CN112957376A - Microbial composition for regulating blood sugar of plateau population and application thereof - Google Patents
Microbial composition for regulating blood sugar of plateau population and application thereof Download PDFInfo
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Abstract
The invention relates to a microbial composition for regulating blood sugar of plateau population and application thereof, wherein the microbial composition comprises safe and effective amounts of two strains of Ackermanella (Akkermansia muciniphila) and Bacteroides thetaiotaomicron (Bacteroides thetaiotaomicron) and/or metabolites thereof. The invention researches the difference of the intestinal microbial flora of the plateau Han population and the plain Han population, finds that the difference of the two strains between the plateau Han population and the plain Han population is obvious, and can adjust the blood sugar of the plateau Han population in a targeted manner and stabilize the intestinal microecological environment at the same time by applying the microbial compound.
Description
Technical Field
The invention relates to a microbial composition for regulating blood sugar of plateau population and application thereof, belonging to the technical field of biological medicine.
Background
There is a large number of commensal flora in the human intestinal tract, which number exceeds 1000 trillion and is about 10 times of the total number of human cells. Meanwhile, the number of microbial genes in the intestinal tract is about 300 ten thousand, which is about more than 100 times of the number of human genome genes, so that the massive microbes form an inseparable symbiotic relationship with the human body, and people can be helped to adapt to changeable environments. The micro-ecological environment in the human body is closely related to the health of the human body, and the change of the micro-ecological environment can influence the clinical indexes of the human body. The maintenance of the stability of the micro-ecological environment in the human body is beneficial to the maintenance of the health of the human body. Can help to maintain the stability of intestinal micro-ecological environment, prevent and treat diseases and improve the function of human body by supplementing probiotics.
Chinese patent document CN110227085A (application number: CN201910639110.5) discloses Akkermanella (Akkermansia muciniphila) and application thereof, and the invention provides that the supplemented Akkermanella (Akkermansia muciniphila) strain has better anti-depression curative effect.
Chinese patent document CN104546935A (application number: CN201410522420.6) discloses Bacteroides thetaiotaomicron and application thereof, wherein the invention mentions that the Bacteroides thetaiotaomicron strain has better functions of reducing redness and swelling of joints and the like.
So far, no report that the intestinal tract is supplemented with two strains of Akkermansia muciniphila and Bacteroides theotiotamoxifen simultaneously to regulate the blood sugar of plateau population is found.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides a microbial composition for regulating the blood sugar of plateau people and application thereof.
The technical scheme of the invention is as follows:
a microbial composition for regulating blood glucose in plateau population, comprising a safe and effective amount of two species of Ackermanella (Akkermansia muciniphila) and Bacteroides thetaiotaomicron (Bacteroides thetaiotaomicron) and/or metabolites thereof.
Preferably, according to the present invention, the microbial composition further comprises a food or pharmaceutically acceptable carrier.
Preferably according to the present invention, the microbial composition is a food composition, a nutraceutical composition, a pharmaceutical composition, a beverage composition or a combination thereof.
Preferably, according to the present invention, the plateau population is a Han population.
Preferably, according to the invention, the microbial composition is an oral formulation,
preferably, the microbial composition is in the form of powder, tablet, sugar-coated agent, capsule, granule, suspension, solution, syrup, drop, sublingual tablet, or their combination.
According to the invention, the microbial composition is applied to regulating the blood sugar of plateau people.
Preferably, according to the present invention, the plateau population is a Han population.
A method for regulating blood sugar of plateau people is to apply the microbial composition to the plateau people.
Preferably, according to the present invention, the plateau population is a Han population.
The invention has the technical characteristics and beneficial effects that:
the invention researches the difference of the intestinal microbial flora of plateau Han population and plain Han population, finds that two strains of Akkermanella (Akkermansia muciniphila) and Bacteroides thetaiotaomicron (Bacteroides thetaiotaomicron) have obvious difference between the plateau Han population and the plain Han population, and the two strains are closely related to clinical indexes related to the blood sugar of the population, so the invention provides a microbial compound containing the two strains of the Akkermanella muciniphila (Akkermansia muciniphila) and the Bacteroides thetaiotaomicron (Bacteroides thetaiotaomicron), and by applying the microbial compound, the blood sugar of the plateau Han population can be pertinently regulated and the intestinal microecological environment can be stabilized.
Drawings
FIG. 1 is a box plot of the abundance of Akkermansia muciniphila species in the gut of three populations; in the figure, Han1000 is plain Han population, H4A4 is plateau Han population in one week, H4LB is plateau Han population more than two months;
FIG. 2 is a box plot of Bacteroides thetaiotaomicron strain abundance in the gut of three groups of people; in the figure, Han1000 is plain Han population, H4A4 is plateau Han population in one week, and H4LB is plateau Han population over two months.
FIG. 3 is a box plot of GLU (blood glucose), a clinical index for three groups of people; in the figure, Han1000 is plain Han population, H4A4 is plateau Han population in one week, and H4LB is plateau Han population over two months.
The specific implementation mode is as follows:
the present invention is further illustrated by the following examples, but the scope of the invention is not limited thereto.
The microorganisms involved in the invention are all the existing strains, can be purchased from the market, and do not relate to the preservation of the microorganisms.
Study subjects: according to STROBE statement design case contrast research, 45 Han male soldiers in China army from a certain month to a certain month in a certain year are taken as an experimental group 1 within one week of service at altitude of more than 3500 m plateau, 22 Han male soldiers in China at altitude of more than 3500 m plateau are taken as an experimental group 2, and 96 plain Han soldiers are matched according to the ages of the soldiers in the experimental group for contrast. Strictly controlling three groups of people to perform the same mixed diet of Chinese troops, keeping the same training environment and training intensity, and stopping smoking and drinking for 3 months. Soldiers with chronic inflammatory disease, oral antibiotics, acute infection and gastrointestinal disease were excluded.
After each study object is brought in, each study object receives a closestool excrement collector, excrement is put into an excrement collecting pipe after the excrement is discharged, the excrement is immediately stored in a refrigerator at the temperature of minus 20 ℃, excrement specimens are transferred to a refrigerator at the temperature of minus 80 ℃ for storage the next day, and the excrement specimens are uniformly detected.
Example 1 determination of differential flora between plains and plateau Hans populations
1.1 extraction of DNA from stool samples
Pipette 1000. mu.L of CTAB lysate into 2.0mL EP tube, add lysozyme, add 500. mu.L of fecal sample into lysate, water bath at 65 ℃ for 20min, mix several times during inversion to allow for sufficient lysis of the sample. The supernatant was centrifuged and an equal volume of phenol (pH8.0) was added: chloroform: isoamyl alcohol (25: 24:1 volume ratio) mixed solution is inverted and mixed evenly, centrifugation is carried out at 12000rpm for 10min, supernatant is taken, and chloroform with the same volume is added: the isoamyl alcohol (24:1 volume ratio) mixed solution is inverted and mixed evenly, centrifuged at 12000rpm for 10min, the supernatant is sucked into a 1.5mL centrifuge tube, added with 0.8 volume of isopropanol and shaken up and down, and precipitated at minus 20 ℃ for overnight. Centrifuge at 12000rpm for 10 minutes and pour out the liquid, taking care not to pour out the pellet. Washing the precipitate with 1mL of 75% ethanol for 2 times, and drying the precipitate on an ultra-clean bench or airing the precipitate at room temperature. The DNA sample was dissolved by adding ddH2O, and 1. mu.L of RNase A was added to digest the RNA and left at 37 ℃ for 15 min. And then, detecting the purity and concentration of the DNA by using agarose gel electrophoresis, taking a proper amount of sample DNA into a centrifugal tube, and diluting the sample to 1 ng/. mu.L by using sterile water.
1.2PCR amplification, sample mixing and purification
Genomic DNA diluted to 1 ng/. mu.L was used as a template, and specific primers 515F and 806R with Barcode, New England Biolabs, were selected from the sequencing region 16S V3-V4
And carrying out PCR by using a High-Fidelity PCR Master Mix with GC Buffer and High-efficiency and High-Fidelity enzyme to ensure the amplification efficiency and accuracy.
Detecting the PCR product by electrophoresis with 2% agarose gel; the PCR products were mixed in equal amounts according to the concentration of the PCR products, and after mixing well, the PCR products were purified by electrophoresis using 1 XTAE and 2% agarose gel, and the target band was recovered by cutting the gel. The product purification kit uses a recovery kit of GeneJET gel from Thermo Scientific company.
1.3 library construction and on-machine sequencing
The construction of the Library is carried out by using Ion Plus Fragment Library Kit 48rxns Library construction Kit of Thermo fisher company, and the constructed Library is subjected to on-machine sequencing by using Ion S5TMXL of Thermo fisher after the constructed Library is qualified through Qubit quantification and Library detection.
1.4 data analysis
1.4.1 sequencing data processing
Cutadapt (V1.9.1, http:// cutapt. readthetadocs. io/en/stable /) is used for carrying out low-quality partial shearing on Reads, then each sample data is split from the obtained Reads according to Barcode, the Barcode and primer sequence are cut off for preliminary quality control to obtain original data, the Reads obtained after the treatment needs to be treated for removing a chimera sequence, the Reads sequence is compared with a species annotation database to detect the chimera sequence, and the chimera sequence is finally removed to obtain final effective data.
1.4.2OTU clustering and species Annotation
All effective data of all samples are clustered by using Upearse software (Upearse v7.0.1001, http:// www.drive5.com/Uparse /), sequences are clustered into OTUs (operational Taxonomic units) by default with 97% consistency, representative sequences of the OTUs are selected at the same time, and the sequences with the highest frequency of occurrence in the OTUs are selected as the representative sequences of the OTUs according to the algorithm principle. And (3) performing species annotation on the OTUS sequences, performing species annotation analysis (setting a threshold value to be 0.8-1) by using a Usearch method and an rdp _16s _ sp.udb database, obtaining taxonomic information, and counting community compositions of all samples at the genus level respectively.
1.4.3 calculation of abundance and comparison of differences in the genus
The abundance of the genera was expressed as the Chao1 estimate and a box plot was plotted versus the difference in abundance of the related genera/species in the three groups of people, the difference comparison being statistically significant for P < 0.05 as the sample is non-normally distributed using the Kruskal-Wallis test.
1.5 sequencing results
Akkermansia muciniphila and Bacteroides theotiotamicron 2 strains in stool samples of the plain Han population and the plateau Han population show that the strains have no obvious difference in the three groups according to the Kruskal-Wallis value.
The results of the five-point scores of Akkermansia muciniphila species in three groups are shown in FIG. 1, and the Kruskal-Wallis values show that the species are not significantly different in all three groups. However, the comparison of median can find that the relative abundance of Akkermansia muciniphila strains is the highest in the Han population in plain, and is reduced after the Han population is in plateau, and the Kruskal-Wallis numerical value shows that the three groups of data have obvious difference;
the results of five-place results of Bacteroides thetaiotaomicron strains in three groups of people are shown in figure 2, the abundance of Bacteroides _ thetaiotaomicron strains in Han people within one week after the plateau is reduced relative to the abundance of strains in the Han people in the plain, and the abundance of Bacteroides _ thetaiotaomicron strains in Han people two months after the plateau is increased back and exceeds the abundance of strains in the Han people in the plain.
Example 2 determination of clinical indices in plain and plateau Han populations
The clinical index is measured from a blood sample. Blood samples were collected as morning fasting venous blood (8ml) using EDTA-K2. Blood samples were centrifuged at 4000r/min for 10min, plasma was separated, and stored at-80 ℃ to hospital 301 (Beijing, approximately 40 m above sea level) for assay. The 76 clinical trial items were determined using a hematology analyzer (Roche-cobas6000, usa) using 2 ml of plasma. In addition, 1.5ml of plasma was used for 11 enzyme-linked immunosorbent assays (ELISA) (expanded Bio, China). The clinical data cover 76 indexes in total.
The results of the five-point scores of GLU (blood glucose) in the clinical index are shown in FIG. 3. Kruskal-Wallis values show that GLU (blood glucose) levels are significantly different in three groups of people. The GLU (blood glucose) levels in chinese populations in plateau continue to decrease and are unstable relative to blood glucose levels in chinese populations in plain.
Example 3 relationship between flora and clinical indices
The R software is adopted to carry out the sperman correlation analysis, the correlation between the Akkermansia muciniphila and bacteria and the theotiotamicron 2 strains and the clinical index is researched, and the research result is shown in the table 1.
TABLE 1.2 Spearmaman correlation analysis results of strains with the clinical index GLU (glucose)
| Bacterial strain | GLU (glucose) |
| Akkermansia muciniphila | Positive correlation |
| Bacteroides thetaiotaomicron | Positive correlation |
It can be known from table 1 that two species of Akkermansia muciniphila and Bacteroides thetaiotaomicron are in positive correlation with GLU (glucose), that is, the two species of Akkermansia muciniphila and Bacteroides thetaiotaomicron can promote the increase of GLU (glucose) content in human body, and the composition of the two species of Akkermansia muciniphila and Bacteroides thetaiotaomicron is supplemented to help plateau han population to stabilize blood sugar and normal metabolism and simultaneously maintain the in vivo intestinal micro-ecological environment of the plateau han population to be stable.
Claims (9)
1. A microbial composition for regulating blood glucose in plateau population, comprising a safe and effective amount of two species of Ackermanella (Akkermansia muciniphila) and Bacteroides thetaiotaomicron (Bacteroides thetaiotaomicron) and/or metabolites thereof.
2. The microbial composition for regulating blood glucose in plateau population according to claim 1, further comprising a food or pharmaceutically acceptable carrier.
3. The microbial composition for regulating blood glucose in plateau population according to claim 1, wherein the microbial composition is a food composition, health composition, pharmaceutical composition, beverage composition or a combination thereof.
4. The microbial composition for regulating blood glucose in plateau population according to claim 1, wherein the microbial composition is an oral preparation; further preferably, the microbial composition is in the form of powder, tablet, sugar-coated agent, capsule, granule, suspension, solution, syrup, drop, sublingual tablet, or a combination thereof.
5. The microbial composition for regulating blood glucose in a plateau population as claimed in claim 1, wherein the plateau population is a Han population.
6. The use of the microbial composition of claim 1 for regulating blood glucose in a plateau population.
7. The use of claim 6, wherein the plateau population is a Han population.
8. A method for regulating blood glucose in a plateau population by administering the microbial composition of claim 1 to the plateau population.
9. The method of claim 8, wherein the plateau population is a Han population.
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| WO2019204985A1 (en) * | 2018-04-24 | 2019-10-31 | 深圳华大生命科学研究院 | Osteoporosis biomarker and use thereof |
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