performed on 5 samples each at 6 points excluding days 0 and 1, for a total of 30 samples. All correlation analyses were performed using Pearson's product-moment correlation coefficient (PCC).

[0089] 2.Results 2-1.DSS induces experimental colitis accompanied by body weight loss and intestinal inflammation Using the DSS-induced colitis model, pathological changes after experimental colitis induction were evaluated. Dextran, a structural analogue of DSS, was administered as a negative control for DSS. Evaluation was performed using changes in mouse body weight and fecal lipocalin-2, an inflammatory marker. As a result, in the DSS-administered group, weight loss was observed from the 5th day, and recovery began from the 9th day onwards (Figure 3(A)). Fecal lipocalin-2 concentrations gradually increased from the first day of DSS administration, peaked on the 7th day, and remained high until the 15th day (Figure 3(B)). On the other hand, in the Dextran-administered group, no dramatic changes were observed in either body weight or fecal lipocalin-2 concentrations over the 15 days (Figures 3(C) and (D)). These results indicate that DSS induces weight loss and an increase in inflammatory markers in the feces, while Dextran does not cause such changes.

[0090] 2.2 DSS causes changes in biogas concentrations Next, to identify biogas components that could serve as biomarkers for IBD, we measured changes in biogas concentrations in mice with experimental colitis.

[0091] In the DSS-administered group, the H2 concentration transiently increased immediately after administration, decreased from the 4th day, reached its lowest value from the 7th to 8th days, and then gradually increased from the 8th day (Fig. 4(A)). On the other hand, in the Dextran-administered group, the H2 concentration transiently peaked on the 1st day, but then stably showed periodic peaks (Fig. 4(F)).

[0092] As shown in Figure 5(A), in the DDS-treated mice, the standard deviation of hydrogen concentration gradually decreased from the first day, reached a low value on the 6th to 8th days, and then showed an upward curve, whereas in the dextran-treated mice in Figure 5(B), the standard deviation of hydrogen concentration fluctuated periodically and showed a similar peak.

[0093] In the DSS-administered group, the NH3 concentration was stable at a high value until the 5th day, then it rapidly decreased after the 5th day when the DSS administration period ended, and increased from the 8th day, after which the concentration continued to fluctuate at a low level (Fig. 4(B)). On the other hand, in the Dextran-administered group, the NH3 concentration increased until the 3rd day, after which it showed a periodic peak state (Fig. 4(G)). In both the DSS-administered group and the Dextran-administered group, the NH3 concentration continued to increase during administration, but immediately returned to a low value every 24 hours. This rapid decrease is thought to be due to the opening of the lid for measurement.

[0094] Regarding changes in sulfur compound levels in the DSS-administered group, the concentrations of H2S, CH3SH, and C2H5SH all gradually increased from day 10 (Fig. 4(C)-(E)). Furthermore, the concentrations of CH3SH and C2H5SH reached their lowest values ​​between days 6 and 9 (Fig. 4(D), (E)). In the Dextran-treated group, although the concentration of C2H5SH showed a transient increase and decrease between days 2 and 5, the sulfur compound levels generally showed stable behavior (Fig. 4(H)-(J)).

[0095] Next, to examine the association with disease, a correlation analysis was performed between biogas and colitis phenotype parameters, namely, body weight change and fecal lipocalin-2 concentration. Since the biogas concentration reflects the total of five mice in a cage, the analysis was performed using the average biogas concentration per 24 hours per mouse and the fecal lipocalin-2 concentration of each mouse. The H2 concentration was positively correlated with body weight (Figure 4(K)), and the strongest negative correlation was observed with lipocalin-2 (Figure 4(L)). A negative correlation was also observed between CH3SH concentration and lipocalin-2 (Figure 4(L)). In addition, a correlation analysis was performed between the standard deviation of each biogas concentration and the fecal lipocalin-2 concentration, and the strongest negative correlation was found between the standard deviation of each of the H2 and CH3SH concentrations and lipocalin-2 (H2, correlation coefficient R = -0.83146319, p = 0.04021337, CH3HS, R = 0.85424181, p = 0.03031982). Next, to confirm the accuracy of predicting the onset of enteritis by biogas concentration measurements, ROC curves were created based on the measured values ​​of each biogas concentration in the DSS-administered group and the Dextran-administered group. As a result, H2 and H2S showed high AUC (Area under the curve) values ​​(Figures 6 (A) and (B)). From this, it is thought that both H2 and CH3SH are useful for predicting pathology (predicting the onset of disease and the effectiveness of treatment), and H2 and H2S are useful for predicting the onset of disease.

[0096] These results suggest that H2 concentration is most strongly correlated with the pathology of DSS-induced colitis, and that CH3SH and H2S concentrations also correlate with the pathology of DSS-induced colitis.

[0097] 2.3 DSS alters the intestinal microbiota It has been reported that the composition of biogas changes depending on the intestinal bacteria. We analyzed the changes in the intestinal microbiota when experimental colitis was induced by DSS. As a result, it was found that the α diversity of the intestinal microbiota gradually decreased after the induction of experimental colitis (Fig. 7(A)). In addition, the β diversity changed rapidly from day 0 to day 1, and then changed gradually (Fig. 7(B)). Further detailed analysis of the composition of the intestinal microbiota (Fig. 8(A)) showed that Akkermansiaceae and Rikenellaceae gradually increased from day 3 to day 7, when the fecal lipocalin-2 concentration began to increase (Fig. 8(B), (C)), while Tannerellaceae decreased from day 3 to day 7 (Fig. 8(D)).

[0098] 2.4 Relative abundance of Rikenellaceae and Akkermansiaceae is associated with disease outcome and H. 2 Strong correlation with concentration Based on the data obtained in the previous section, the correlation between the relative abundance of each intestinal bacteria and the body weight change, fecal lipocalin concentration, and H2 concentration in colitis was evaluated (Figure 9). As a result, the relative abundance of Akkermansiaceae and Rikenellaceae showed a positive correlation with lipocalin-2 and a negative correlation with body weight change. On the other hand, the relative abundance of Tannerellaceae showed a negative correlation with lipocalin-2 and a positive correlation with body weight change. In addition, the relative abundance of Tannerellaceae and Oscillospiraceae showed a positive correlation with H2 concentration, and Tannerellaceae in particular showed the strongest positive correlation. Conversely, the relative abundance of Akkermansiaceae and Rikenellaceae showed a negative correlation with H2 concentration. From the above, it was found that the relative abundance of specific intestinal bacteria showed opposite correlations with the progression of inflammation and H2 concentration, respectively (Figure 10).

[0099] In this study, the H2 concentration decreased with the progression of experimental colitis, suggesting that H2 concentration may be used as a biomarker to predict the onset of IBD. Furthermore, the relative abundance of specific gut bacteria correlated with the pathology of colitis and changes in H2 concentration, suggesting that they may mediate the negative correlation between the onset and progression of colitis and H2 concentration.

Claims

1. 1. A method for aiding in the diagnosis of enteritis in a subject, comprising: Measuring the concentration of hydrogen in the gas mixture emitted from the subject; and Comparing said measurements or the standard deviation of said measurements with a reference value. A method comprising:

2. 2. The method of claim 1, wherein the enteritis comprises inflammatory bowel disease.

3. The method of claim 1 , wherein the gas mixture exhaled by the subject comprises exhaled breath.

4. the reference value being an average value of the concentration of hydrogen in the gas mixture excreted from patients with enteritis, a median value of the concentration of hydrogen in the gas mixture excreted from patients with enteritis, a value lower than the average value or the median value, a cut-off value of the concentration of hydrogen in the gas mixture that distinguishes between patients with enteritis and healthy subjects, or a standard deviation of the concentration of hydrogen in the gas mixture excreted from patients with enteritis, The method according to any one of claims 1 to 3, further comprising determining that there is a high possibility of colitis when the measured hydrogen concentration or the standard deviation of the measured hydrogen concentration is equal to or less than the reference value.

5. The reference value is an average value of the concentration of hydrogen in the gas mixture exhaled from a healthy subject, a median value of the concentration of hydrogen in the gas mixture exhaled from a healthy subject, a value higher than the average value or the median value, or a standard deviation of the concentration of hydrogen in the gas mixture exhaled from a healthy subject; The method according to any one of claims 1 to 3, further comprising determining that the possibility of suffering from enteritis is low when the measured value of the hydrogen concentration or the standard deviation of the measured value is equal to or greater than the reference value.

6. the reference value being a value of the concentration of hydrogen in a gas mixture emitted from the subject, previously measured in the subject; The method according to any one of claims 1 to 3, further comprising determining that there is a high possibility that enteritis is progressing when a concentration of hydrogen in a gas mixture discharged from the subject measured at a certain time point in the subject is lower than the reference value, or determining that there is a high possibility that enteritis is not progressing when a concentration of hydrogen in a gas mixture discharged from the subject measured at a certain time point in the subject is equal to or higher than the reference value.

7. The subject is a patient with enteritis, the reference value being a measurement of the concentration of hydrogen in the gas mixture excreted from the subject prior to administration of a drug; measuring the concentration of hydrogen includes measuring the concentration of hydrogen in a gas mixture excreted from the subject after administering the drug; The method according to any one of claims 1 to 3, further comprising determining that the therapeutic effect of the drug is high when a measured value of a concentration of hydrogen in the gas mixture discharged from the subject after administration of the drug is higher than the reference value, or determining that the therapeutic effect of the drug is low when a measured value of a concentration of hydrogen in the gas mixture discharged from the subject after administration of the drug is equal to or lower than the reference value.

8. A method for screening for a substance effective in treating enteritis, comprising the steps of: administering a test substance to a subject suffering from enteritis; comparing the concentration of hydrogen in the gas mixture exhaled from the subject prior to administration of the test substance with the concentration of hydrogen in the gas mixture exhaled from the subject after administration of the test substance; and selecting the test substance as a candidate effective for treating enteritis when the concentration of hydrogen in the gas mixture exhaled from the subject after administration of the test substance is higher than the concentration of hydrogen in the gas mixture exhaled from the subject before administration of the test substance. The method further comprising:

9. 10. The method of claim 1 or 8, further comprising measuring a concentration of methanethiol in the gas mixture exhausted from the subject.

10. 10. The method of claim 1 or 8, further comprising measuring the concentration of hydrogen sulfide in the exhaust gas mixture of the subject.

11. The method of claim 1 or 8, further comprising measuring the concentration or amount of lipocalin-2 in the subject's stool.

12. 10. The method of claim 1 or 8, further comprising measuring the relative abundance of at least one enterobacteria selected from the group consisting of Akkermansiaceae, Rikenellaceae, and Tannerellaceae in the feces of the subject.

Images