CN113616715A - Fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora thereof - Google Patents
Fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora thereof Download PDFInfo
- Publication number
- CN113616715A CN113616715A CN202111009083.7A CN202111009083A CN113616715A CN 113616715 A CN113616715 A CN 113616715A CN 202111009083 A CN202111009083 A CN 202111009083A CN 113616715 A CN113616715 A CN 113616715A
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- traditional chinese
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- chinese medicine
- liquid
- oral liquid
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Abstract
The invention discloses a fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora thereof, which is prepared by uniformly mixing 1-1.5 kg of a traditional Chinese medicine composition, 0.8-1.2L of a compound probiotic bacterial liquid, 0.5-1 kg of brown sugar and 0.3-0.6 kg of poplar flower, adjusting the water content to 55-65%, hermetically fermenting at 37 ℃ for 6-8 days to complete traditional Chinese medicine fermentation, continuously adding 0.8-1.2L of the compound probiotic bacterial liquid and 0.5-0.7L of deionized water into fermented traditional Chinese medicine when preparing a traditional Chinese medicine extracting solution, standing and soaking at 23-26 ℃ after uniformly mixing for 24-30 h, stirring irregularly during standing and soaking, and filtering by using a filter screen to remove filter residues to obtain the fermented traditional Chinese medicine oral liquid; the fermented traditional Chinese medicine oral liquid fully exerts the effects of probiotics, traditional Chinese medicines and fermentation products thereof, can remarkably treat piglet diarrhea, improve piglet intestinal flora, reduce bacterial drug resistance and antibiotic drug residue caused by feed antibiotic abuse, and ensure food production safety; the preparation method of the fermented traditional Chinese medicine oral liquid is simple and is suitable for industrial production and market popularization and application.
Description
Technical Field
The invention belongs to the technical field of composite probiotic preparations, and particularly relates to a fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora of piglets.
Background
The piglets are the core of modern pig raising production, and the health condition of the piglets has extremely important influence on the later growth and development. Because the development of the immune system of the piglets is not complete, the invasion of harmful microorganisms can easily cause diarrhea of the piglets. In one month after the birth of piglets, the development function of digestive glands and thermoregulation centers is incomplete and not perfect, such as temperature change and overhigh milk fat rate, which easily causes dyspepsia and diarrhea. The diarrhea is a common disease in pig farm cultivation, can cause the survival rate of piglets to be reduced, influences the composition of intestinal flora, and causes the diarrhea easily due to the abnormal intestinal flora, thereby bringing huge loss to the breeding industry.
To reduce this risk, antibiotics are used in large quantities in livestock and poultry farming as growth-promoting factors, making a great contribution to the farming industry. However, the long-term use of antibiotics in large quantities destroys the micro-ecological balance of the organism, resulting in the generation of drug-resistant bacteria and the variation of bacteria. Seriously affecting the survival rate of piglets and easily causing endogenous infection or secondary infection of animals. The fade-out and the comprehensive prohibition of antibiotics in the breeding industry are great trends. Therefore, there is a need to develop a new formulation that is an effective antibiotic substitute for controlling diarrhea in piglets.
The probiotics have various probiotic properties, and can generate antibacterial substances and competitively adhere to intestinal epithelial cells with pathogenic microorganisms to prevent the adhesion and colonization of the pathogens in the intestinal tract; the probiotics can improve the survival rate of intestinal epithelial cells during virus infection, protect the normal physiological structure of the intestinal tract and stimulate protective immune response to regulate the immunity of the organism; the probiotics can protect and resist the invasion of pathogenic microorganisms by adjusting the intestinal microecological balance; the probiotic metabolite organic acid, digestive enzyme, antibacterial substance, etc. have effects of antagonizing, inhibiting the growth of pathogenic microorganism, and promoting or improving host immunity. And the composite probiotics have better beneficial effects on the aspects of intestinal tract shape, disease resistance and organism immunity of livestock and poultry.
The Chinese herbal medicine is a natural plant, has the characteristics of no toxic or side effect, no drug resistance, no pollution, no drug residue and the like, has the functions of enhancing the animal immunity, improving the anti-stress capability of animals, effectively regulating the organism metabolism, improving the quality of livestock and poultry and the like, and is popular with most farmers.
The probiotics and the Chinese herbal medicines are green feed additives which are generally accepted at present, and the number of the viable probiotics of the fermented traditional Chinese medicine oral liquid is more than or equal to 108CFU/mL, including probiotic organic acid, digestive enzyme, antibacterial substance and other metabolites, and composite Chinese herbal medicine active ingredients for fermentation conversion and dissolution. After the probiotics are fermented, certain traditional Chinese medicine components are decomposed and converted into new medicine components, particularly, multiple enzymes generated by the probiotics can promote effective components of the Chinese herbal medicines to be fully absorbed and utilized by animal organisms, the Chinese herbal medicines contain rich nutrition and can promote a large amount of proliferation of the probiotics, the Chinese herbal medicines supplement each other, and the efficacy of the Chinese herbal medicines, namely the natural medicine, is better exerted. The Chinese herbal medicine fermented by the composite probiotics has the effect of the Chinese herbal medicine and also has the probiotic auxiliary function of microorganisms. Thereby adjusting the flora balance in the intestinal tract of the pigletFurther, the newborn piglets have good health condition. The microbial liquid can improve the intestinal flora of animals, enhance the inhibition of animals on harmful microorganisms, improve the immunity of organisms and the like.
Disclosure of Invention
The invention provides fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora thereof, which is prepared by uniformly mixing 1-1.5 kg of traditional Chinese medicine composition, 0.8-1.2L of composite probiotic bacteria liquid, 0.5-1 kg of brown sugar and 0.3-0.6 kg of poplar flower, adjusting the water content to 55-65%, and performing sealed fermentation at 37 ℃ for 6-8 days to complete traditional Chinese medicine fermentation; when the traditional Chinese medicine extracting solution is prepared, 0.8-1.2L of compound probiotic bacteria liquid and 0.5-0.7L of deionized water are continuously added into the fermented traditional Chinese medicine, the mixture is uniformly mixed and then is kept stand and soaked at 23-26 ℃ for extraction for 24-30 h, the mixture is kept stand and soaked for indefinite stirring to facilitate the dissolution of substances, a 80-mesh filter screen is used for filtering and removing filter residues to obtain the fermented traditional Chinese medicine oral liquid, the number of effective viable bacteria in the fermented traditional Chinese medicine oral liquid is more than or equal to 108CFU/mL。
The composite probiotic bacterial liquid of the invention has the effective viable count of 108~109CFU/mL Lactobacillus salivarius (Lactobacillus salivarius) Bacterial liquid, effective viable count 108~109CFU/mL lactococcus lactis (Lactococcus lactis) Bacterial liquid, effective viable count 108~109CFU/mL Lactobacillus plantarum (Lactobacillus plantarum) The bacterial liquid is prepared from the following components in percentage by volume (0.9-1.2): (0.9-1.1): (0.9-1.1) to obtain a mixed bacterial liquid, and then mixing the mixed bacterial liquid and the effective viable count of 108~109CFU/mL Bacillus licheniformis (C: (C)Bacillus licheniformis) And (3) uniformly mixing the mixed bacterial liquid and the bacillus licheniformis bacterial liquid according to the volume ratio of the bacterial liquid being 4-6: 1.
Wherein Lactobacillus salivarius (Lactobacillus salivarius) The preservation number is CGMCC No.20700 (202011523897.8), lactococcus lactis (Lactococcus lactis) The preservation number is CGMCC No.20699 (202011337808.0), lactobacillus plantarum (A)Lactobacillus plantarum) The preservation number is CGMCC No.21266 (202110637585.8), and Bacillus licheniformis (Bacillus licheniformis)Bacillus licheniformis) The preservation number is CGMCC No.21267 ()202110289676.7), all of which are disclosed in other patent applications.
The traditional Chinese medicine composition comprises, by weight, 4-6 parts of liquorice, 4-6 parts of coptis chinensis, 5-7 parts of sweet wormwood, 5-7 parts of immature bitter orange, 5-7 parts of hawthorn, 5-7 parts of platycodon grandiflorum, 5-7 parts of honeysuckle stem, 5-7 parts of radix bupleuri, 5-7 parts of dandelion, 5-7 parts of folium isatidis, 5-7 parts of houttuynia cordata, 5-7 parts of scutellaria baicalensis, 5-7 parts of cape jasmine, 5-7 parts of selfheal, 5-7 parts of rhizoma atractylodis, 9-11 parts of golden cypress, 20-25 parts of rhizoma alismatis, 30-35 parts of poria cocos and 30-35 parts of astragalus membranaceus.
The method of the invention has the following characteristics:
(1) the fermented traditional Chinese medicine oral liquid fully exerts the effects of probiotics, traditional Chinese medicines and fermentation products thereof, can obviously treat piglet diarrhea and improve piglet intestinal flora; on the 6 th-20 th day of piglet day age, the fermented traditional Chinese medicine oral liquid is drenched into the stomach of the piglet, the drenched dose is 15mL each time, the oral liquid is taken once a day, and the treatment is carried out for three days;
(2) the number of the probiotics viable bacteria of the fermented traditional Chinese medicine oral liquid is more than or equal to 108CFU/mL, including probiotic organic acid, digestive enzyme, antibacterial substance and other metabolites, and composite Chinese herbal medicine active ingredients for fermentation conversion and dissolution; after the probiotics are fermented, certain traditional Chinese medicine components are decomposed and converted into new medicine components, particularly, multiple enzymes generated by the probiotics can promote the effective components of the Chinese herbal medicines to be fully absorbed and utilized by animal organisms, the Chinese herbal medicines contain rich nutrition and can promote the large-scale proliferation of the probiotics, the Chinese herbal medicines supplement each other, and the efficacy of the natural medicine of the Chinese herbal medicines is better exerted; the Chinese herbal medicine fermented by the composite probiotics has the effect of the Chinese herbal medicine and also has the probiotic auxiliary function of microorganisms;
(3) the traditional Chinese medicine composition disclosed by the invention contains abundant nutrients, prebiotics and traditional Chinese medicine effective components for inhibiting pathogenic bacteria, can improve the growth performance of animals by improving intestinal flora, enhance the resistance of organisms to pathogenic bacteria, stimulate the generation of immune response of the organisms, maintain the higher-level immune response of the organisms, obviously improve the intestinal micro-ecological environment of the organisms under the condition of pathogenic bacterial infection, and improve the resistance of the organisms to the pathogenic bacteria;
(4) the fermented traditional Chinese medicine oral liquid can inhibit the propagation of harmful pathogenic bacteria, reduce the addition of antibiotics serving as preparations for preventing and treating animal diseases in the feed in long-term sub-therapeutic dose and ultrahigh dose, and avoid the harm of bacterial drug resistance, antibiotic drug residue, food safety and the like caused by the abuse of feed antibiotics. The fermented traditional Chinese medicine oral liquid can obviously improve the intestinal mucosa immunity level, reduce or avoid intestinal diseases and maintain the micro-ecological balance of the intestinal tract.
Drawings
FIG. 1 is the composition at the level of the phylum Enterobacteriaceae of example 3;
FIG. 2 is the composition at the level of the gut flora of example 3.
Detailed Description
The technical scheme of the present invention is further described in detail by the following examples, but the content of the present invention is not limited thereto, and the methods in the examples are all conventional methods unless otherwise specified, and materials, reagents and the like used therein are commercially available or formulated by conventional methods unless otherwise specified.
Example 1: preparation of fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora thereof
(1) Recovering strains stored according to the proportion of 50% of glycerol to bacteria liquid =7:3 in water bath at 37 ℃ for 1min, streaking the recovered lactobacillus salivarius, lactococcus lactis and lactobacillus plantarum on an MRS culture medium by using an inoculating loop, streaking the bacillus licheniformis strain on an LB culture medium by using an inoculating loop, and then placing the strains in a constant-temperature incubator at 37 ℃ for inverted culture for 12 h; respectively selecting single colony, inoculating to sterile liquid culture medium, and culturing in shaking bed to OD600The value is 0.6-0.8, the bacterial liquid is subcultured by 1% of inoculum size, and then the subcultured bacterial liquid is inoculated into the sterile liquid culture medium with the required dosage and cultured at 37 ℃, so that the effective viable count of 10 is obtained8~109CFU/mL lactobacillus salivarius bacterial liquid and effective viable count of 108~109CFU/mL lactococcus lactis bacterial liquid with effective viable count of 108~109CFU/mL lactobacillus plantarum, effective viable count 108~109CFU/mL of Bacillus licheniformis liquid;
(2) mixing lactobacillus salivarius, lactococcus lactis and lactobacillus plantarum according to the volume ratio of 1:1:1 to prepare a mixed bacterial liquid, and then mixing the mixed bacterial liquid and the bacillus licheniformis bacterial liquid according to the volume ratio of 4:1 to prepare a composite probiotic bacterial liquid;
(3) the traditional Chinese medicine composition comprises, by weight, 4 parts of liquorice, 6 parts of coptis chinensis, 5 parts of sweet wormwood, 5 parts of immature bitter orange, 6 parts of hawthorn, 6 parts of platycodon grandiflorum, 5 parts of honeysuckle stem, 5 parts of radix bupleuri, 6 parts of dandelion, 6 parts of folium isatidis, 5 parts of houttuynia cordata, 5 parts of scutellaria baicalensis, 6 parts of cape jasmine, 7 parts of selfheal, 7 parts of rhizoma atractylodis, 9 parts of phellodendron, 20 parts of rhizoma alismatis, 35 parts of poria cocos and 35 parts of astragalus membranaceus;
(4) uniformly mixing 1kg of the traditional Chinese medicine composition, 0.8L of compound probiotic bacteria liquid, 0.5kg of brown sugar and 0.6kg of poplar flower, adjusting the water content to 60%, hermetically fermenting for 6-8 days at 37 ℃ to complete traditional Chinese medicine fermentation, continuously adding 0.8L of compound probiotic bacteria liquid and 0.5L of deionized water into the fermented traditional Chinese medicine when preparing a traditional Chinese medicine extracting solution, uniformly mixing, standing at 23 ℃, soaking and extracting for 30 hours (stirring irregularly during the period to facilitate dissolution of substances), and filtering by using a 80-mesh filter screen to remove filter residues to obtain the fermented traditional Chinese medicine oral liquid.
Example 2: preparation of fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora thereof
(1) Recovering strains preserved according to the volume ratio of 50% of glycerol to bacterium liquid =7:3 in water bath at 37 ℃ for 1min, scribing lactobacillus salivarius, lactococcus lactis and lactobacillus plantarum strains on an MRS culture medium by using an inoculating loop after recovery, scribing bacillus licheniformis strains on an LB culture medium by using the inoculating loop, and then placing the strains in a constant-temperature incubator at 37 ℃ for inverted culture for 12 h; respectively picking single colony to sterile liquid medium for shake culture to OD600The value is 0.6-0.8, the bacterial liquid is subcultured by 1% of inoculum size, and then the subcultured bacterial liquid is inoculated into an aseptic liquid culture medium for culture at 37 ℃ to obtain the effective viable count of 108~109CFU/mL lactobacillus salivarius bacterial liquid and effective viable count of 108~109CFU/mL lactococcus lactis bacterial liquid with effective viable count of 108~109CFU/mL lactobacillus plantarum, effective viable count 108~109CFU/mL of Bacillus licheniformis liquid;
(2) mixing lactobacillus salivarius, lactococcus lactis and lactobacillus plantarum according to the volume ratio of 0.9:1:1.1 to prepare a mixed bacterial liquid, and then mixing the mixed bacterial liquid and the bacillus licheniformis bacterial liquid according to the volume ratio of 5:1 to prepare a composite probiotic bacterial liquid;
(3) the traditional Chinese medicine composition comprises, by weight, 6 parts of liquorice, 4 parts of coptis chinensis, 6 parts of sweet wormwood, 6 parts of immature bitter orange, 7 parts of hawthorn, 7 parts of platycodon grandiflorum, 7 parts of honeysuckle stem, 7 parts of radix bupleuri, 7 parts of dandelion, 7 parts of folium isatidis, 6 parts of houttuynia cordata, 7 parts of scutellaria baicalensis, 6 parts of cape jasmine, 6 parts of selfheal, 6 parts of rhizoma atractylodis, 11 parts of golden cypress, 25 parts of rhizoma alismatis, 30 parts of poria cocos and 30 parts of astragalus membranaceus;
(4) uniformly mixing 1.5kg of the traditional Chinese medicine composition, 1.0L of compound probiotic bacteria liquid, 0.8kg of brown sugar and 0.3kg of poplar flower, adjusting the water content to 65%, sealing and fermenting at 37 ℃ for 7 days, continuously adding 1.0L of compound probiotic bacteria liquid and 0.7L of deionized water into the fermented material, sealing and standing at 25 ℃ after uniformly mixing, extracting for 24 hours (stirring irregularly during the period to facilitate the dissolution of substances), and filtering to remove filter residues to obtain the fermented traditional Chinese medicine oral liquid.
Example 3: tolerance of 4 strains in composite probiotic bacterial liquid to simulated gastrointestinal fluid
Preparing artificial gastric juice: 1.64mL of 1mol/L diluted hydrochloric acid is taken in a container, 80mL of water is added, 1g of pepsin is added, the pH value is adjusted to 3.0, the mixture is uniformly mixed and the volume is adjusted to 100mL, and a 0.22 mu m filter membrane is used for sterilization; preparing artificial gastric juice with pH of 4.0 by the same method, and placing in a refrigerator at 4 deg.C for use;
preparing artificial intestinal juice: taking 0.68g of monopotassium phosphate, adding 80mL of water for dissolution, adding 1g of trypsin, and adjusting the pH value to 6.8 by using 0.lmol/L sodium hydroxide solution; diluting with appropriate amount of water to 100mL, filtering with 0.22 μm filter membrane for sterilization to obtain simulated intestinal juice, and placing in refrigerator at 4 deg.C;
recovering strains stored according to the proportion of 50% of glycerol to bacteria liquid =7:3 in a water bath at 37 ℃ for one minute, streaking the recovered lactobacillus salivarius, lactococcus lactis and lactobacillus plantarum on an MRS culture medium by using an inoculating loop, streaking the bacillus licheniformis strain on an LB culture medium by using an inoculating loop, and then placing the strains in a constant-temperature incubator at 37 ℃ for inverted culture for 12 hours; respectively picking single colony to sterile liquid medium for shake culture to OD600And (3) carrying out subculture on the bacterial liquid with the inoculation amount of 1%, inoculating lactobacillus salivarius, lactococcus lactis and lactobacillus plantarum to the same MRS liquid culture medium for mixed culture, and inoculating bacillus licheniformis to an LB liquid culture medium. Shaking culture at 37 deg.C and 150rpm to OD600The value is 0.6 to 0.8; respectively taking 10mL of activated mixed bacteria and bacillus licheniformis suspension, centrifuging at 13000r/min for 10min, collecting thalli, respectively adding 10mL of simulated gastrointestinal fluid, performing vortex mixing uniformly, performing standing culture at 37 ℃ for 4h, determining the number of viable bacteria before and after treatment of the artificial gastric juice and the artificial intestinal juice by using a plate counting method for 0h and 4h of the mixed bacteria by using an MRS agar culture medium, and determining the number of viable bacteria before and after treatment of the artificial gastric juice and the artificial intestinal juice by using an LB agar culture medium for 0h and 4h of the bacillus licheniformis, wherein the experimental results are shown in Table 1;
survival rate ═ N1/N0]×100%(N0: the number of viable bacteria is 0 h; n1: number of viable bacteria after digestion of simulated gastrointestinal fluid)
TABLE 1 survival rate of mixed lactic acid bacteria and spores artificially simulated gastrointestinal fluids
Antibacterial experiment of fermented traditional Chinese medicine oral liquid:
the antibacterial effect of the fermented traditional Chinese medicine oral liquid is measured by a double-layer flat-plate oxford cup diffusion method, a sterilized LB agar solid culture medium is poured into a culture dish to be cooled, 4mL of LB agar semisolid culture medium which is sterilized and cooled to about 50 ℃ is respectively and uniformly mixed with 100 mu L of escherichia coli, shigella, staphylococcus aureus and salmonella and poured onto the LB solid culture medium, and the mixed liquid is uniformly distributed on the LB solid culture medium. Placing the sterilized Oxford cup on the mixed solution by using sterile forceps after the mixed solution is solidified, and adding 200 mu L of fermented traditional Chinese medicine oral liquid filtered by a 0.22 mu m filter membrane into the Oxford cup; diffusing in a refrigerator at 4 ℃ for 4h, culturing at 37 ℃ for constant temperature for 12h, and observing the size of a bacteriostatic zone; the diameter of the inhibition zone is measured by using a vernier caliper (two effective figures are reserved), the experiment is independently repeated for three times, the diameter of the oxford cup is 8mm, and the experimental result is shown in table 2;
TABLE 2 treatment of diarrhea in piglets and improvement of antibacterial property of fermented traditional Chinese medicine oral liquid for intestinal flora
As shown in the experimental results in tables 1 and 2, the fermented traditional Chinese medicine oral liquid has good gastrointestinal liquid tolerance and excellent characteristics of inhibiting pathogenic bacteria in a broad spectrum.
Example 3: excrement form observation and flora diversity analysis of fermented traditional Chinese medicine oral liquid on diarrhea piglets in treatment process
Stool form of diarrhea piglets: water-like, non-textured, flat spread;
stool form of healthy piglets: the paste is dry, is obviously agglomerated and is sticky and not easy to separate;
according to the feces forms of the diarrhea piglets and the healthy piglets, determining the diarrhea piglets as experimental objects, taking the feces samples of the diarrhea piglets as a disease group, determining the healthy piglets according to the feces forms of the piglets as a health group, taking the feces samples of the healthy piglets as a health group, feeding the diarrhea piglets with fermented traditional Chinese medicine oral liquid in a feeding mode of pouring 15mL/d, and finding the feces forms after feeding for two days: the soil is very moist and has obvious texture, residues are left on the ground after being picked up, and the shape of the excrement cannot be maintained; the method can be obviously different from the feces form of the diarrhea piglets, feces samples are continuously taken and named as a "treat group", the feces samples are continuously fed for the third day in the same feeding mode, the feces form of the piglets is found to be similar to that of the healthy piglets, the feces samples are taken and named as a "recovery group", and three parallel serial numbers of each group are sent to the Shanghai Meiji biological medicine science and technology Limited company for 16Sr RNA microbial diversity sequencing.
An Illumina PE300 sequencing platform is adopted to sequence a sample, and a Meiji biological cloud platform is adopted to comprehensively analyze the number of operable classification units (OTUs) of intestinal flora, the diversity of alpha and beta and the phylum and the genus of different bactria.
And 4 groups of piglet intestinal flora are subjected to alpha and beta diversity analysis, the coverage rate of each group reaches over 0.99, the data coverage is high, and the reliability is good, and the alpha diversity index shannon index and the chao index are expressed in a mode of mean value plus or minus standard deviation.
Shannon index Student T test analysis shows that the diversity of intestinal flora of piglets in the treatment group is obviously higher than that in the disease group (P <0.05), the diversity of the intestinal flora of piglets in the recovery group is extremely lower than that in the treatment group: (P <0.001);
Analysis of the chao index Student T test shows that the intestinal flora abundance of the piglet in the disease group is very significantly lower than that in the health group (P <0.01), the intestinal flora abundance of the young animals in the recovery group is obviously lower than that of the health group (a)P <0.05), the intestinal flora abundance of the piglet in the treatment group is remarkably higher than that in the disease group (P <0.01), the abundance of intestinal flora of piglets in the treatment group is remarkably higher than that in the recovery groupP <0.001) and the results are shown in table 3, wherein the same letter indicates no significant difference after each index value, and the difference is significant when the letters are different.
TABLE 3 analysis of alpha and beta diversity of piglet intestinal flora
The Shannon index is a statistic of flora diversity, and the larger the index value is, the higher diversity is indicated; the Chao index is a statistic of flora abundance, the larger index value represents more species, the larger value represents Coverage rate, and the larger value represents better Coverage.
And (3) analyzing the flora structure: the research is divided into four groups, the average value of each group of three is taken in parallel, according to the sequencing result, the relative abundance distribution of the phylum classification species 7 before ranking is shown in figure 1, and the Yunnan black pig intestinal flora mainly comprises Firmicutes of Firmicutes, bacteroidetes bacteroidata, actinomycetes, Proteobacteria, fusobacteria Fusobacteriota, desulfobacterium desulfobactrobacteria and synergestita; the relative abundance of each group at 7 gate levels is shown in table 4, expressed as mean plus minus standard deviation; one-way ANOVA (One-way ANOVA) showed significant differences in pairwise comparisons between Actinobacteriota, Desulfobacterota and Synergestota groups of ActinobacteriotaP <0.05)。
TABLE 4 relative abundance values of the intestinal flora of each group of piglets at the phylum level
Note: after the horizontal relative abundance value of the same gate, the same letter indicates that no significant difference exists among groups, and different letters indicate that significant difference exists among groups
In the genus level, the dominant genus of the intestinal tract of Yunnan black piglet is LactobacillusLactobacillusBacteroides genusBacteroides、norank_f__Muribaculaceae、norank_f__Eubacterium_coprostanoligenes_ group、Olsenella、unclassified_f__LachnospiraceaeBifidobacterium, BifidobacteriumBifidobacterium、Christensenellaceae_R-7_group、Escherichia-ShigellaEnterococcus genusEnterococcus. The relative abundance distribution of different sets of dominance levels is shown in FIG. 2; the relative abundance values are shown in Table 5, and One-way ANOVA (One-way ANOVA) is shown in LactobacillusLactobacillus、norank_f__Muribaculaceae、Olsenella、unclassified_ f__Lachnospiraceae、BifidobacteriumThe relative abundance of bacteria in different groups differed significantly (P <0.05)。
TABLE 5 relative abundance values of the intestinal flora of each group of piglets at the genus level
Note: after the relative abundance value of the same genus level, the same letter indicates that no significant difference exists among groups, and different letters indicate that significant difference exists among groups
The most abundant relative species among the phylum and genus levels of dominance are Firmicutes and lactobacilliLactobacillusAnd in Firmicutes, LactobacillusLactobacillusThe relative abundance of bacteria in the recovery group is higher than that of the treatment group than that of the area group, and the comparison between the recovery group and the health group has no significant difference.
The diarrhea of the piglets isCommon diseases of pig farm cultivation, intestinal flora balance has important significance for maintaining health state of organisms. Bacteria in the intestinal tract can participate in the metabolism of nutrient substances, Firmicute bacteria in Firmicute are related to energy absorption, and the experimental result shows that: the relative abundance of the bacteria in the phylum firmicutes of the disease group is 58.73%, the relative abundance of the bacteria in the phylum firmicutes of the disease group is 78.24%, the relative abundance of the bacteria in the phylum regulus of the recovery group is 87.44%, and the relative abundance of the bacteria in the phylum heamostemensis of the disease group is 89.93%; the fermented traditional Chinese medicine oral liquid prepared by the invention can improve the relative abundance of Firmicute bacteria in Firmicute of Firmicute group when being fed to Yunnan black pigs, and the relative abundance of Firmicute bacteria in Firmicute group and health group has no obvious difference after treatment. After the fermented traditional Chinese medicine oral liquid is fed, the relative abundance of actinomycetemcomia and desulfobacterium desulfobacticola in recovery group is obviously higher than that in health groupP <0.001), the relative abundance of the Synergestotia bacteria in the recovery group is obviously higher than that in the health group (B)P <0.05)。
recure group lactobacillusLactobacillusThe relative abundance of bacteria is significantly higher than that of the disease group (P <0.05), the relative abundance distribution of the lactobacillus bacteria in different groups well explains the diversity and abundance difference of the intestinal flora of Yunnan black pigs among different groups. The reason why the Yunnan black pig intestinal flora diversity and richness of the traditional Chinese medicine oral liquid is higher than that of the disease group is that the traditional Chinese medicine oral liquid is fed by the traditional Chinese medicine oral liquid fermented for two days, and the reason why the Yunnan black pig intestinal flora diversity recovery group is lower than that of the traditional Chinese medicine oral liquid fermented for two days is that the traditional Chinese medicine oral liquid fermented for two days can inhibit the growth of various pathogenic bacteria in the treatment process. The reason why the Yunnan black pig intestinal flora abundance disease group is lower than the health group is that the intestinal microecology of the black pig in the health group is more abundant; the reason that the recure group with the abundance of the Yunnan black pig intestinal flora is lower than the health group is probably because the fermented traditional Chinese medicine oral liquid changes the structure of the black pig intestinal flora in the treatment process, and although the recure group has lower abundance, the probiotic property is stronger;
recure group bifidobacteriumBifidobacteriumThe relative abundance of bacteria is significantly higher than that of the health group (P <0.001)。Escherichia-ShigellaIs common bacteria causing bacterial diarrhea, and the relative abundance of the bacteria is reduced from 5.59 percent of disease group to 0.26 percent of recovery group and is lower than 1.16 percent of health group by feeding the bacteria through fermented traditional Chinese medicine oral liquid.
In conclusion, the fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora of the piglet can obviously improve the relative abundance of probiotic bacteria in piglet intestinal tracts, simultaneously improve the relative abundance of intestinal microorganisms related to the digestion and absorption capacity of carbohydrate, the intestinal inflammation resistance and the mucosal immunity capacity, reduce the relative abundance of conditional pathogenic bacteria causing diarrhea, and has good antidiarrheal effect and short treatment period.
Claims (3)
1. A fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora thereof is characterized in that: uniformly mixing 1-1.5 kg of traditional Chinese medicine composition, 0.8-1.2L of compound probiotic bacteria liquid, 0.5-1 kg of brown sugar, 0.3-0.6 kg of poplar flower, adjusting the water content to 55-65%, hermetically fermenting for 6-8 days at 37 ℃ to complete traditional Chinese medicine fermentation, when preparing a traditional Chinese medicine extracting solution, continuously adding 0.8-1.2L of compound probiotic bacteria liquid and 0.5-0.7L of deionized water into the fermented traditional Chinese medicine, standing and soaking at 23-26 ℃ for 24-30 hours after uniformly mixing, stirring irregularly during standing and soaking, and filtering and removing filter residues by using a filter screen to obtain the fermented traditional Chinese medicine oral liquid.
2. The fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora according to claim 1, is characterized in that: the composite probiotic bacteria liquid has effective viable bacteria count of 108~109CFU/mL Lactobacillus salivarius (Lactobacillus salivarius) Bacterial liquid, effective viable count 108~109CFU/mL lactococcus lactis (Lactococcus lactis) Bacterial liquid, effective viable count 108~109CFU/mL Lactobacillus plantarum (Lactobacillus plantarum) The bacterial liquid is prepared from the following components in percentage by volume (0.9-1.2): (0.9-1.1): (0.9-1.1) to obtain a mixed bacterial liquid, and then mixing the mixed bacterial liquid and the effective viable count of 108~109CFU/mL Bacillus licheniformis (C: (C)Bacillus licheniformis) And (3) uniformly mixing the mixed bacterial liquid and the bacillus licheniformis bacterial liquid according to the volume ratio of the bacterial liquid being 4-6: 1.
3. The fermented traditional Chinese medicine oral liquid for treating piglet diarrhea and improving intestinal flora according to claim 1, is characterized in that: the traditional Chinese medicine composition comprises, by weight, 4-6 parts of liquorice, 4-6 parts of coptis chinensis, 5-7 parts of sweet wormwood, 5-7 parts of immature bitter orange, 5-7 parts of hawthorn, 5-7 parts of platycodon grandiflorum, 5-7 parts of honeysuckle stem, 5-7 parts of radix bupleuri, 5-7 parts of dandelion, 5-7 parts of folium isatidis, 5-7 parts of houttuynia cordata, 5-7 parts of scutellaria baicalensis, 5-7 parts of cape jasmine, 5-7 parts of selfheal, 5-7 parts of rhizoma atractylodis, 9-11 parts of golden cypress, 20-25 parts of rhizoma alismatis, 30-35 parts of poria cocos and 30-35 parts of astragalus membranaceus.
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