CN115141766A - Application of composite microecological preparation in preparation of preparation for repairing intestinal tract injury caused by weaning stress of piglets - Google Patents
Application of composite microecological preparation in preparation of preparation for repairing intestinal tract injury caused by weaning stress of piglets Download PDFInfo
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- CN115141766A CN115141766A CN202210600031.5A CN202210600031A CN115141766A CN 115141766 A CN115141766 A CN 115141766A CN 202210600031 A CN202210600031 A CN 202210600031A CN 115141766 A CN115141766 A CN 115141766A
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- 208000027418 Wounds and injury Diseases 0.000 title claims abstract description 22
- 208000014674 injury Diseases 0.000 title claims abstract description 22
- 241000193171 Clostridium butyricum Species 0.000 claims abstract description 31
- 241000186604 Lactobacillus reuteri Species 0.000 claims abstract description 31
- 229940001882 lactobacillus reuteri Drugs 0.000 claims abstract description 31
- 241000186606 Lactobacillus gasseri Species 0.000 claims abstract description 27
- 150000001875 compounds Chemical class 0.000 claims abstract description 12
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- 238000006243 chemical reaction Methods 0.000 claims description 3
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Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
- A23K10/16—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions
- A23K10/18—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions of live microorganisms
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/30—Feeding-stuffs specially adapted for particular animals for swines
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/60—Feeding-stuffs specially adapted for particular animals for weanlings
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2400/00—Lactic or propionic acid bacteria
- A23V2400/11—Lactobacillus
- A23V2400/145—Gasseri
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2400/00—Lactic or propionic acid bacteria
- A23V2400/11—Lactobacillus
- A23V2400/173—Reuteri
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/145—Clostridium
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/225—Lactobacillus
Abstract
The invention discloses application of a composite microecological preparation in preparation of a preparation for repairing intestinal tract injury caused by piglet weaning stress. The composite microecological preparation comprises lactobacillus reuteri, clostridium butyricum and lactobacillus gasseri. The compound microecological preparation comprises lactobacillus gasseri, lactobacillus reuteri and clostridium butyricum, and can be used for feeding weaned pigs in different modes of drinking water, mixing materials or spraying feed, so that the intestinal colonization of the weaned pigs can be realized, the intestinal microbial flora structure and intestinal metabolites can be regulated, harmful bacteria can be inhibited, the generated microbial secondary metabolite butyric acid can improve the feed digestion utilization rate, bacterin can repair intestinal mucosa injury, the immunity of the piglets can be improved, the problems of piglet diarrhea, high mortality, low weaning weight and the like in the current weaned pig breeding are solved, and the urgent need of the sustainable development of the healthy breeding of live pigs is met.
Description
The technical field is as follows:
the invention belongs to the field of livestock feeding, and particularly relates to application of a composite microecological preparation in preparation of a preparation for repairing intestinal injury caused by weaning stress of piglets.
Background art:
the piglet breeding is an important link in the live pig breeding, and the breeding and management in the piglet stage directly influences the indexes of the live pigs, such as slaughter time, slaughter rate, slaughter weight and the like, so that the production benefit of the live pig breeding is influenced. With the continuous improvement of the modernization level of pig raising, at present, domestic and foreign farms improve the annual birth times and the breeding efficiency of sows by shortening the weaning days of piglets (from 35-42 days to 21-28 days or even earlier), and meanwhile, the early weaning of the piglets can improve the utilization rate of a delivery house, the health level of a swinery and save the feed of the sows. However, the digestive system and the immune function of the piglets are not mature in the early weaning period, so that the piglets have the weaning stress problems of diarrhea, intestinal flora disorder, damaged intestinal structure and function and intestinal barrier, reduced immunity, slow weight increment, increased death rate and the like, and huge economic loss is caused to breeding enterprises. At present, the weaning stress of piglets is mainly regulated and controlled by adding antibiotics, high-dose trace elements and the like, so that the problems of antibiotic residue of animal products, intestinal micro-ecological imbalance, increase of drug resistance of pathogenic bacteria and the like are brought while the weaning stress of the piglets is relieved and diarrhea is reduced, the breeding environment is seriously polluted, and the serious threat to the human health is formed. Therefore, the method has the important effects of regulating and controlling intestinal health of piglets to relieve weaning stress, improving the digestibility of nutrient substances, and searching for an environment-friendly and effective antibiotic substitute to realize green and healthy development of animal husbandry and reduce bacterial drug resistance.
Intestinal microorganisms play an important role in regulating and controlling various physiological processes of animal nutrient digestion, intestinal barrier, immune response, endocrine and the like, and intestinal flora imbalance and translocation can cause intestinal cell apoptosis, damage to intestinal physical barrier and intestinal immune dysfunction, so that animals grow and develop slowly, and diarrhea and even death are realized. In vivo and in vitro experimental studies show that the microbial preparation can inhibit harmful bacteria by various mechanisms, including occupying and competing nutrients, directly inhibit specific pathogenic microorganisms, generate bacteriocin, polypeptide and other direct antibacterial compounds, generate short-chain fatty acids, reduce the pH of intestinal tracts, regulate immune response, regulate and control intestinal epithelial gene expression such as mucin and the like, and prevent the harmful microorganisms from invading. The weaning stress of piglets is especially a serious problem of causing the damage of intestinal canal structure and function and intestinal canal barrier, causing enteritis, causing diarrhea of piglets and seriously causing death. Aiming at the problem of intestinal tract injury caused by piglet weaning stress, the invention screens a functional strain capable of repairing the intestinal tract injury caused by piglet weaning stress, prepares a composite microecological preparation for repairing the intestinal tract injury caused by piglet weaning stress by optimizing a fermentation process, and repairs the intestinal tract injury caused by piglet weaning stress by regulating and controlling an intestinal tract flora structure and intestinal tract metabolites of the piglets, thereby solving the problems of piglet diarrhea, high mortality and the like in the current piglet weaning stress stage culture, replacing the use of antibiotics in the culture and promoting the green, healthy and sustainable development of the live pig culture industry.
The invention content is as follows:
the invention aims to provide application of a composite microecological preparation in preparation of a preparation for repairing intestinal injury caused by weaning stress of piglets. The composite microecological preparation can repair intestinal tract injury caused by piglet weaning stress by regulating and controlling piglet intestinal tract flora structures and intestinal tract metabolites, solves the problems of piglet diarrhea, high mortality and the like in the current piglet weaning stress stage culture, replaces the use of antibiotics in the culture, and promotes the green, healthy and sustainable development of the live pig breeding industry.
Therefore, the invention provides an application of a composite microecological preparation in preparing a preparation for repairing intestinal tract injury caused by piglet weaning stress, wherein the composite microecological preparation comprises lactobacillus reuteri, clostridium butyricum and lactobacillus gasseri.
Preferably, the composite microecological preparation is prepared by mixing lactobacillus reuteri liquid, clostridium butyricum liquid and lactobacillus gasseri liquid according to the volume ratio of 300-600:200-400:100-300, and mixing.
Preferably, the viable count of the lactobacillus reuteri bacterial liquid is more than or equal to 1.0 multiplied by 10 9 cfu/mL; the viable count of the clostridium butyricum liquid is more than or equal to 5.0 multiplied by 10 8 cfu/mL; the viable count of the Lactobacillus gasseri liquid is more than or equal to 1.0 multiplied by 10 9 cfu/mL。
Preferably, the composite microecological preparation comprises lactobacillus reuteri (1.0-1.8) multiplied by 10 9 cfu/mL, clostridium butyricum (4.0-6.0). Times.10 8 cfu/mL and Lactobacillus gasseri (1.0-1.8). Times.10 9 cfu/mL。
Preferably, the lactobacillus gasseri is prepared by the following method:
transferring the lactobacillus gasseri seed liquid into a fermentation tank containing a secondary fermentation culture medium for secondary fermentation according to the inoculation amount of 3-8% by volume, wherein the liquid loading amount of the fermentation tank is 70-80%, the fermentation temperature is 30 +/-1 ℃, the fermentation is performed in a closed standing culture mode for 20-24 hours, the fermentation can be finished when the pH value is detected to be reduced to 4.0-4.5 and no change occurs, and thus the lactobacillus gasseri is prepared;
the second-stage fermentation medium contains 20g of molasses, 5g of ammonium nitrate, 3g of beef extract, 5g of yeast powder, 5g of lactose and 5g of sodium chloride per liter, and the pH value is 6.8.
Preferably, the lactobacillus reuteri is prepared by the following method:
transferring the lactobacillus reuteri seed liquid into a fermentation culture medium containing glycerol-producing dehydratase for fermentation by an inoculation amount of 3-6% in volume ratio, wherein the liquid loading amount of a fermentation tank is 50-65%, the fermentation temperature is 28-32 ℃, the micro-aerobic culture is carried out, the rotating speed is 80-100rpm, the pH is adjusted by NaOH, the pH of the whole fermentation enzyme is controlled to be 5.5-7.0, after the fermentation time is 18-25h, the sterilized and cooled glycerol is fed into a bacterial liquid in a variable-speed feeding manner, and the volume ratio of the glycerol solution to the bacterial liquid is as follows: 1-5: 99-95, enzyme conversion temperature: the fermentation can be finished when the pH is detected to be reduced to 4.0-4.5 and is not changed at 28-32 ℃ for 1-5h, thereby preparing the lactobacillus reuteri;
the glycerol producing dehydratase fermentation medium contains 25g of molasses, 4g of beef extract and 4g of yeast powder per liter, and has the pH value of 6.0.
Preferably, the clostridium butyricum is prepared by the following method:
transferring the clostridium butyricum seed liquid into a fermentation tank containing a secondary fermentation culture medium for secondary fermentation by using the inoculation amount of 3-8% of the volume ratio, wherein the liquid loading amount of the fermentation tank is 70-80%, the fermentation temperature is 37 +/-1 ℃, the fermentation is performed in a closed standing culture mode for 40-52 hours, and the fermentation can be finished when the pH is detected to be reduced to 4.0-4.5 and is not changed any more, so that the clostridium butyricum is prepared;
the second-stage fermentation medium contains 3g of peptone, 20g of yeast extract powder, 5g of glucose, 8g of soluble starch, 5g of sodium chloride, 0.2g of magnesium sulfate heptahydrate, 0.5g of L-cysteine hydrochloride and 0.2g of manganese sulfate monohydrate per liter, and the pH value is 7.0 +/-0.2.
The compound microecological preparation comprises lactobacillus gasseri, lactobacillus reuteri and clostridium butyricum, and can be used for feeding weaned pigs in different modes of drinking water, mixing materials or spraying feed, and the like, so that the compound microecological preparation can be planted in the intestinal tracts of the weaned pigs, regulating and controlling the intestinal microbial flora structure and intestinal metabolites, inhibiting harmful bacteria, and producing a microbial secondary metabolite butyric acid which can improve the digestion utilization rate of the feed, repair intestinal mucosa injury by bacterin, improve the immunity of the piglets, solve the problems of diarrhea, high mortality, low weaning weight and the like of the piglets faced by the current weaned pig breeding, and meet the urgent need of the sustainable development of the healthy breeding of live pigs.
Compared with the prior art, the invention has the following advantages:
(1) The invention discovers that the effect of the lactobacillus reuteri, the clostridium butyricum and the lactobacillus gasseri used in a composite way is better than that of a single microbial inoculum, and the lactobacillus gasseri, the lactobacillus reuteri and the clostridium butyricum have different effects.
(2) Compared with the traditional methods of using antibiotics and the like, the composite microecological preparation for repairing intestinal tract injury caused by piglet weaning stress can repair intestinal tract mucosa injury, improve piglet immunity, reduce diarrhea and death rate of weaned piglets and improve the feed digestion and utilization rate.
(3) The invention relates to a composite microbial ecological agent for repairing intestinal injury caused by piglet weaning stress, belonging to a green environment-friendly animal microbial agent.
The specific implementation mode is as follows:
the following examples are further illustrative of the present invention and are not intended to be limiting thereof.
1. Preparation method of lactobacillus gasseri for repairing intestinal injury caused by piglet weaning stress
1 Strain activation
Firstly, preparing a strain, aseptically starting a freeze-dried preserved strain lactobacillus gasseri, streaking and inoculating the strain to an improved MRS agar test tube inclined plane, and culturing for 22-28h at 30 ℃; then transferring the strain into an improved MRS liquid culture medium, and statically culturing the strain for 24 to 30 hours at the temperature of 30 ℃ to form a triangular flask liquid strain.
2 amplification culture
2.1 first order amplification culture
Transferring the triangular flask liquid strain into a 30L seed tank containing a primary fermentation culture medium in an inoculation amount of 3-6% by volume, wherein the liquid loading amount of the fermentation tank is 70-80%, the fermentation temperature is 30 +/-1 ℃, and the fermentation time is 18-24h, and preparing the primary seed liquid.
2.2 two-stage amplification culture
Transferring the primary seed liquid into a 300L fermentation tank containing a secondary fermentation culture medium by an inoculation amount of 3-8% in volume ratio for secondary fermentation, wherein the liquid loading amount of the fermentation tank is 70-80%, the fermentation temperature is 30 +/-1 ℃, the fermentation is performed in a closed standing culture mode for 20-24h, and the fermentation can be finished when the pH is detected to be reduced to 4.0-4.5 and not to be changed any more. Thus preparing the lactobacillus gasseri liquid microbial preparation.
3 culture Medium
3.1 the improved MRS liquid culture medium has the following first-level fermentation culture media: in a 1L system: adding 10g of peptone, 5g of yeast powder, 10g of beef extract, 5g of glucose, 5g of sodium acetate, 2g of diammonium hydrogen citrate, 2g of dipotassium hydrogen phosphate, 0.2g of magnesium sulfate, 0.05g of manganese sulfate, 801mL of tween and 10g of calcium carbonate into 1000mL of water, and adjusting the pH value to 6.8; sterilizing at 115 deg.C for 30min. The agar medium was 20g/L agar added.
3.2 the secondary fermentation culture medium comprises: in a 1L system: adding molasses 20g, ammonium nitrate 5g, beef extract 3g, yeast powder 5g, lactose 5g, and sodium chloride 5g into water 1000ml, and adjusting pH to 6.8; sterilizing at 115 deg.C for 30min.
2. Preparation method of lactobacillus reuteri for repairing intestinal injury caused by weaning stress of piglets
1 Strain activation
Firstly, preparing a strain, starting a freeze-dried preserved strain lactobacillus reuteri in an aseptic mode, streaking and inoculating the strain to an MRS agar test tube inclined plane, and culturing for 24-28h at 37 ℃; then transferring the strain into an MRS liquid culture medium, and carrying out static culture at 37 ℃ for 20-30h to form a triangular flask liquid strain.
2 amplification culture
2.1 first order amplification culture
Transferring the triangular flask liquid strain into a 30L seed tank containing a primary fermentation culture medium by an inoculation amount of 3-6% by volume, wherein the liquid loading amount of the fermentation tank is 70-80%, the fermentation temperature is 37 +/-1 ℃, and the fermentation time is 16-20h, and preparing primary seed liquid;
2.2 two-stage amplification culture
Transferring the first-stage seed liquid into a fermentation medium containing glycerol-producing dehydratase for fermentation by using an inoculation amount of 3-6% in volume ratio, wherein the liquid loading amount of a fermentation tank is 50-65%, the fermentation temperature is 28-32 ℃, the micro-aerobic culture is carried out, the rotating speed is 80-100rpm, the pH is adjusted by using 6mol/L NaOH, the pH of the whole fermentation enzyme is controlled to be 5.5-7.0, after the fermentation time is 18-25h, sterilized and cooled glycerol (the concentration of the glycerol is 150-450 mmol/L) is fed into a bacterial liquid in a variable-speed feeding mode, and the volume ratio of the glycerol solution to the bacterial liquid is as follows: 1-5: 99-95, enzyme conversion temperature: at 28-32 ℃ for 1-5h. The fermentation can be terminated by detecting the pH drop to 4.0-4.5 and no further change. Thus preparing the liquid microbial preparation of the lactobacillus reuteri.
3 culture Medium
3.1, the first-level fermentation culture medium of the MRS liquid culture medium is: in a 1L system: adding 10g of peptone, 5g of yeast powder, 10g of beef extract, 5g of sodium acetate, 2g of diammonium hydrogen citrate, 2g of dipotassium hydrogen phosphate, 0.2g of magnesium sulfate, 0.05g of manganese sulfate, 801mL of tween and 10g of calcium carbonate into 1000mL of water, and adjusting the pH value to 6.8; sterilizing at 115 deg.C for 30min. The agar medium was 20g/L agar added.
3.2 the glycerol producing dehydratase fermentation medium is: in a 1L system: adding 25g of molasses, 4g of beef extract and 4g of yeast powder into 1000ml of water at the pH of 6.0, and sterilizing at 115 ℃ for 30min.
Preparation method of clostridium butyricum for repairing intestinal injury caused by weaning stress of piglets
1 Strain activation
Firstly, preparing a strain, starting a freeze-dried preserved strain clostridium butyricum aseptically, streaking and inoculating the strain to an RCM agar test tube inclined plane, and carrying out anaerobic culture at 37 ℃ for 18-24h; then transferring the strain into an RCM liquid culture medium, and carrying out anaerobic static culture at 37 ℃ for 20-28h to form a triangular flask liquid strain.
2 amplification culture
2.1 first order amplification culture
Transferring the triangular flask liquid strain into a 30L seed tank containing a primary fermentation culture medium at an inoculum size of 3-6% by volume, wherein the liquid loading capacity of the fermentation tank is 70-80%, the fermentation temperature is 37 +/-1 ℃, and performing closed static culture for 18-24h to prepare a primary seed solution.
2.2 two-stage amplification culture
Transferring the primary seed liquid into a 300L fermentation tank containing a secondary fermentation culture medium by an inoculation amount of 3-8% in volume ratio for secondary fermentation, wherein the liquid loading amount of the fermentation tank is 70-80%, the fermentation temperature is 37 +/-1 ℃, the fermentation is performed in a closed standing culture mode for 40-52h, and the fermentation can be finished when the pH is detected to be reduced to 4.0-4.5 and not to be changed any more. Thereby preparing the clostridium butyricum liquid microbial preparation.
3 culture Medium
3.1 the RCM culture medium and the first-stage fermentation culture medium are all as follows: in a 1L system
Adding water to 1000ml, pH7.0 + -0.2; sterilizing at 115 deg.C for 30min.
The agar medium was 20g/L agar added.
3.2 the secondary fermentation medium comprises: in a 1L system
Adding water to 1000ml, pH7.0 + -0.2; sterilizing at 115 deg.C for 30min
The four-repairing piglet weaning stress induced intestinal injury three functional microbial inoculums (lactobacillus reuteri, clostridium butyricum and lactobacillus gasseri) are optimally combined, and the total amount is calculated by 1L:
300-600mL of lactobacillus reuteri (viable count is more than or equal to 1.0 multiplied by 10) 9 cfu/mL), 200-400mL of clostridium butyricum (viable count is more than or equal to 5.0 multiplied by 10) 8 cfu/mL) and 100-300mL of Lactobacillus gasseri (viable count is more than or equal to 1.0 multiplied by 10) 9 cfu/mL)。
In the following examples, the liquid microbial preparations of Lactobacillus reuteri, clostridium butyricum and Lactobacillus gasseri described above were each adjusted or concentrated to a concentration that satisfies the requirements by using a medium used in the secondary scale-up culture.
Example 1:
the preparation method of the composite microecological preparation for repairing intestinal injury caused by piglet weaning stress comprises the following steps:
in terms of 1L: the liquid microbial preparations of the lactobacillus reuteri, the clostridium butyricum and the lactobacillus gasseri are respectively 500mL, 300mL and 200mL and are mixed to obtain the composite microbial ecological preparation for repairing the intestinal injury caused by the weaning stress of the piglets, and the total viable count of the composite microbial ecological preparation is 3.1 multiplied by 10 9 cfu/mL。
Cultivation test:
the lactobacillus reuteri liquid microbial preparation, the clostridium butyricum liquid microbial preparation, the lactobacillus gasseri liquid microbial preparation and the composite microbial preparation prepared by the invention are uniformly sprayed on a weaned pig basic feed according to the proportion of 1 percent by mass to feed weaned pigs, the experimental conditions of each group are consistent, and each group contains 10 weaned pigs.
After 21d cultivation test, the result shows that: compared with Ctr0 (the diarrhea rate of Ctr0 group is 25.89%, the mortality rate is 14.08%, the height of duodenum villi is 398.11 μm, the crypt depth is 234.54 μm, the height of jejunum villi is 487.14 μm, the crypt depth is 198.78 μm, the height of ileum villi is 239.56 μm, and the crypt depth is 165.60 μm), the diarrhea rate of the test group I, II, III, IV fed with the microecological preparation is respectively reduced by 39.35%, 42.16%, 46.10% and 89.797% compared with the control group, the mortality rate is respectively reduced by 38.12%, 32.04%, 40.33% and 71.27% compared with Ctr0, the height of Ctduodenum villi is respectively increased by 12.95%, 9.12%, 11.97% and 37.99% compared with Ctr0, the crypt depth of duodenum is respectively reduced by 15.08%, 14.96%, 15.78% and 28.30%, the height of jejunum villi is respectively increased by 12.70.9.9%, and 13.9.9.99%, and 13.9.9% compared with Ctr 0%, and 13.9.9.9% respectively compared with Ctr 0%. The results show that: the compound microbial preparation for repairing intestinal tract injury caused by piglet weaning stress can reduce the diarrhea rate of weaned piglets, improve the survival rate of the weaned piglets and have obvious repairing effect on the injury of the intestinal tract structure, and the effect of the compound microbial preparation is superior to that of a single microbial preparation.
Example 2:
the preparation method of the composite microecological preparation for repairing intestinal injury caused by piglet weaning stress comprises the following steps:
in terms of 1L: the liquid microbial preparations of the lactobacillus reuteri, the clostridium butyricum and the lactobacillus gasseri are respectively 300mL, 400mL and 300mL and are mixed to prepare the composite microecological preparation for repairing the intestinal injury caused by the weaning stress of the piglets, and the total viable count of the composite microecological preparation is 2.6 multiplied by 10 9 cfu/mL。
Cultivation test:
the lactobacillus reuteri liquid microbial preparation, the clostridium butyricum liquid microbial preparation, the lactobacillus gasseri liquid microbial preparation and the composite microbial preparation prepared by the invention are uniformly sprayed on a weaned pig basic feed according to the proportion of 1 percent by mass to feed weaned pigs, the experimental conditions of each group are consistent, and each group contains 10 weaned pigs.
After 21d cultivation test, the result shows that: compared with Ctr0 (the diarrhea rate of Ctr0 group is 23.77%, the fatality rate of disease is 13.19%, the content of intestinal mucosa proinflammatory factors IL-1 beta is 3709.41pg/g, the content of IL-2 is 423.78pg/g, the content of IL-6 is 291.15pg/g, the content of IL-10 is 149.36 pg/g), the diarrhea rate of the weaned piglets fed with the microecological preparation in the test groups I, II, III and IV is respectively reduced by 51.78%, 48.61%, 49.33% and 81.43% compared with the control group, the fatality rate is respectively reduced by 48.17%, 41.88%, 45.03% and 83.25% compared with Ctr0, the content of the intestinal mucosa proinflammatory factors IL-1 beta is respectively reduced by 23.81%, 28.57%, 19.05% and 47.62% compared with the control group, the content of IL-2 is respectively reduced by 18.62%, 15.58%, 16.35% and 44.55% compared with the control group, the content of IL-6 is respectively reduced by 23.03%, and the content of the anti-inflammatory factors is respectively increased by 23.35.59%, and 20.35.55% compared with the control group, and the content of IL-6 is respectively increased by 20.35.35.35.35.35%. The results show that: the compound microbial preparation for repairing intestinal tract injury caused by piglet weaning stress can reduce the diarrhea rate of weaned piglets, improve the survival rate of the weaned piglets and have obvious repairing effect on the injury of intestinal mucosa, and the effect of the compound microbial preparation is superior to that of a single microbial preparation.
Example 3:
the preparation method of the composite microecological preparation for repairing intestinal injury caused by piglet weaning stress comprises the following steps:
in terms of 1L: mixing 600mL, 200mL and 200mL of microbial preparations of Lactobacillus reuteri, clostridium butyricum and Lactobacillus gasseri respectively to obtain the productCompound microecological preparation for treating intestinal tract injury caused by weaning stress of piglets, with total viable count of 4.2 × 10 9 cfu/mL。
And (3) breeding test:
uniformly spraying the liquid microbial preparation of lactobacillus reuteri, the liquid microbial preparation of clostridium butyricum, the liquid microbial preparation of lactobacillus gasseri and the composite microbial preparation prepared by the invention on a basic feed of weaned pigs according to the proportion of 1 percent by mass for feeding the weaned pigs, wherein the experimental conditions of all groups are consistent, and 10 weaned pigs are fed in each group.
After 21d cultivation test, the result shows that: compared with Ctr0 (the diarrhea rate of Ctr0 group is 24.11%, the fatality rate of disease is 13.68%, the daily gain rate of weight is 14.57%, the jejunal mucosa diamine oxidase activity is 171.82U/kg, and the ileal mucosa diamine oxidase activity is 62.63U/kg), the diarrhea rates of the weaned piglets of the test groups I, II, III and IV fed with the microecological preparation are respectively reduced by 51.83%, 48.93%, 49.48% and 85.71% compared with the control group, the fatality rates are respectively reduced by 46.77%, 44.78%, 48.76% and 81.59% compared with Ctr0, the daily gain rates are respectively improved by 35.31%, 38.63%, 41.05% and 71.21% compared with Ctr0, the jejunal mucosa diamine oxidase activity is respectively reduced by 42.03%, 43.91%, 44.28% and 70.21% compared with Ctr0, and the ileal mucosa diamine oxidase activity is respectively reduced by 43.93%, 46.51%, 42.74% and 70.17% compared with Ctr 0. The results show that: the compound microbial preparation for repairing intestinal tract injury caused by piglet weaning stress can reduce the diarrhea rate of the weaned piglets, improve the daily gain and survival rate of the weaned piglets, and has obvious repairing effect on the injury of intestinal mucosa, and the effect of the compound microbial preparation is superior to that of a single microbial agent.
Claims (7)
1. The application of the composite microecological preparation in preparing the preparation for repairing intestinal tract injury caused by piglet weaning stress is characterized in that the composite microecological preparation comprises lactobacillus reuteri, clostridium butyricum and lactobacillus gasseri.
2. The use according to claim 1, wherein the composite microecological preparation is prepared by mixing the lactobacillus reuteri liquid, the clostridium butyricum liquid and the lactobacillus gasseri liquid according to a volume ratio of 300-600:200-400:100-300, and mixing.
3. The application of repairing intestinal injury caused by piglet weaning stress according to claim 2, wherein the viable count of the lactobacillus reuteri bacterial liquid is more than or equal to 1.0 x 10 9 cfu/mL; the viable count of the clostridium butyricum liquid is more than or equal to 5.0 multiplied by 10 8 cfu/mL; the viable count of the Lactobacillus gasseri liquid is more than or equal to 1.0 multiplied by 10 9 cfu/mL。
4. The use of the compound microecological preparation for repairing intestinal tract injury caused by piglet weaning stress according to claim 1, wherein the compound microecological preparation comprises lactobacillus reuteri (1.0-1.8) x 10 9 cfu/mL, clostridium butyricum (4.0-6.0). Times.10 8 cfu/mL and Lactobacillus gasseri (1.0-1.8). Times.10 9 cfu/mL。
5. The use according to claim 1, 2, 3 or 4, wherein the Lactobacillus gasseri is prepared by the following method:
transferring the lactobacillus gasseri seed liquid into a fermentation tank containing a secondary fermentation culture medium by an inoculation amount of 3-8% in volume ratio for secondary fermentation, wherein the liquid loading amount of the fermentation tank is 70-80%, the fermentation temperature is 30 +/-1 ℃, the fermentation time is 20-24 hours, and the fermentation can be finished when the pH is detected to be reduced to 4.0-4.5 and is not changed any more, so that the lactobacillus gasseri is prepared;
the second-stage fermentation medium contains 20g of molasses, 5g of ammonium nitrate, 3g of beef extract, 5g of yeast powder, 5g of lactose and 5g of sodium chloride per liter, and the pH value is 6.8.
6. The use according to claim 1, 2, 3 or 4, wherein the Lactobacillus reuteri strain is prepared by the following method:
transferring the lactobacillus reuteri seed liquid into a fermentation culture medium containing glycerol-producing dehydratase for fermentation by an inoculation amount of 3-6% in volume ratio, wherein the liquid loading amount of a fermentation tank is 50-65%, the fermentation temperature is 28-32 ℃, the micro-aerobic culture is carried out, the rotating speed is 80-100rpm, the pH is adjusted by NaOH, the pH of the whole fermentation enzyme is controlled to be 5.5-7.0, after the fermentation time is 18-25h, the sterilized and cooled glycerol is fed into a bacterial liquid in a variable-speed feeding manner, and the volume ratio of the glycerol solution to the bacterial liquid is as follows: 1-5: 99-95, enzyme conversion temperature: the fermentation can be finished when the pH is detected to be reduced to 4.0-4.5 and is not changed at 28-32 ℃ for 1-5h, thereby preparing the lactobacillus reuteri;
the glycerol producing dehydratase fermentation medium contains 25g of molasses, 4g of beef extract and 4g of yeast powder per liter, and has the pH value of 6.0.
7. The use according to claim 1, 2, 3 or 4, wherein the Clostridium butyricum is prepared by the following method:
transferring the clostridium butyricum seed liquid into a fermentation tank containing a secondary fermentation culture medium for secondary fermentation by using the inoculation amount of 3-8% of the volume ratio, wherein the liquid loading amount of the fermentation tank is 70-80%, the fermentation temperature is 37 +/-1 ℃, the fermentation is performed in a closed standing culture mode for 40-52 hours, and the fermentation can be finished when the pH is detected to be reduced to 4.0-4.5 and is not changed any more, so that the clostridium butyricum is prepared;
the second-stage fermentation medium contains 3g of peptone, 20g of yeast extract powder, 5g of glucose, 8g of soluble starch, 5g of sodium chloride, 0.2g of magnesium sulfate heptahydrate, 0.5g of L-cysteine hydrochloride and 0.2g of manganese sulfate monohydrate per liter, and the pH value is 7.0 +/-0.2.
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| CN101974463A (en) * | 2010-10-19 | 2011-02-16 | 黑龙江八一农垦大学 | Lactobacillus reuteri and composite viable bacteria preparation thereof |
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