CN112680500A

CN112680500A - Reagent and biomarker for detecting type 1 diabetes and application of reagent and biomarker

Info

Publication number: CN112680500A
Application number: CN202011606047.4A
Authority: CN
Inventors: 牟丽莎; 蔡志明; 曹梦涛; 彭园征; 陆赢; 陈姣; 蒲祖辉; 梁樱莹
Original assignee: Shenzhen Second Peoples Hospital
Current assignee: Shenzhen Second Peoples Hospital
Priority date: 2020-12-30
Filing date: 2020-12-30
Publication date: 2021-04-20

Abstract

The application discloses a reagent and a biomarker for detecting type 1 diabetes and application thereof. The reagent for detecting the type 1 diabetes mellitus is a reagent for detecting the abundance of intestinal microbiota of a to-be-detected object; wherein the gut microbiota comprises Peste des petits ruminants, Clerodendron, butyric acid bacilli, Oscillatoria, Dulbecco and Zuricobacter. The reagent for detecting the type 1 diabetes can effectively reflect the state of the type 1 diabetes by specifically detecting a plurality of key intestinal microbiota, thereby providing scientific reference basis for the detection and treatment effect of the type 1 diabetes.

Description

Reagent and biomarker for detecting type 1 diabetes and application of reagent and biomarker

Technical Field

The application relates to the technical field of diabetes detection, in particular to a reagent and a biomarker for detecting type 1 diabetes and application thereof.

Background

Type 1 diabetes (abbreviated T1DM) is a chronic disease in which autoimmunity destroys beta cells, resulting in little or no insulin production. The incidence of T1DM has increased over the last several decades. However, a number of factors, including genetic and environmental factors, play a crucial role in the pathophysiology of T1 DM. T1DM is not the only autoimmune disease with increased incidence in recent decades, and other immune-mediated diseases, including inflammatory bowel disease and allergic diseases, have also been shown to have increased incidence. Therefore, this increased incidence is not only due to genetic changes, but also environmental factors are one of the important influencing factors, which play a key role in triggering the imbalance of the immune system and in making the individual susceptible to autoimmunity.

In recent decades, the intestinal microbiota has changed dramatically due to changes in living conditions and increased use of antibiotics. Evidence from humans and animals suggests a close association between T1DM and intestinal microorganisms. Studies have shown that patients with T1DM have an abnormal intestinal microbiota compared to healthy individuals. Although gut microbes are important for the development of the mucosal immune system, the relationship of gut microbes to T1DM is not clear.

For some patients with T1DM, they do not have the ability to control blood glucose stably and have severe hypoglycemic episodes and hypoglycemic coma, and clinical islet transplantation is effective in controlling diabetes. Over 1500 diabetic patients showed lower risk and longer effective results after clinical islet transplantation than pancreas transplantation. Furthermore, pancreas transplantation carries significant surgical risks and has potentially serious complications and immunosuppression. The results of the three-phase clinical multicenter test also prove the safety and effectiveness of the clinical islet transplantation. Individuals with T1DM exhibit an abnormal gut microbiota, although pancreas or islet transplantation therapy is effective in controlling diabetes; however, it is not clear whether pancreas or islet transplantation therapy can restore abnormal gut microbiota.

In summary, although studies have shown that patients with T1DM exhibit an abnormal intestinal microbiota; however, the relationship between gut microbes and T1DM, and the effect of diabetes treatment on gut microbes, are not known.

Disclosure of Invention

The purpose of the application is to provide a novel reagent and a biomarker for detecting type 1 diabetes and application thereof.

In order to achieve the purpose, the application adopts the following technical scheme:

the first aspect of the application discloses a reagent for detecting type 1 diabetes, which is a reagent for detecting the abundance of intestinal microbiota of a subject to be detected; the intestinal microflora includes ruminant (ruminococcus), ozonobacterium (Odoribacter), butyric acid bacillus (Butyricicoccus), oscillatorium (Oscillibacter), Duboiella (Dubosiella) and Bacillus thuringiensis (Turcibacter).

The reagent can reflect the state of type 1 diabetes to a certain extent by detecting the abundance of intestinal microorganisms such as ruminant coccus, ozonobacterium, butyric acid bacillus, oscillatorium, dobesia and zurich bacillus, and provides a reference basis for the diagnosis and treatment effect of type 1 diabetes.

In one implementation of the present application, the gut microbiota further comprises bacteroides, firmicutes and melanin bacteria.

It should be noted that the research of the present application finds that the ruminant, the ozonobacterium, the butyric acid bacillus, the quiver bacillus, the dobbella and the zurich bacillus can be used as the biomarker of the type 1 diabetes; further, bacteroides, firmicutes and melanobacteria can also be used as biomarkers for type 1 diabetes; therefore, in the detection of type 1 diabetes, bacteroides, firmicutes and melanin bacteria can be further detected.

In one implementation of the application, the reagent comprises a nucleic acid combination or a polypeptide combination that detects the abundance of the gut microbiota to be described. Wherein, the nucleic acid combination can be a primer or a combination of a primer and a probe; the polypeptide combination may be a specific antibody or polypeptide.

In one implementation of the application, the reagent comprises a medium or combination of media to detect the abundance of the gut microbiota to be described. The culture medium or the culture medium combination can be a conventional microorganism quantitative detection realized by biological or chemical reaction.

It is important to note that the key to the present application is the inventive discovery that ruminants, ozonobacteria, butyric acid bacilli, treetobacter, dobbella and zurich bacilli, as well as bacteroides, firmicutes and melanobacteria, can serve as biomarkers for type 1 diabetes; as to how to specifically detect the abundance of these intestinal microbiota, reference may be made to the prior art, such as designing specific primers, probes, or specific antibodies, or other methods that can achieve quantitative detection of microorganisms, and are not specifically limited herein.

A second aspect of the application discloses the use of gut microbiota as a biomarker for the detection of type 1 diabetes or the prediction of diabetes status, wherein gut microbiota comprises ruminants, foetobacter, butyric acid bacilli, fibrillation bacilli, dipteria and zurich bacilli.

Preferably, for the application of the present application, the intestinal microbiota further comprises bacteroides, firmicutes and melanin bacteria.

A third aspect of the present application discloses a biomarker for detection of type 1 diabetes or prediction of diabetes status, the biomarker being gut microbiota including ruminant, ozonobacteria, butyric acid bacillus, fibrillation bacillus, dewarpilla and zurich bacillus.

Preferably, the gut microbiota in the biomarker further comprises bacteroides, firmicutes and melanin bacteria.

In a fourth aspect of the present application, the use of the biomarkers of the present application as targets for the treatment of diabetes is disclosed.

It will be appreciated that the gut microbiota of the present application is closely related to the state of diabetes and, therefore, these gut microbiota may also be targeted for the treatment of diabetes.

The fifth aspect of the application discloses application of the biomarker in preparation of a type 1 diabetes detection reagent, preparation of a diabetes treatment drug or development of a diabetes treatment drug.

The application of the reagent in preparing the type 1 diabetes mellitus detection reagent mainly refers to a detection reagent for specifically detecting the abundance of the intestinal microbiota serving as the biomarker, such as a primer, a probe, a polypeptide and the like. The application in preparing the diabetes treatment medicine mainly refers to the preparation of corresponding medicines by directly taking the intestinal microbiota as a target. The application in the development of the diabetes treatment drug mainly means that the effect of the drug on diabetes is evaluated by comparing and analyzing the abundance changes of the intestinal microbiota before and after the drug is used, so that reference is provided for the treatment effect, and the drug screening and development effects are further realized.

Due to the adoption of the technical scheme, the beneficial effects of the application are as follows:

the reagent for detecting the type 1 diabetes can effectively reflect the state of the type 1 diabetes by specifically detecting a plurality of key intestinal microbiota, thereby providing scientific reference basis for the detection and treatment effect of the type 1 diabetes.

Drawings

FIG. 1 is a time chart of stool samples collected from five groups of mice in the examples of the present application;

FIG. 2 is a fluorescent microscope observation result after the insulin staining in the example of the present application;

FIG. 3 is a graph of insulin concentrations measured by ELISA in mice in the examples of the present application;

FIG. 4 shows the results of blood glucose concentration tests of five groups of mice treated in the present example;

FIG. 5 shows the results of the weight test of five groups of mice treated in the present example;

FIG. 6 is a graph showing the results of pancreatic islet transplantation in the Homo-Tx group and the Allo-Tx group according to the present example;

FIG. 7 is a graph showing the results of fecal microbiological analysis of mice in the diabetic group and the control group in the examples of the present application;

FIG. 8 is a graph showing the quantitative analysis of the types of microorganisms detected in the feces of mice in the diabetic group and the control group in the examples of the present application;

FIG. 9 is a graph showing the results of analysis of relative abundance of fecal microorganisms at the phylum level in mice of the diabetic group and the control group in the examples of the present application;

FIG. 10 is a graph showing the results of analysis of the abundance of Bacteroides and Myxoplasma in mouse feces of the diabetic group and the control group according to the example of the present application;

FIG. 11 is a graph showing the results of analysis of the abundance of microorganisms in mouse feces of the diabetic group and the control group according to the example of the present application;

FIG. 12 is a graph showing the results of microbiological analysis of stool samples from five groups of mice in the examples of the present application;

FIG. 13 is a graph showing the quantitative analysis of the types of microorganisms detected in fecal samples of five groups of mice according to the example of the present application;

FIG. 14 shows the results of relative abundance analysis of microorganisms in stool samples of five groups of mice in examples of the present application;

FIG. 15 shows the results of analysis of the relative abundance of Bacteroides, Mycobacteria and melanocytes in stool samples from five groups of mice in the examples of the present application;

FIG. 16 shows the analysis results of the relative abundance of ruminant bacteria in stool samples of five groups of mice according to the example of the present application;

FIG. 17 shows the results of analysis of the relative abundance of Oesophagostomum in fecal samples from five groups of mice in the examples of the present application;

FIG. 18 shows the analysis results of the relative abundance of butyric acid bacteria in fecal samples of five groups of mice according to the example of the present application;

FIG. 19 shows the analysis results of relative abundance of Oscillatoria in stool samples of five groups of mice in the examples of the present application;

FIG. 20 shows the analysis results of the relative abundance of Dunaliella in stool samples of five groups of mice according to the example of the present application;

FIG. 21 shows the results of relative abundance analysis of Bacillus thuringiensis from stool samples of five groups of mice according to the examples of the present application.

Detailed Description

Existing studies have shown differences between the gut microbiota of patients with T1DM and healthy persons, but, in particular, the relationship of gut microbiota to T1DM is not clear, nor is the effect of diabetes treatment on gut microbiota clear.

The inventive discovery of this application suggests that specific gut microbiota including ruminants, ozonobacteria, butyrobacterium, oscillatorium, dewarpia and zurich can be used as biomarkers for type 1 diabetes. Further, these gut microbiota may also include bacteroides, firmicutes and melanin bacteria. The state of the type 1 diabetes can be reflected to a certain extent by detecting the intestinal microbiota, and a reference basis is provided for the detection and treatment effect of the type 1 diabetes.

The present invention will be described in further detail with reference to the following detailed description and accompanying drawings. The following examples are intended to be illustrative of the present application only and should not be construed as limiting the present application. Unless otherwise specified, the instruments and materials used in the following examples are the ones conventionally used in laboratories.

Examples

Materials and methods

1. Test animal

Wild type male C57BL/6 and Balb/C mice were purchased from the Guangdong medical laboratory animal center for 6-8 weeks. Only male mice were used in this study, as glucose stimulated insulin release was independent of sex. All mice were bred under Specific Pathogen Free (SPF) conditions at shenzhen hospital, beijing university. The animal experimental scheme is approved by the ethical committee of biomedical research of Shenzhen hospital institution of Beijing university.

2. Induction of diabetes in mice

Streptozotocin (STZ, Sigma, St.Louis, Mo., USA) was dissolved in 0.5M sodium citrate buffer, and the single-use intraperitoneal injection dose of C57BL/6 mice was 250 mg/kg. Two days after injection, the blood glucose level of the mice was measured, and mice with blood glucose level higher than 16.8mmol/L and kept stable for 5 days were defined as diabetic mice.

The experiments were divided into five groups: control group (Ctrl group), normal C57BL/6 mice, without STZ treatment; (ii) diabetic group (DM group), STZ-induced diabetic mice; (III) Insulin treatment group (Insulin group), STZ induced diabetic mice, each injected with 0.8U Insulin (Wanbang, Jiangsu, China), and blood glucose was tested after half an hour after each Insulin injection. Within 6 hours after the insulin injection of the mice, the blood sugar reaches 11.1 mmol/L; (IV) isogenic transplantation group (Homo-Tx group), STZ induced C57BL/6 diabetic mice transplanted C57BL/6 mouse islets; (V) allogenic-Tx group, STZ-induced C57BL/6 diabetic mice transplanted Balb/C mouse islets without immunosuppressive agents.

3. Islet isolation, purification and culture

Islets were isolated from anesthetized C57BL/6 or Balb/C donor mice and perfused in situ through common bile duct with collagenase type V (Sigma-Aldrich, USA) at 2mg/mL into the pancreas. The perfused pancreas was removed and further digested with additional 2mg/mL collagenase type V in a 37 ℃ standing water bath in a water bath. The enzyme digestion was then stopped by briefly shaking with a vortex shaker, placing on ice, and then washing. Islets were purified by centrifugation on a discontinuous gradient (density 1.119, 1.083 and 1.077g/mL) Histopaque cell isolate (Sigma-Aldrich, USA). Islets were manually picked from digested pancreatic tissue under a dissecting microscope. The islets were then brought to 37 deg.C, 95% air and 5% CO₂The culture was performed in an incubator of 5mL of complete medium per dish (60mm dish) at a density of about 150 islets per dish.

4. Islet cell viability assessment

Fluoroscein Diacetate (FDA) (Sigma-Aldrich, USA) 1. mu.L and Propylidine Iodide (PI) (Sigma-Aldrich, USA) 10. mu.L were added to 1mL of PBS. Mixing with islets, and examining the islets with a fluorescence microscope.

5. Glucose stimulated insulin secretion test

In this experiment, 10 islets of similar size were manually selected, 95% air and 5% CO at 37 ℃₂The incubator of (2) for cultivation. After 30min pre-incubation, islets were incubated with Kreb's solution containing low glucose (2.8mM) or high glucose (28mM) for 60 min. The insulin concentration in the islet culture was determined by mouse insulin ELISA (MercodiaAB, Sweden).

6. Islet transplantation

Approximately 300 islets were transplanted under the renal capsule of each diabetic recipient. A small pocket is made at one end of the kidney. The islets are sucked into the tip of the lance head. The tip is inserted into the space between the renal capsule and the kidney and the islets are slowly expelled as the tip is withdrawn. The kidney is placed back into the abdominal cavity and the wound is then sutured closed.

7. DNA extraction from stool specimens and 16S rRNA sequences

Stool specimens were collected from each group of mice according to a schedule, as shown in fig. 1. In general, fecal pellets from the allogeneic-transplant group were collected at 2 weeks after islet transplantation, when rejection is occurring. The other groups of fecal pellets were harvested at week 4 after islet transplantation. Fecal DNA extraction kits (Tiangen Biotechnology, China) were used to isolate fecal specimen DNA according to standard procedures. The V3-V4 region of the bacterial 16S rRNA gene was PCR amplified with the following primers: 338F 5 '-barcode-ACCTAC GGAGGA GCAGCA-3' and 806R 5 '-GGACTACHVGGTWTC TAAT-3', wherein the barcode is an 8 base sequence unique to each sample. Sequencing was performed using illumina miseq platform.

8. Histological analysis

Diabetic mice transplanted with Balb/c mouse islets were sacrificed at week 2 and all other mice were sacrificed at week 4. The kidney containing the graft was fixed in 4% paraformaldehyde solution for at least 48h, paraffin sections were cut at 4 μm, stained with hematoxylin and eosin, and the graft size and envelope thickness were determined. Sections were used for immunohistochemical staining using an anti-insulin antibody (CST, usa) and an AlexaFluor 546-labeled donkey anti-goat IgG secondary antibody (Jackson ImmunoResearch, usa). Sections were stained with 300nM DAPI (ThermoFisher science, USA) in PBS, incubated for 20min, washed three times with PBS, and then photographed with a LeicaFDM2500 microscope.

9. Statistical analysis

The experiment was performed with GraphPad software (7 th edition) to analyze differences between groups using a two-tailed t-test. Statistical significance was defined as P < 0.05. P <0.05, p <0.01, p < 0.001.

Second, result in

1. Islet transplantation and insulin treatment for normalization of STZ-induced hyperglycemia in diabetic mice

In order to study the changes of intestinal microorganisms in diabetic mice and whether hyperglycemia treatment by insulin or pancreatic islets affects the intestinal microorganisms, five experimental groups were designed: (i) control (Ctrl group), i.e., control, normal C57BL/6 mice, without STZ treatment; (ii) diabetic group (DM group), STZ-induced diabetic mice; (iii) in the Insulin group, STZ induced diabetic mice, each mouse was injected with 0.8U of Insulin (Jiangsu Wanbang, China), and blood glucose was tested half an hour after each injection. Mice reached glucose levels <11.1mmol/L within 6 hours after insulin injection; (iv) group of homograft (Homo-Tx), STZ induced C57BL/6 diabetic mice transplanted C57BL/6 islets; (v) allograft (Allo-Tx group), STZ induced the transplantation of islets from C57BL/6 diabetic mice into Balb/C mice without immunosuppression, as shown in FIG. 1. The islet viability is above 95%, the stimulation index is above 2, and the islet transplantation is performed as shown in fig. 2 and fig. 3. Diabetic mice had elevated blood glucose levels, above 30mmol/L, and both insulin and islet transplantation showed the ability to lower blood glucose to normal levels, as shown in FIG. 4. The body weight of the diabetic mice was significantly reduced compared to the control mice. However, insulin or islet treatment may eliminate the decrease in body weight, as shown in fig. 5. These results indicate that islet transplantation or insulin treatment can correct STZ-induced hyperglycemia. Immunohistochemistry was also performed in this experiment, and it was found that transplanted islets remained morphologically intact in the kidney and expressed secreted insulin, as shown in fig. 6.

2. Diabetic mice display a similar bacterial diversity as non-diabetic mice

Stool samples were collected from the above five groups according to the schedule for this experiment, and bacterial DNA was isolated using the TIANAmp Stool DNA kit and 16S rRNA sequencing was performed to analyze symbiotic microflora as shown in FIG. 1. The results show that the fecal bacterial composition, and in particular the relative proportions, are different in the diabetic mice compared to the control non-diabetic mice, as shown in figure 7; but the two groups were of similar bacterial species, as shown in figure 8. The relative abundance of fecal microorganisms at the portal level showed differences between the two groups, as shown in figure 9. Diabetic mice had a significant reduction in bacteroides, and a significant increase in firmicutes, as compared to control mice, as shown in figure 10. The findings of this trial are consistent with the change in the values of firmicutes and bacteroides in patients with T1 DM. By more in-depth analysis, the present trial found that many genus bacteria in diabetic mice were altered compared to the non-diabetic control group, which further indicated a higher degree of gut microbiota dysbiosis, as shown in fig. 11. The changes for the first ten genera of bacteria are as follows: ectopic propionibacterium, lactobacillus, aldehydis, unidentified streptomycete, osmidobacter, bacteroides, unidentified ruminant coccus, helicobacter pylori, vibrio desulfovibrio, and dobesia.

3. Effects of islet transplantation and insulin treatment on microbiota

There is a large body of evidence that the composition of the gut microbiota in the development of diabetes has relevant characteristics; however, previous studies have not investigated the relationship between the microbiota of diabetic mice and islet transplantation. Thus, the assay performed 16S rRNA sequencing to analyze these changes. The results show that islet transplantation at least partially restores the dysregulated microbial composition found in diabetic mice, as shown in figure 12. The observed species were not different between groups as shown in figure 13. Insulin or islet treatment partially rescued bacteroides depletion, and insulin or islet treatment also mitigated firmicutes and melanobacteria increase, as shown in fig. 14 and 15. The data from this trial indicate that insulin or islet transplantation treatment can partially restore the disregulated bacteria, however, the commensal bacteria are still different from those of the control mice.

4. The symbiotic flora can be used as biomarker for prognosis of STZ-induced diabetes

Further analysis was performed at the genus level based on 16S rRNA sequencing, which indicated that many bacteria from different phyla were significantly increased in diabetic mice and recovered after diabetic mice received daily insulin treatment or islet transplantation treatment. The assay was performed on ruminants, ozonobacteria, butyrobacterium, oscillatoria, dobberella and zurich and analyzed for their relative abundance in the treated group compared to the untreated group as shown in fig. 16 to 21. Therefore, these commensal bacteria can serve as a valuable biomarker for the prognosis of STZ-induced diabetes.

Third, discussion and conclusion

The gut microbiota is very important for both humans and animals, and they are involved in the development of disease and influence its development. There is currently no study on changes in gut microbiota in diabetic mice treated with insulin or islet transplantation. In the study of this experiment, DNA of intestinal microbiota was isolated from diabetic mice treated with insulin or islet transplantation and sequenced using 16S rRNA. The results showed that diabetic mice did present with intestinal microbial abnormalities including an increase in firmicutes and a decrease in bacteroides, etc., as was the case with T1DM patients and diabetic rats. The insulin or islet transplantation therapy can partially recover abnormal microorganisms, and is beneficial to recovery of intestinal microorganisms. The abundance of melanin bacteria in the islet transplantation group was lower than that in the insulin group. It has also been found that certain bacteria, including ruminants, ozonobacteria, butyrobacterium, quiverful, dobbella and zurich, can be used as biomarkers for STZ-induced diabetes, mimicking human diabetes.

To study changes in microflora, diabetic mice were transplanted with islets from C57BL/6 mice or Balb/C mice. Islet transplantation in Balb/C mice maintained normoglycemia for 2 weeks prior to rejection by the immune system in C57BL/6 recipient mice. Although the time of sample collection varied between the transplanted groups of C57BL/6 or Balb/C mice, the intestinal microflora of diabetic mice after islet insulin was similar, unlike diabetic mice treated with insulin. This result indicates that immune rejection of the transplanted organ or cell does not interfere with intestinal microorganisms. Transplantation of islets into a recipient may affect the gut microbiota in other ways besides secreting insulin. However, restoring blood glucose levels is not sufficient to completely rescue the abnormal intestinal microorganisms.

Since a correlation of gut microbiota with STZ-induced diabetes development was observed in this study, this experiment suggests that bacteroides, firmicutes and melanin bacteria may also be biomarkers. Previous studies have shown that firmicutes and bacteroides are associated with obesity and diabetes. Firmicutes are defined as "fatty bacteria" and bacteroides as "lean bacteria". With increasing firmicutes/bacteroides ratio, the inflammatory response and BMI levels rise, and insulin resistance is more likely to develop, which will ultimately lead to the development of diabetes. Previous studies have not reported the association of melanocytes with diabetes.

This experiment also found a panel of commensal microbiota including ruminant, ozonella, butyric acid, oscillatoria, Debrella and Zuricella, isolated from the control and insulin/transplant groups. Among these specific bacteria, the ruminant family has been shown to be positively associated with diabetes, which may be associated with infection and inflammation. Since butyric acid bacteria secrete butyric acid and increase insulin sensitivity, they are negatively associated with Low Density Lipoprotein (LDL), blood Glucose (GLU), Uric Acid (UA), Total Cholesterol (TC), Body Mass Index (BMI), and Diabetes Mellitus (DM). Other bacteria including Bacillus thuringiensis, Clerodendrium and Oscillatoria have not been reported to be associated with diabetes. Previous studies have shown that, with respect to bacillus thuringiensis, it can reduce susceptibility to salmonella infection. In studies based on a model of inflammation tumorigenesis, abundant malodorous bacteria were detected. In patients with atrial fibrillation, there is a dramatic decrease in the number of oscillators.

Among the above bacteria, some specific bacteria were significantly increased in diabetic mice, including bacteroides, osmidium, and butyric acid coccus. However, insulin or islet transplantation can restore these bacteria to normal levels. It was also found in this trial that some bacteria were reduced in diabetic mice, including Dulbecco's bacteria and Bacillus thuringiensis. Also, insulin or islet transplantation restores the abundance of these reduced bacteria. Thus, these bacteria are biomarkers for monitoring islet transplantation function or the effectiveness of insulin therapy.

This study showed that the STZ-induced intestinal microbiota of diabetic mice had significantly changed and that insulin and islet transplantation treatments could partially correct the dysregulated microbiota. In STZ-induced diabetic mice, the abundance of some bacteria was up-regulated, including fetobacter, butyrobacter; while other bacteria, including Dulbecco and Bacillus thuringiensis, are down-regulated in abundance. However, insulin or islet transplantation restored these bacteria to normal levels. Thus, these gut microbiota may also be targets for the treatment of diabetes. This trial suggests that the microbiota directed against diabetic patients may further enhance the beneficial metabolic effects obtained by insulin treatment or islet transplantation. Therefore, these specific bacteria can be used as biomarkers for predicting the diabetic state of diabetic mice.

In summary, the intestinal microbiota of the ruminants, the fetobacter, the butyric acid bacilli, the oscillatoria, the dobbella, the zurich bacilli, the bacteroides, the firmicutes and the melanin bacteria can be used as biomarkers of type 1 diabetes, and the state of type 1 diabetes can be detected by detecting the abundance of the intestinal microbiota, so that a reference basis is provided for the detection and treatment effect of type 1 diabetes.

The present invention has been described in terms of specific examples, which are provided to aid understanding of the invention and are not intended to be limiting. For a person skilled in the art to which the invention pertains, several simple deductions, modifications or substitutions may be made according to the idea of the invention.

Claims

1. A reagent for detecting type 1 diabetes, comprising: the reagent is used for detecting the abundance of the intestinal microbiota of the object to be detected; the gut microbiota includes ruminant, ozonobacteria, butyric acid bacillus, quiver bacillus, Dulbengiella and Zuricella.

2. The reagent according to claim 1, characterized in that: the gut microbiota further comprises bacteroides, firmicutes and melanin bacteria.

3. The reagent according to claim 1 or 2, characterized in that: the reagent comprises a nucleic acid combination or a polypeptide combination for detecting the abundance of the intestinal microbiota to be detected.

4. The reagent according to claim 1 or 2, characterized in that: the reagents include a medium or combination of media to detect the abundance of the gut microbiota.

5. Use of the gut microbiota as a biomarker for the detection of type 1 diabetes or the prediction of the diabetic status, characterized in that: the gut microbiota includes ruminant, ozonobacteria, butyric acid bacillus, quiver bacillus, Dulbengiella and Zuricella.

6. Use according to claim 5, characterized in that: the gut microbiota further comprises bacteroides, firmicutes and melanin bacteria.

7. A biomarker for detection of type 1 diabetes or prediction of diabetes status, characterized by: the biomarker is gut microbiota including ruminant, ozonobacteria, butyric acid bacillus, oscillatoria, dewarpiella and zurich bacillus.

8. The biomarker of claim 7, characterized in that: the gut microbiota further comprises bacteroides, firmicutes and melanin bacteria.

9. Use of a biomarker according to claim 7 or 8 as a target for the treatment of diabetes.

10. The use of the biomarker according to claim 7 or 8 in the preparation of a type 1 diabetes detection reagent, the preparation of a diabetes treatment drug, or the development of a diabetes treatment drug.