CN103555640B - Bacillus subtilis and application thereof to livestock breeding - Google Patents
Bacillus subtilis and application thereof to livestock breeding Download PDFInfo
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Abstract
The invention aims to provide bacillus subtilis WL-04 and application thereof to livestock breeding. The preservation number of the bacillus subtilis WL-04 is CCTCC NO:M 2013343. The bacillus subtilis WL-04 can be used for remarkably inhibiting the growth of Escherichia coli, salmonella, clostridium pefringens and other conditional pathogenic bacteria of the intestinal canal of livestock, and particularly, the bacteriostasis rate of the bacillus subtilis WL-04 to Escherichia coli and clostridium perfingens is above 75%; the enzyme production is stable; the average activity of alpha-amylase is 22U/mL, and the average activity of neutral protease is 170U/mL; the reproduction of probiotics of intestinal canal can be promoted; high temperature for pelletizing can be endured; the influence of inverse environments with low-pH gastric fluid and high-concentration cholate is endured; the production performance of breeding animals can be effectively improved; immune organ development can be promoted. The bacillus subtilis WL-04 can be widely applied to livestock feeds as a feed additive.
Description
Technical field
The invention belongs to microbe to screen and applied technical field, be specifically related to a kind of subtilis WL-04 bacterial strain and the application as fodder additives thereof.
Background technology
Nowadays; mass-producing and intensive culture development are rapidly; there is the problem of some breeding environment Quality Downs such as stocking density is excessive, breeding house environment ventilation is bad thereupon; the puzzlement of the various disease of alimentary tract of livestock husbandry and birds caused by conditionality malignant bacteria to cultivation is also more and more outstanding, as the necrotic enteritis etc. that colibacillosis, salmonellosis, clostridium perfringens cause.For reaching the object of disease preventing and treating or promotion growth of animal, a large amount of microbiotic is applied to multiple links of Production of Livestock and Poultry.Antibiotic life-time service and abuse bring many negative effects and serious consequence, as low in fowl poultry immune, bacterial drug resistance, medicine be residual the problem such as to exceed standard, particularly some pathogenic bacterium are zoonosis type, if produce the superbacteria of superpower resistance, serious public health security problem will be brought.Therefore, in breeding production few with or forbid microbiotic and become social consensus.
Through technological development for many years and accumulation, probiotics micro-ecological formulation becomes the first-selection of substitute antibiotics.Probiotics self secretion many kinds of substance, the growth of antagonism pathogenic micro-organism, regulates animal immune, to have no drug resistance, noresidue, deeply popular to people.Wherein genus bacillus the most widely used a kind of probiotics especially.
But the prebiotic performance of different Bacillus strain differs greatly.At present, studying more is take single performance as sieve bacterium index, such as investigates bacteriostatic activity or enzymatic productivity etc. merely, to bacterial strain overall performance to consider report less.
Summary of the invention
The object of this invention is to provide a bacillus subtilis WL-04 and the application in livestock and poultry cultivation thereof, this bacterium has remarkable restraining effect to animal and bird intestines conditionality pathogenic bacterium colibacillosis, salmonellosis, clostridium perfringens, and has promoter action to the breeding of probiotic bacterium; This bacterium can produce multiple enzyme, gemma form can tolerate the adverse environments such as the high temperature in the artificial hydrochloric acid in gastric juice of higher concentration, cholate and feed granulating process, it can be used as fodder additives to be applied in animal and fowl fodder, the outburst effectively can taking precautions against livestock and poultry intestinal canal diseases also can improve the production performance of cultivated animals.
One aspect of the present invention provides a bacillus subtilis WL-04(Bacillus subtilis WL-04), be preserved in the China typical culture collection center of Wuhan, China Wuhan University on July 16th, 2013, its deposit number is CCTCC NO:M2013343.
Subtilis WL-04 of the present invention is in the application of feed additive field.
Another aspect of the present invention provides a kind of microbial preparation comprising above-mentioned subtilis.
Above-mentioned microbial preparation is the bacterium powder of subtilis WL-04, by mass percentage for the ratio of 0.1%-1% is added in animal and fowl fodder.
Subtilis WL-04 of the present invention, has significant restraining effect to the growth of animal and bird intestines conditionality pathogenic bacterium as intestinal bacteria, Salmonellas, clostridium perfringens, wherein to intestinal bacteria and clostridium perfringens bacteriostasis rate higher, reach more than 75%; Produce enzyme more stable, alpha-amylase activity average out to 22U/mL; Neutral protease vigor average out to 170U/mL; Promote the breeding of beneficial bacteria of intestinal tract; Feed granulating high temperature can be tolerated; Tolerate the impact in inverse ring border of low pH gastric juice, high density cholate, and effectively can improve the production performance of cultivated animals, Promote immunity allelotaxis, can be used as fodder additives and be widely used in animal and fowl fodder.
Embodiment
The invention provides a strain and there is Efficient antibacterial activity (can effectively suppress colibacillosis, salmonellosis, clostridium perfringens), the stable genus bacillus producing amylase and proteolytic enzyme, antibiotic additive agent for feeding as an alternative, can play a role in animal intestinal, balance intestinal flora, suppresses the outburst of enteron aisle conditionality pathogenic bacterium, secretes multiple enzyme and VITAMIN etc., thus improve the production performance of animal, safeguard that animal body is healthy.
Below in conjunction with embodiment, bacterial strain of the present invention is described in detail.
Embodiment 1 subtilis WL-04 Isolation and ldentification
1, the separation screening of genus bacillus
By the mud gathered in Pingdu broiler breeding field, Qingdao City, add in sterilized water with the ratio of 1:10, vibration, mixing, be prepared into suspension, through 80 DEG C of water bath processing 10min, kill most of heat labile bacterial strain, the preliminary genus bacillus obtaining good heat resistance.
2, there is the genus bacillus screening suppressing pathogenic bacteria and high enzymatic productivity
1) genus bacillus bacterium solution preparation: by the genus bacillus be separated, be inoculated in respectively in nutrient broth, 37 DEG C, 200rpm, shaking culture 24h;
2) antibacterial experiment in vitro-micro-meat soup method: by centrifugal for the bacillus liquid cultivating 24h, collection supernatant, get aseptic 96 well culture plates, genus bacillus supernatant liquor and pathogenic bacteria liquid (testing pathogenic bacteria used is intestinal bacteria (ATCC25922), Salmonellas (ATCC14028), clostridium perfringens (ATCC13124)) is added in 1:1 ratio, at 37 DEG C, jointly 6h is hatched under 200rpm condition, calculate bacteriostasis rate, formula is as follows:
A is control group OD
600, A0 is 0h data; A2 is 6h data;
B is genus bacillus supernatant treatment group OD
600, B0 is 0h data; B2 is 6h data;
3) enzymatic productivity measures: by centrifugal for the bacillus liquid cultivating 24h, collection supernatant, carry out the detection (GB/T24401-2009) of alpha-amylase activity, and the detection of neutral protease vigor (GB/T28715-2012);
4) combine the situation analysis suppressing pathogenic bacteria and enzymatic productivity, be finally separated to a bacillus and do to investigate further.
3, Bacillus Determination
1) above-mentioned genus bacillus physiological and biochemical property is as shown in table 1:
Table 1: the genus bacillus physiological and biochemical analysis result of separation
2) utilize test kit to extract above-mentioned subtilis genomic dna, utilize round pcr amplification 16S rRNA sequence, obtaining length after order-checking is the fragment of 1406bp.By BLAST compare of analysis, this bacterium and subtilis homology the highest, in conjunction with Physiology and biochemistry result, identify that this genus bacillus is subtilis.By this bacterium called after subtilis WL-04(Bacillus subtilis WL-04), and on July 16th, 2013, be preserved in the China typical culture collection center of Wuhan, China Wuhan University, deposit number is CCTCC NO:M2013343.
Embodiment 2: subtilis WL-04 suppresses animal source Analysis of pathogenic bacteria
1, Bacteria culturing is indicated: random choose is separated the pathogenic bacteria from sick chicken, chitling road, intestinal bacteria, Salmonellas, clostridium perfringens, totally 5 strains (Qingdao Agricultural University's Preventive Veterinary Medicine laboratory is so kind as to give), wherein intestinal bacteria, Salmonellas are inoculated in nutrient broth, 37 DEG C, 200rpm, cultivates 15h; Clostridium perfringens is inoculated in cooked meat medium, 37 DEG C, cultivates 15h;
2, subtilis WL-04 bacterium liquid is prepared: picking list bacterium colony, is inoculated in nutrient broth, 37 DEG C, 200rpm, cultivates 24h;
3, antibacterial experiment in vitro-micro-meat soup method: by centrifugal for the bacillus liquid cultivating 24h, collection supernatant, get aseptic 96 well culture plates, add genus bacillus supernatant liquor and pathogenic bacteria liquid in 1:1 ratio, at 37 DEG C, jointly hatch 6h under 200rpm condition, calculate bacteriostasis rate, formula is as follows:
A is control group OD
600, A0 is 0h data; A2 is 6h data;
B is genus bacillus supernatant treatment group OD
600, B0 is 0h data; B2 is 6h data;
Subtilis WL-04 is as shown in table 2 to the zoogenous intestinal bacteria of 4 strain, Salmonellas, clostridium perfringens growth-inhibiting situation:
Table 2: the subtilis WL-04 bacteriostatic experiment result that the present invention is separated
+++ represent that bacteriostasis rate is greater than 75%; ++ represent that bacteriostasis rate is between 50-75%;
From antibacterial experiment in vitro result in upper table, subtilis WL-04 of the present invention significantly suppress the growth be separated from zoogenous 5 pathogen strain bacterium, wherein to 2 strain intestinal bacteria and 1 strain clostridium perfringens bacteriostasis rate higher, reach more than 75%, show that subtilis WL-04 has significant restraining effect to animal source intestinal bacteria, Salmonellas, clostridium perfringens.
Embodiment 3: subtilis WL-04 produces enzyme performance analysis
1, subtilis WL-04 bacterium liquid is prepared: picking list bacterium colony, is inoculated in nutrient broth, 37 DEG C, 200rpm, cultivates 24h;
2, obtaining fermented supernatant fluid: by cultivating the bacillus liquid of 24h, centrifugal under 10000rpm, 10min condition, collecting supernatant;
3, produce enzyme situation: the detection (GB/T24401-2009) carrying out alpha-amylase activity, and the detection of neutral protease vigor (GB/T28715-2012), carry out 3 batches of mensuration altogether, result is as shown in table 3;
Table 3: subtilis WL-04 produces enzyme analysis
Test again through three batch weight, carry out enzyme activity determination analysis, it is more stable that subtilis WL-04 produces enzyme, alpha-amylase activity average out to 22U/mL; Neutral protease vigor average out to 170U/mL.In feed, macromolecules starch particle, protein are difficult to directly be absorbed by animal and utilize, and these macromole can be decomposed into the small molecules nutritive substance easily absorbed by α-amylase and neutral protease effectively.Subtilis WL-04 of the present invention stably can secrete high enzyme α-amylase alive and neutral protease, therefore, is expected to be developed as fodder additives, for improving the utilization ratio of starch in feed, protein.
Embodiment 4: subtilis WL-04 resistance is analyzed
1, subtilis WL-04 bacterium liquid viable bacteria gauge number (control group): by bacterium liquid stroke-physiological saline solution ten times of gradient dilutions, selects the extent of dilution bacterium liquid be applicable to, coats on nutrient agar plate and cultivate, counting viable bacteria;
2, Analysis of Heat Tolerance: get subtilis WL-04 bacterium liquid 1mL and join in the triangular flask containing 99mL sterilized water, respectively after bath temperature 75 DEG C, 80 DEG C, 95 DEG C process 10min, totally 3 treatment group, counting viable bacteria;
3, stomach juice-resistant analysis: get subtilis WL-04 bacterium liquid 1mL and join respectively in the triangular flask of the artificial simulation gastric juices of the different pH of 99mL, pH is followed successively by 1,2,3, totally 3 treatment group, in 37 DEG C of water bath processing 2h, counting viable bacteria;
4, bile tolerance analysis: get subtilis WL-04 bacterium liquid 1mL and join in the triangular flask of the artificial simulation intestinal juice of 99mL various biliary salt concn, gallbladder salinity is followed successively by 0.3%, 0.5%, 1%, totally 3 treatment group, in 37 DEG C of water bath processing 4h, counting viable bacteria;
Table 4: subtilis WL-04 resistance toheat is analyzed
As shown in Table 4, subtilis WL-04 all can keep high survival rate in different high temperature bath process, illustrates that this bacterium is to hot environment better tolerance.
Table 5: subtilis WL-04 tolerates hydrochloric acid in gastric juice, cholate analysis
As shown in Table 5, after the artificial simulation gastric juices process of different pH, when pH >=3, subtilis WL-04 survival rate is 100.0%, as pH=2, subtilis WL-04 survival rate slightly reduces, and is 99.2%, grow up cultivated animals if the Gastric pH scope of pig, chicken is about 2 ~ 4 fluctuations, and experiment shows that subtilis WL-04 acid resistance is good.
Through the process of the artificial simulation intestinal juice of various biliary salt concn, along with the increase of gallbladder salinity, subtilis WL-04 survival rate has certain decline, but survival rate is all more than 95%, illustrates that this bacterium has the ability of very strong tolerance cholate.
In feeding animals, active bacteria formulation must experience competence exertion effect after the barrier of hydrochloric acid in gastric juice and enteron aisle cholate.Subtilis WL-04 of the present invention can tolerate the inverse ring border of acid and cholate, contributes to its widely using in feed additive field.
Embodiment 5: subtilis WL-04 is on the impact of growth of meat chicken performance and Development of Immune Organs
Purchase the white plumage meat cock of the substantially identical Roche 308 of 1 age in days healthy state, be divided into 3 groups at random, comprise 1 control group and 2 experimental group, each treatment group 16 repetition, each repetition 8 plumage chicken, the production test phase is 35 days, wherein control group is general goods daily ration of feeding, experimental group is the general goods daily ration of subtilis WL-04 bacterium powder of Different adding amount of feeding, bacterium powder addition is 0.1%(mass ratio), 0.5%(mass ratio), measure 1) changing conditions such as food consumption, feed-weight ratio, body weight in process of the test; 2) observe broiler chicken Development of Immune Organs situation, experimental result is as shown in table 6:
Table 6: the bacterium powder feeding dorking experiment statistics result made
By broiler chicken 0-35 days feeding experiment results of table 6, compare with day weight gain and feedstuff-meat ratio, the experimental group 1 and 2 of adding subtilis WL-04 is all better than control group, but between two experimental group, difference is not remarkable, illustrates that the subtilis WL-04 that feeds can promote growth of meat chicken; And in Immune Organs Index, experimental group 2 is optimum, be secondly experimental group 1, control group is the poorest, illustrates that subtilis addition is 0.5% to 0.1% development growth more contributing to broiler chicken immune organ.
Comprehensive analysis, feed after adding subtilis WL-04, well improve the production performance of broiler chicken, and addition 0.1% effectively can improve the production efficiency of broiler chicken, add the economic benefit of raiser, when addition reaches 0.5%, more remarkable to the Development of Immune Organs promoter action of broiler chicken.
Embodiment 6: the impact of subtilis WL-04 on Intestine of Broiler Flora dynamics of feeding
Purchase the white plumage meat cock of the substantially identical Roche 308 of 1 age in days healthy state, be divided into 3 groups at random, comprise 1 control group and 2 experimental group, each treatment group 16 repetition, each repetition 8 plumage chicken, control group is general goods daily ration of feeding, experimental group is the general goods daily ration of subtilis WL-04 bacterium powder of Different adding amount of feeding, bacterium powder addition is 0.1%(mass ratio), 0.5%(mass ratio), 1-7d is for raise the phase in advance, test period is 14d, 21d, 28d, collect fresh chicken manure sample, measure wherein microorganism species changing conditions, experimental result is as shown in table 7:
Table 7: the intestinal microflora change of subtilis WL-04 feeding dorking test
From table 7 test-results, in administering transgenic, 1) the bacterium amount change detection process of probiotic lactic acid bacillus, experimental group 1 and 2 bacterium amount is significantly higher than control group, and between two experimental group, difference is not remarkable; 2) enteric pathogenic bacteria intestinal bacteria, Salmonellas and clostridium perfringens bacterium amount change detection process, experimental group 1 and 2 bacterium amount is significantly lower than control group, and between two experimental group, difference is not remarkable.Explanation is fed in process, has promoter action, and plays restraining effect to enteron aisle conditionality pathogenic bacterium, be obviously better than control group after adding subtilis WL-04 of the present invention to the proliferation of probiotics of Intestine of Broiler.Add 0.1% or 0.5% subtilis WL-04 microbial inoculum feeding dorking, in regulating intestinal canal flora, show good result.
Claims (3)
1. a subtilis, is characterized in that, described subtilis deposit number is CCTCC NO:M 2013343.
2. a microbial preparation, described microbial preparation comprises subtilis according to claim 1.
3. microbial preparation as claimed in claim 2, it is characterized in that, described microbial preparation is the bacterium powder of subtilis according to claim 1.
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| CN105039223B (en) * | 2015-08-14 | 2018-05-25 | 山东宝来利来生物工程股份有限公司 | One plant has the bacillus subtilis for inhibiting C.perfringens effect and its application |
| CN109337841B (en) * | 2017-12-27 | 2021-01-12 | 山东农业大学 | Bacillus subtilis BYS2 with efficient antibacterial performance |
| CN109679879B (en) * | 2019-01-25 | 2021-07-30 | 石河子大学 | Bacterial strain, microbial inoculum and application |
| CN110272854B (en) * | 2019-07-29 | 2021-07-27 | 北京林业大学 | Bacillus subtilis strain and application thereof |
| SG10201910683UA (en) * | 2019-11-14 | 2021-06-29 | Agency Science Tech & Res | An isolated bacillus subtilis strain, uses thereof and processes using said isolated strain |
| CN112760251A (en) * | 2021-01-05 | 2021-05-07 | 安徽农业大学 | Compound bacillus probiotics and preparation and application thereof |
| CN113151069B (en) * | 2021-04-01 | 2022-07-26 | 安杰利(重庆)生物科技有限公司 | Bacillus subtilis and application thereof in preparation of antibacterial peptide and feed |
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103039698A (en) * | 2012-12-31 | 2013-04-17 | 泰山医学院 | Preparation method of bacillus subtilis powder and application thereof |
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2013
- 2013-11-18 CN CN201310577204.7A patent/CN103555640B/en active Active
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103039698A (en) * | 2012-12-31 | 2013-04-17 | 泰山医学院 | Preparation method of bacillus subtilis powder and application thereof |
Non-Patent Citations (2)
| Title |
|---|
| Isolation and characterization of a novel analyte from Bacillus subtilis SC-8 antagonistic to bacillus cereus;Nam Keun Lee,et al;《Journal of Bioscience and Bioengineering》;20101231(第3期);298-303 * |
| 高产中性蛋白酶菌株选育及产酶特性研究;陈鸿图;《中国优秀硕士学位论文全文数据库》;20130115(第2013/01期);B018-94 * |
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