CN113025527A - Milk-derived streptococcus salivarius for predicting infant obesity and separation method thereof - Google Patents
Milk-derived streptococcus salivarius for predicting infant obesity and separation method thereof Download PDFInfo
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Abstract
The invention provides milk-derived streptococcus salivarius for predicting infant obesity and an isolation method thereof. The milk-derived Streptococcus salivarius is Streptococcus salivarius strain SHDP5A1, the 16SrRNA gene sequence is shown as SEQ ID NO:1, the milk-derived Streptococcus salivarius is classified and named as Streptococcus salivarius, the preservation number is CCTCC NO: M2021060, the preservation date is 2021, 01, 14 days, the preservation unit is China center for type culture preservation, and the preservation unit address is Wuhan university, Wuhan City, China. The milk-derived streptococcus salivarius for predicting the obesity of the infants is used as facultative anaerobe, is easy to colonize in intestinal tracts, creates conditions for maturation and stability of later-stage intestinal flora, and can predict the obesity of offspring by detecting the abundance of streptococcus salivarius in the early life stage.
Description
Technical Field
The invention relates to the technical field of biomedicine, in particular to milk-derived streptococcus salivarius for predicting infant obesity and a separation method thereof.
Background
Childhood obesity is currently a common metabolic disease in the clinic, manifested by body weight beginning to deviate from the normal growth curve during infancy. An increase in the prevalence of obesity in children can lead to an increase in the prevalence of chronic diseases in adulthood, such as impaired glucose tolerance and hypertension. Therefore, it is necessary to predict obesity after birth of infants.
The obesity of offspring is under the combined action of genetic factors and environmental factors, the genetic factors determine the genetic action of an obesity gene through the discovery of certain genes (FTO genes), and the environmental factors possibly have more obvious effect on the obesity of the offspring and are the most widely researched at present, such as diet, living environment, exercise, early life nutritional state, intestinal flora and the like. But the prediction effect of a certain key index on the offspring obesity is lacked at present.
The nutrition source of the offspring in the early life is breast milk firstly, and the colonization of flora in the breast milk in the offspring intestinal tract plays an important role in nutrition absorption and metabolism, but the prediction effect on the offspring obesity by sequencing the flora in the early life is lacked in the clinical application at present, and the separation of probiotics in the breast milk is mainly bifidobacteria and lactobacilli in the traditional sense, and the bifidobacteria and the lactobacilli have no relevance to the offspring obesity. Streptococcus salivarius is an important early-life colonization bacterium, and thermophilic subspecies of streptococcus salivarius are currently applied to dairy fermentation and industrial production, and are rarely researched in breast milk.
Disclosure of Invention
The invention aims to overcome the defects in the prior art and provide milk-derived Streptococcus salivarius (Streptococcus salivarius) for predicting infant obesity and an isolation method thereof.
The invention adopts the following technical scheme for solving the technical problems:
in a first aspect of the present invention, there is provided a milk-derived streptococcus salivarius for predicting obesity in infants, wherein the 16S rRNA gene sequence is as follows:
AGGACGAACGCTGGCGGCGTGCCTAATACATGCAAGTAGAACGCTGAAGAGAGGAGCTTGCTCTTCTTGGAAGAGTTGCGAACGGGTGAGTAACGCGTAGGTAACCTGCCTTGTAGCGGGGGATAACTATTGGAAACGATAGCTAATACCGCATAACAATGGGTGACCCATGTCATTCATTTGAAAGGGGCAACTGCTCCACTACAAGATGGACCTGCGTTGTATTAGCTAGTAGGTGAGGTAACGGCTCACCTAGGCGACGATACATAGCCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCGGCAATGGGGGCAACCCTGACCGAGCAACGCCGCGTGAGTGAAGAAGGTTTTCGGATCGTAAAGCTCTGTTGTAAGTCAAGAACGAGTGTGAGAGTGGAAAGTTCACACTGTGACGGTAGCTTACCAGAAAGGGACGGCTAATTACGTGCCCAGCAGCCGCGGTAATACGTAGGTCCCGAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGTTTGATAAGTCTGAAGTTAAAGGCTGTGGCTCAACCATAGTTCGCTTTGGAAACTGTCAAACTTGAGTGCAGAAGGGGAGAGTGGAATTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAGGAACACCGGTGGCGAAAGCGGCTCTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAGGTGTTGGGTCCTTTCCGGGATTCAGTGCCGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCCGATGCTATTTCTAGAGATAGGAAGTTACTTCGGTACATCGGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCCTATTGTTAGTTGCCATCATTCAGTTGGGCACTCTAGCGAGACTGCCGGTAATAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGTTGGTACAACGAGTTGCGAGTCGGTGACGGCGAGCTAATCTCTTAAAGCCAATCTCAGTTCGGATTGTAGGCTGCAACTCGCCTACATGAAGTCGGAATCGCTAGTAATCGCGGATCAGCACGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTATGGAGCCAGCCGCCTAAGGTGGGATAGATGATTGGGGTGAAGT(SEQ ID NO:1)。
furthermore, the milk-derived Streptococcus salivarius is Streptococcus salivarius strain SHDP5A1 which is named as Streptococcus salivarius salinivirus in a classification way, the preservation number is CCTCC NO: M2021060, the preservation date is 2021, 01, 14 days, the preservation unit is China center for type culture Collection, and the preservation unit address is Wuhan university, Wuhan City, China.
Further, the milk-derived streptococcus salivarius is isolated from human fresh breast milk.
The second aspect of the present invention provides the method for separating milk-derived streptococcus salivarius described above, comprising the steps of:
step one, diluting a fresh breast milk sample, and coating a part of the diluted sample on a KF streptococcus agar plate for culture;
and step two, after the culture is finished, selecting a single colony and identifying to obtain the lactogenic streptococcus salivarius.
Further, in step one, the fresh breast milk is diluted 101-103And (4) doubling.
Further, the culture condition in the step one is 37 ℃ and anaerobic, and the culture time is 2-3 days.
By adopting the technical scheme, compared with the prior art, the invention has the following technical effects:
the milk-derived streptococcus salivarius for predicting the obesity of the infants is used as facultative anaerobe, is easy to colonize in intestinal tracts, creates conditions for maturation and stability of later-stage intestinal flora, and can predict the obesity of offspring by detecting the abundance of streptococcus salivarius in the early life stage.
Drawings
The milk-derived Streptococcus salivarius SHDP5A1 for predicting infant obesity, which is disclosed by the invention, is preserved and named as Streptococcus salivarius by classification, the preservation number is CCTCC NO: M2021060, the preservation date is 2021, 01, 14 days, the preservation unit is China center for type culture Collection, and the preservation unit address is Wuhan university, Wuhan City, China.
FIG. 1 is a graph showing the results of comparing the structures of breast milk flora in an obese group and a normal group of pregnant women in accordance with an embodiment of the present invention;
fig. 2 is a graph showing the results of comparing the intestinal flora structure of infants aged 6 months in the practice of the present invention in an obese group and a normal group of pregnant women.
Detailed Description
The invention provides milk-derived streptococcus salivarius SHDP5A1 for predicting infant obesity, which is preserved at present, wherein the preservation number is CCTCC NO: M2021060, the preservation date is 2021, 01, 14 days, the preservation unit is China center for type culture preservation, and the preservation unit address is Wuhan university in Wuhan City.
In a preferred embodiment of the invention, the milk-derived streptococcus salivarius is isolated from human fresh breast milk.
The present invention will be described in detail and specifically with reference to the following examples and drawings so as to provide a better understanding of the invention, but the following examples do not limit the scope of the invention.
Example 1
The embodiment provides a method for separating milk-derived streptococcus salivarius SHDP5a1, wherein the preservation number of the milk-derived streptococcus salivarius is CCTCC NO: m2021060, wherein the preservation date is 2021, 14 days 01 and 14, the preservation unit is China center for type culture Collection, and the preservation unit address is Wuhan university in Wuhan City; the specific separation method is as follows:
dilution of fresh breast milk sample 10-103After doubling, 50-200 mul of the bacillus subtilis is taken and coated on a commercial KF streptococcus agar plate, anaerobic culture is carried out for 2-3 days at 37 ℃, and a single colony is selected and identified. The 16S rRNA gene sequence of the strain is identified as follows:
AGGACGAACGCTGGCGGCGTGCCTAATACATGCAAGTAGAACGCTGAAGAGAGGAGCTTGCTCTTCTTGGAAGAGTTGCGAACGGGTGAGTAACGCGTAGGTAACCTGCCTTGTAGCGGGGGATAACTATTGGAAACGATAGCTAATACCGCATAACAATGGGTGACCCATGTCATTCATTTGAAAGGGGCAACTGCTCCACTACAAGATGGACCTGCGTTGTATTAGCTAGTAGGTGAGGTAACGGCTCACCTAGGCGACGATACATAGCCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCGGCAATGGGGGCAACCCTGACCGAGCAACGCCGCGTGAGTGAAGAAGGTTTTCGGATCGTAAAGCTCTGTTGTAAGTCAAGAACGAGTGTGAGAGTGGAAAGTTCACACTGTGACGGTAGCTTACCAGAAAGGGACGGCTAATTACGTGCCCAGCAGCCGCGGTAATACGTAGGTCCCGAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGTTTGATAAGTCTGAAGTTAAAGGCTGTGGCTCAACCATAGTTCGCTTTGGAAACTGTCAAACTTGAGTGCAGAAGGGGAGAGTGGAATTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAGGAACACCGGTGGCGAAAGCGGCTCTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAGGTGTTGGGTCCTTTCCGGGATTCAGTGCCGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCCGATGCTATTTCTAGAGATAGGAAGTTACTTCGGTACATCGGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCCTATTGTTAGTTGCCATCATTCAGTTGGGCACTCTAGCGAGACTGCCGGTAATAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGTTGGTACAACGAGTTGCGAGTCGGTGACGGCGAGCTAATCTCTTAAAGCCAATCTCAGTTCGGATTGTAGGCTGCAACTCGCCTACATGAAGTCGGAATCGCTAGTAATCGCGGATCAGCACGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTATGGAGCCAGCCGCCTAAGGTGGGATAGATGATTGGGGTGAAGT。
verification examples
The effect of milk-derived streptococcus salivarius SHDP5a1 in predicting obesity in infants is verified by detecting the abundance of streptococcus salivarius in breast milk and infant feces and detecting the weight of infants. The specific implementation process and results are as follows:
10 pregnant women with normal weight during pregnancy and 15 obese pregnant women (BMI)>27kg/m2) The study was included, and the milk was collected after the delivery of the baby by breast feeding, signing an informed consent.
Firstly, cleaning the breast of a lying-in woman with physiological saline, extracting 20ml of milk by using a breast pump, immediately sending 5ml of milk to a laboratory for dilution, coating 50-200 mu l of milk on a commercial KF (KF) streptococcus agar plate, carrying out anaerobic culture at 37 ℃ for 2-3 days, selecting a single colony for colony identification, sending the rest 15ml of milk to a refrigerator for freezing, carrying out follow-up visit on indexes such as weight and the like at the age of 6 months of an infant, collecting excrement again, and sending the excrement into the refrigerator for freezing. All samples were subjected to 16S rRNA gene sequence sequencing, and the results are shown in FIGS. 1 and 2.
According to follow-up record and analysis of body weight, the average body weight of the infants at the age of 6 months of the obese pregnant woman is higher than that of the normal pregnant woman delivery group (9.08kg vs. 8.37kg); as can be seen from fig. 1 and 2, the abundance of streptococcus salivarius in the breast milk is lower than that of the normal pregnant group (1.79% vs. 2.73%) (fig. 1), and also lower abundance (10.88% vs. 16.61%) is present in the intestine of infants at the age of 6 months (fig. 2).
The embodiments of the present invention have been described in detail, but the embodiments are merely examples, and the present invention is not limited to the embodiments described above. Any equivalent modifications or alterations to this practice will occur to those skilled in the art and are intended to be within the scope of this invention. Accordingly, equivalent changes and modifications made without departing from the spirit and scope of the present invention should be covered by the present invention.
Sequence listing
<110> tenth people hospital in Shanghai City
<120> milk-derived streptococcus salivarius for predicting obesity in infants and method for isolating same
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1477
<212> DNA
<213> 16S rRNA sequence of Streptococcus salivarius strain SHDP5A1
<400> 1
aggacgaacg ctggcggcgt gcctaataca tgcaagtaga acgctgaaga gaggagcttg 60
ctcttcttgg aagagttgcg aacgggtgag taacgcgtag gtaacctgcc ttgtagcggg 120
ggataactat tggaaacgat agctaatacc gcataacaat gggtgaccca tgtcattcat 180
ttgaaagggg caactgctcc actacaagat ggacctgcgt tgtattagct agtaggtgag 240
gtaacggctc acctaggcga cgatacatag ccgacctgag agggtgatcg gccacactgg 300
gactgagaca cggcccagac tcctacggga ggcagcagta gggaatcttc ggcaatgggg 360
gcaaccctga ccgagcaacg ccgcgtgagt gaagaaggtt ttcggatcgt aaagctctgt 420
tgtaagtcaa gaacgagtgt gagagtggaa agttcacact gtgacggtag cttaccagaa 480
agggacggct aattacgtgc ccagcagccg cggtaatacg taggtcccga gcgttgtccg 540
gatttattgg gcgtaaagcg agcgcaggcg gtttgataag tctgaagtta aaggctgtgg 600
ctcaaccata gttcgctttg gaaactgtca aacttgagtg cagaagggga gagtggaatt 660
ccatgtgtag cggtgaaatg cgtagatata tggaggaaca ccggtggcga aagcggctct 720
ctggtctgta actgacgctg aggctcgaaa gcgtggggag cgaacaggat tagataccct 780
ggtagtccac gccgtaaacg atgagtgcta ggtgttgggt cctttccggg attcagtgcc 840
gcagctaacg cattaagcac tccgcctggg gagtacgacc gcaaggttga aactcaaagg 900
aattgacggg ggcccgcaca agcggtggag catgtggttt aattcgaagc aacgcgaaga 960
accttaccag gtcttgacat cccgatgcta tttctagaga taggaagtta cttcggtaca 1020
tcggtgacag gtggtgcatg gttgtcgtca gctcgtgtcg tgagatgttg ggttaagtcc 1080
cgcaacgagc gcaaccccta ttgttagttg ccatcattca gttgggcact ctagcgagac 1140
tgccggtaat aaaccggagg aaggtgggga tgacgtcaaa tcatcatgcc ccttatgacc 1200
tgggctacac acgtgctaca atggttggta caacgagttg cgagtcggtg acggcgagct 1260
aatctcttaa agccaatctc agttcggatt gtaggctgca actcgcctac atgaagtcgg 1320
aatcgctagt aatcgcggat cagcacgccg cggtgaatac gttcccgggc cttgtacaca 1380
ccgcccgtca caccacgaga gtttgtaaca cccgaagtcg gtgaggtaac cttatggagc 1440
cagccgccta aggtgggata gatgattggg gtgaagt 1477
Claims (6)
1. The milk-derived streptococcus salivarius for predicting infant obesity is characterized in that the 16SrRNA gene sequence of the milk-derived streptococcus salivarius is shown as SEQ ID NO. 1.
2. The milk-derived Streptococcus salivarius according to claim 1, wherein the milk-derived Streptococcus salivarius is Streptococcus salivarius strain SHDP5A1 which is classified and named as Streptococcus salivarius, and has a preservation number of CCTCC NO: M2021060, a preservation date of 2021, 01, 14 days, a preservation unit of China center for type culture Collection, and a preservation unit address of Wuhan university, Wuhan City, China.
3. Milk-derived streptococcus salivarius according to claim 1 wherein the milk-derived streptococcus salivarius is isolated from human fresh breast milk.
4. A method for the isolation of milk-derived streptococcus salivarius according to any one of claims 1 to 3 comprising the steps of:
step one, diluting a fresh breast milk sample, and coating a part of the diluted sample on a KF streptococcus agar plate for culture;
and step two, after the culture is finished, selecting a single colony and identifying to obtain the milk-derived streptococcus salivarius.
5. The method according to claim 4, wherein the fresh human milk is diluted 10 in step one1-103And (4) doubling.
6. The method for separating milk-derived streptococcus salivarius according to claim 4 wherein the culture conditions in step one are 37 ℃ and anaerobic, and the culture time is 2-3 days.
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2021
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