Lactobacillus plantarum GBW-LP001 capable of highly producing lactic acid and antibacterial agent alternative thereof and application

文档序号：102248 发布日期：2021-10-15 浏览：27次中文

阅读说明：本技术 一株高产乳酸的植物乳杆菌gbw-lp001及其替抗菌剂和应用 (Lactobacillus plantarum GBW-LP001 capable of highly producing lactic acid and antibacterial agent alternative thereof and application ) 是由冯海霞吕伟王金龙赵斌师冬云于 2021-06-29 设计创作，主要内容包括：本发明提供了一株高产乳酸的植物乳杆菌GBW-LP001及其替抗菌剂和应用。所述植物乳杆菌GBW-LP001的分类命名为植物乳杆菌Lactobacillus plantarum,保藏编号为CGMCC No.22078,其16S rRNA的核苷酸序列如SEQ ID No.1所示,并具有培养24h后产乳酸量大于18.1g/L的作用,将其作为雏禽动物的开口料具有很好的使用效果。本发明还利用植物乳杆菌GBW-LP001与凝结芽孢杆菌E21、丁酸梭菌GBW-N1、葡萄糖氧化酶共同制备得到了一种新的替抗菌剂,能够用于禽类动物的养殖,其完全能够替代抗生素取得很好的养殖效果,增加养殖效益。(The invention provides lactobacillus plantarum GBW-LP001 capable of highly producing lactic acid, and a substituted antibacterial agent and application thereof. The classification of the Lactobacillus plantarum GBW-LP001 is named as Lactobacillus plantarum Lactobacillus plantarum The preservation number is CGMCC No.22078, the nucleotide sequence of 16S rRNA is shown in SEQ ID No.1, and the lactic acid yield is more than 18 after 24h of culture.1g/L, and has good use effect when being used as an opening material for young poultry animals. The invention also utilizes the lactobacillus plantarum GBW-LP001, the bacillus coagulans E21, the clostridium butyricum GBW-N1 and the glucose oxidase to jointly prepare a novel antibacterial agent, can be used for breeding poultry animals, can completely replace antibiotics to obtain a good breeding effect, and increases the breeding benefit.)

1. A lactobacillus plantarum GBW-LP001 capable of highly producing lactic acid is characterized in that the lactobacillus plantarum GBW-LP001 is named as lactobacillus plantarum in a classification modeLactobacillus plantarumThe microbial inoculum is preserved in the China general microbiological culture Collection center with the preservation number of CGMCC No. 22078.

2. Lactobacillus plantarum GBW-LP001 according to claim 1, characterized in that the nucleotide sequence of the 16S rRNA of Lactobacillus plantarum GBW-LP001 is shown in SEQ ID No. 1.

3. Lactobacillus plantarum GBW-LP001 according to claim 1, wherein the colonies of Lactobacillus plantarum GBW-LP001 are very regular circles, milky white, 2-3mm in diameter, smooth in surface, regular in edges, convex in the middle, opaque; the thallus is in a short rod shape, two ends are blunt and round, and the thallus is single, paired or chain-shaped, and has no flagellum and no spore.

4. Lactobacillus plantarum GBW-LP001 according to claim 1, wherein the lactic acid production of Lactobacillus plantarum GBW-LP001 after 24h cultivation is 18.1g/L or more.

5. An antibacterial agent comprising lactobacillus plantarum GBW-LP001 according to any one of claims 1 to 4, further comprising bacillus coagulans, clostridium butyricum and glucose oxidase.

6. The alternative antibacterial agent according to claim 5, wherein the Bacillus coagulans is Bacillus coagulans E21 with the preservation number of CGMCC No.13044, and the Clostridium butyricum is Clostridium butyricum GBW-N1 with the preservation number of CGMCC No. 14499.

7. The alternative antibacterial agent according to claim 6, wherein the viable bacteria content of Lactobacillus plantarum GBW-LP001 is 50 hundred million CFU/g, the viable bacteria content of Bacillus coagulans E21 is 50 hundred million CFU/g, the viable bacteria content of Clostridium butyricum GBW-N1 is 10 hundred million CFU/g, and the enzyme activity of glucose oxidase is 1000U/g.

8. Use of lactobacillus plantarum GBW-LP001 according to any one of claims 1-4 or the alternative antibacterial agent according to any one of claims 5-7 for the preparation of a mouth-opening or spray for hatchling.

9. The use according to claim 8, wherein the powder of Lactobacillus plantarum GBW-LP001 is used in an amount: adding 50g of lactobacillus plantarum GBW-LP001 bacterial powder into 10000 young birds.

10. Use of lactobacillus plantarum GBW-LP001 according to any one of claims 1 to 4 or a antibacterial agent according to any one of claims 5 to 7 for the preparation of a feed additive for improving the growth performance of poultry.

Technical Field

The invention belongs to the technical field of microorganisms, and relates to lactobacillus plantarum GBW-LP001 capable of highly producing lactic acid, a substituted antibacterial agent and application thereof.

Background

The production of commercial feed containing the growth-promoting drug feed additive is gradually stopped (short for 'forbidden resistance'). Probiotics, especially lactic acid bacteria, have become new anti-bacterial products and gradually highlight their unique advantages.

Lactobacillus plantarum is one of the lactobacilli, and is widely found in intestinal tracts of humans and animals, silage, fermented foods and dairy products. Usually, lactobacillus plantarum is found only in the late fermentation period, mainly because lactobacillus plantarum has high acid resistance and can endure various adverse environments. At present, lactobacillus plantarum is used as a probiotic for human, is used for reducing cholesterol and blood fat and losing weight, but is rarely used for breeding livestock and poultry animals.

At present, the lactobacillus plantarum is used for drinking water feeding in the breeding field of meat poultry. However, in the process of transporting the chicks from the poultry farm to the farm, a blank period of probiotics exists, and the blank period is the beginning and the key period of establishing intestinal flora from sterility to the intestinal flora of the chicks, so that the problem of how to increase the use of the probiotics after the chicks are shelved is very valuable to research, and most of microorganisms contacted with the chicks after the chicks are shelved are probiotics. Moreover, the stability of the lactobacillus plantarum is always a difficult problem, and the lactobacillus plantarum is short in storage time at normal temperature. Therefore, the stability of the lactobacillus plantarum is the first to solve, and the application range of the strain in the animal husbandry industry is expanded secondarily.

Disclosure of Invention

In order to overcome the defects and defects in the prior art, the invention provides a lactobacillus plantarum GBW-LP001 capable of highly producing lactic acid, the lactobacillus plantarum GBW-LP001 has excellent stability and stress resistance, a substitute antibacterial agent is prepared by using the lactobacillus plantarum GBW-LP001 and other strains, and the agent can be used for breeding livestock and poultry with remarkable effect.

In order to achieve the purpose of the invention, the invention is realized by the following technical scheme:

the invention provides Lactobacillus plantarum GBW-LP001 capable of highly producing lactic acid, which is classified and named as Lactobacillus plantarum and is preserved in China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No. 22078.

Further, the nucleotide sequence of 16S rRNA of Lactobacillus plantarum GBW-LP001 is shown as SEQ ID No. 1.

Further, the nucleotide sequence of pheS of lactobacillus plantarum GBW-LP001 is shown as SEQ ID No. 2; the nucleotide sequence of pyrG is shown as SEQ ID No. 3; the nucleotide sequence of uvrC is shown in SEQ ID No. 4; the nucleotide sequence of rpoA is shown in SEQ ID No. 5.

Furthermore, the bacterial colony of the lactobacillus plantarum GBW-LP001 is in a very regular circle shape, is milky white, has the diameter of 2-3mm, has a smooth surface and neat edges, and is convex and opaque in the middle; the thallus is in a short rod shape, two ends are blunt and round, and the thallus is single, paired or chain-shaped, and has no flagellum and no spore.

Furthermore, the lactic acid yield of the lactobacillus plantarum GBW-LP001 after 24 hours of culture is more than or equal to 18.1 g/L.

The invention also provides an antibacterial agent containing the lactobacillus plantarum GBW-LP001, and the antibacterial agent also comprises bacillus coagulans, clostridium butyricum and glucose oxidase.

Further, the bacillus coagulans adopts bacillus coagulans E21 with the preservation number of CGMCC No.13044, and the clostridium butyricum adopts clostridium butyricum GBW-N1 with the preservation number of CGMCC No. 14499.

Furthermore, the viable bacteria content of the lactobacillus plantarum GBW-LP001 is 50 hundred million CFU/g, the viable bacteria content of the bacillus coagulans E21 is 50 hundred million CFU/g, the viable bacteria content of the clostridium butyricum GBW-N1 is 10 hundred million CFU/g, and the enzyme activity of the glucose oxidase is 1000U/g.

The invention also provides application of the lactobacillus plantarum GBW-LP001 or the antibacterial agent in preparation of an opening material or a spray for breeding poultries.

Further, when in use, the lactobacillus plantarum GBW-LP001 bacterial powder is used in the following amount: adding 50g of lactobacillus plantarum GBW-LP001 bacterial powder into 10000 young birds.

Further, the chicks are chicks.

The invention also provides the application of the lactobacillus plantarum GBW-LP001 or the antibacterial agent in preparing feed additives or drinking water additives for improving the growth performance of poultry.

Furthermore, when the antibacterial agent is used as a feed additive, the dosage of the antibacterial agent is 500g per ton of feed per day according to the feed intake of the poultry all day.

Further, when the antibacterial agent is used as a drinking water additive, the dosage of the antibacterial agent is 250g/m³And (4) adding water.

Compared with the prior art, the invention has the following advantages and technical effects:

1. the lactobacillus plantarum GBW-LP001 with high lactic acid yield is extracted from intestinal sections of healthy white feather broilers, has good stability and stress resistance, is higher than those of similar products in the market, has acid resistance, bile salt resistance and acid yield obviously better than those of similar products in the market, is safe and reliable, can be used as an opening material for chicks, and can obviously improve the survival rate and the marketing weight of the broilers through experimental verification, thereby improving the breeding benefit of the broilers.

2. The invention utilizes lactobacillus plantarum GBW-LP001, bacillus coagulans E21, clostridium butyricum GBW-N1 and glucose oxidase to prepare a novel alternative antibacterial agent, and experiments prove that the alternative antibacterial agent can completely replace antibiotics to be used in daily breeding of broilers and meat ducks, so that the growth performance of the broilers and the meat ducks is improved, the survival rate is improved, the feed-meat ratio is reduced, the slaughter weight is increased, and the breeding benefit is improved.

Drawings

FIG. 1 is the colony morphology of Lactobacillus plantarum GBW-LP001 on MRS plates.

FIG. 2 shows morphological characteristics of Lactobacillus plantarum GBW-LP001 under a microscope.

FIG. 3 shows the use of Lactobacillus plantarum GBW-LP001 as starter feed in a breeder's chicken house using the breeding data.

FIG. 4 is field 'substitution antibiotic' test data of the substitution antibiotic agent in the later stage of broiler chicken breeding; wherein A is the stool form of the test group, B is the stool form of the control group 1, C is the stool form of the control group 2, and D is the daily gain data of the broiler chickens.

Detailed Description

The technical solution of the present invention will be further described in detail with reference to the following specific examples.

In the following examples, unless otherwise specified, the experimental methods used were all conventional methods, and materials, reagents and the like used were all available from biological or chemical reagents companies.

The formulations of the media required in the following examples are as follows:

1. MRS solid medium: 10.0g/L of peptone, 5.0g/L of beef extract powder, 4.0g/L of yeast extract powder, 20.0g/L of glucose, 2.0g/L of dipotassium phosphate, 2.0g/L of triammonium citrate, 5.0g/L of sodium acetate, 0.2g/L of magnesium sulfate, 0.05g/L of manganese sulfate, 15.0g/L of agar, 801.0 g/L of tween, and the pH value of 6.5 +/-0.2 (25 ℃).

2. MRS liquid medium: 10.0g/L of peptone, 5.0g/L of beef extract powder, 4.0g/L of yeast extract powder, 20.0g/L of glucose, 2.0g/L of dipotassium phosphate, 2.0g/L of triammonium citrate, 5.0g/L of sodium acetate, 0.2g/L of magnesium sulfate, 0.05g/L of manganese sulfate, 801.0 g/L of tween, pH value of 6.5 +/-0.2 (25 ℃), and 0.5g/L of L-cysteine hydrochloride.

The above culture medium is sterilized at 116 deg.C for 30min before use, and then stored at room temperature.

Example 1: screening, separation and molecular identification of lactobacillus plantarum GBW-LP001

1. Screening and purifying of lactobacillus plantarum GBW-LP001 strain

Sampling different intestinal sections of healthy white feather broilers, separating and purifying the samples on an MRS liquid culture medium and an MRS solid culture medium for multiple times to obtain a lactobacillus strain with good acid productivity, and naming the single colony as GBW-LP 001.

As shown in figure 1, the bacterial colony of the bacterial strain GBW-LP001 on an MRS plate is in a very regular circle shape, is milky white, has the diameter of 2-3mm, has a smooth surface and neat edges, and is convex in the middle and opaque; the thallus under the microscope is in a short rod shape, two ends are blunt and round, and the thallus is single, paired or chain-shaped, and has no flagellum and no spore.

2. Identification of Lactobacillus plantarum GBW-LP001

(1)16S rRNA gene identification: the DNA of the strain GBW-LP001 is used as a template, 16S rRNA universal primers 27F and 1492R are used for amplification, amplified fragments are sent to Shanghai workers for sequence determination, the obtained nucleotide sequence is shown as SEQ ID No.1, the obtained sequence is compared by BLAST, and the result shows that the homology of the GBW-LP001 and a Lactobacillus plantarum sequence in GenBank is 99.93 percent, so that the strain is preliminarily determined to be Lactobacillus plantarum.

(2) Other conserved genes were further identified: the method comprises the steps of designing proper primers (the primer sequences are shown in table 1) by utilizing 4 conserved genes of pheS gene (phenylalanyl-tRNA synthetase alpha subunit), pyrG gene (CTP synthetase), uvrC gene (excision endonuclease subunit) and rpoA gene (RNA polymerase alpha subunit), carrying out PCR amplification, sending amplified fragments to Shanghai to carry out sequence determination, and respectively obtaining nucleotide sequences shown as SEQ ID No.2-SEQ ID No.5, wherein the sequence shows that the homology comparison similarities of the amplified fragments with Lactobacillus plantarum sequences in GenBank are respectively 99.86%, 99.41%, 100% and 99.88% through BLAST comparison. Therefore, it was finally confirmed that the strain was Lactobacillus plantarum.

Table 1: conserved gene name and primer

(3) Strain preservation

And (3) performing strain preservation on the screened strain GBW-LP001, wherein the preservation unit of the lactobacillus plantarum GBW-LP001 is as follows: china general microbiological culture Collection center (CGMCC); address: western road No.1, north west city of township, beijing, institute of microbiology, china academy of sciences; the preservation date is as follows: 26/03/2021; the preservation number of the Lactobacillus plantarum is CGMCC No. 22078.

Example 2: stability and stress resistance analysis of Lactobacillus plantarum GBW-LP001

1. Stability analysis of Lactobacillus plantarum GBW-LP001

Fermenting 1000 hundred million CFU/g pure bacterial powder of lactobacillus plantarum GBW-LP001 in a fermentation tank at 37 deg.C for about 12h, wherein the carrier is skimmed milk powder, and lyophilizing. The same line 1 in the market is lactobacillus plantarum produced by a certain Shanghai company, and the bacterial count is 2000 hundred million CFU/g; the market peer 2 is Lactobacillus plantarum produced by a certain company in Jiangsu, and the bacterial count is 1000 hundred million CFU/g; the market peer 3 is Lactobacillus plantarum produced by Taiwan, the bacterial load is 1000 hundred million CFU/g; three market samples were available commercially.

Sealing different lactobacillus plantarum pure bacteria powder, and placing the sealed lactobacillus plantarum pure bacteria powder at the temperature of 20+1 ℃ to track the number of viable bacteria. The results are shown in table 2, the bacterial load of lactobacillus plantarum GBW-LP001 after 1 month of storage is 83.9% of the original bacterial load, and the bacterial load after 2 months is 48.6% of the original bacterial load, although the number of viable bacteria is reduced during storage, the bacterial loads are all higher than those of market isors 1, 2 and 3, which indicates that the stability of lactobacillus plantarum GBW-LP001 after 1 month and 2 months of storage is better than that of the existing lactobacillus plantarum products.

Table 2: stability of Lactobacillus plantarum GBW-LP001

2. Acid resistance of Lactobacillus plantarum GBW-LP001

Firstly, preparing artificial gastric juice with pH value of 3.0, containing 1% pepsin for standby. Weighing different lactobacillus plantarum powder, adding the powder into artificial gastric juice according to the mass-volume ratio of 1: 100, and oscillating the powder at the constant temperature of 37 ℃ and 200rpm for 1 h; meanwhile, blank control without artificial gastric juice is made, and the bacteria content is respectively determined. The bacteria retention rate is the bacterial quantity after the artificial gastric juice treatment/blank bacterial quantity multiplied by 100 percent. As shown in table 3, the bacteria retention rates of GBW-LP001 in artificial gastric juice at pH 3.0 for 1h and 2h were 96.6% and 72.4%, respectively, which are higher than 3 market peers, indicating that lactobacillus plantarum GBW-LP001 has better acid resistance than the existing lactobacillus plantarum products.

Table 3: stress resistance of Lactobacillus plantarum GBW-LP001 under acidic conditions

3. Bile salt resistance of Lactobacillus plantarum GBW-LP001

First, PBS buffer solution with pH of 6.8 is prepared and sterilized for 30min at 121 ℃ for standby. Then, 0.3% of pig bile salt was added to the PBS buffer to prepare a PBS buffer containing 0.3% of bile salt. Weighing different lactobacillus plantarum powder, adding the powder into PBS buffer solution containing 0.3% bile salt according to the mass-volume ratio of 1: 100, and oscillating at the constant temperature of 37 ℃ and 200rpm for 4 hours; meanwhile, the blank control without the bile salt solution is used for measuring the bacteria content. And (5) calculating the survival rate of the bacteria by taking the number of bacteria measured in the blank as a control.

The results are shown in Table 4, the bacterial retention rate of GBW-LP001 in 0.3% bile salt after being treated for 4h at 37 ℃ is 28.9%, which is obviously higher than that of 3 market lines, and the results show that the Lactobacillus plantarum GBW-LP001 has certain choline resistance and the effect is superior to that of the existing Lactobacillus plantarum product.

Table 4: stress resistance of Lactobacillus plantarum GBW-LP001 under bile salt condition

4. Acid productivity of Lactobacillus plantarum GBW-LP001

Respectively selecting single colonies of different lactobacillus plantarum, inoculating the single colonies into an MRS liquid culture medium, culturing at 37 ℃ for 24 hours, taking a proper amount of culture solution to detect the pH value, and detecting the lactic acid yield by using a lactic acid test kit (Nanjing constructed biology). As shown in Table 5, the pH of the bacterial liquid of Lactobacillus plantarum GBW-LP001 was 3.74, the lactic acid production amount was 18.1g/L, the pH was lower than 3 market peers, and the lactic acid production amounts were all higher than 3 market peers, indicating that Lactobacillus plantarum GBW-LP001 had more excellent acid production performance.

Table 5: acid production performance of Lactobacillus plantarum GBW-LP001

Example 3: application case of lactobacillus plantarum GBW-LP001 in culture site

Lactobacillus plantarum GBW-LP001 used as chick starter feed in chicken house

Test site: fuxi (Weihai) farming-grazing development Co Ltd

Test protocol:

1. test materials: lactobacillus plantarum GBW-LP001 with the strain content of 2000 hundred million CFU/g; edible carmine pigment: procurement by Tianjin food company; PBS buffer: the Qingdao root research and development center provides; spraying equipment: is provided by a Fox breeding poultry factory.

2. Application protocol

2.1 preparation of probiotic spray solution

Taking out Lactobacillus plantarum GBW-LP001 bacterial powder from a refrigerator, adding 50g of bacterial powder into 10000 chicks according to the using amount of the bacterial powder, dissolving the bacterial powder into 2100mL of PBS buffer solution, adding 1g of carmine, and uniformly mixing. Adding the prepared probiotic spraying solution into spraying equipment.

2.2, grouping

300 ten thousand chicks in the test group use the lactobacillus plantarum GBW-LP001 bacterial powder; the control group 300 ten thousand chicks were not used.

2.3 spraying

And adjusting a spraying device, spraying the chicks for three times, stopping for 5s, and simultaneously controlling the speed of the chick conveying belt to enable the chicks in each paper box to stay for 5s under the spraying device.

3. Effects of use

After spraying, the probiotic spraying solution forms fog in the spraying equipment and can enter the bodies of the chickens through a respiratory system, meanwhile, red small water drops are formed on the feathers of the chickens, the chickens can peck the water drops mutually, and the probiotics further enter the bodies of the chickens.

The farming and pasturing company adopts Cobb ground flat breeding, and after the chicks are hatched and sprayed, the comprehensive data of 600 ten thousand broilers which are out of the farm are counted after two months, wherein the weight of the farm is the average weight of each chicken, the input-output ratio is the ratio of input/output, the input means the cost of each chicken for increasing probiotics, and the output means the yield increase rate of each chicken after the same number of chicks are out of the farm.

The production performance data of broiler breeding after chicks are bred is shown in fig. 3, and it can be seen that the broiler survival rate of a test group is improved by 0.91% compared with that of a control group, the slaughter weight is increased by 79g, the feed-weight ratio is reduced by 0.02%, the input-output ratio is 1: 33, which shows that after lactobacillus plantarum GBW-LP001 is used, the growth performance of the broiler is obviously improved, and the broiler breeding effect is very considerable because the broiler breeding performance data is safe and reliable for the chicks.

Second, the use of 'replacing antibiotics' in the later stage of the cultivation of the meat poultry

A novel antibacterial agent is prepared by utilizing lactobacillus plantarum GBW-LP001, bacillus coagulans E21, clostridium butyricum GBW-N1 and glucose oxidase, wherein the viable bacteria content of the lactobacillus plantarum GBW-LP001 is 50 hundred million CFU/g, the viable bacteria content of the bacillus coagulans E21 is 50 hundred million CFU/g (the bacillus coagulans preservation number is CGMCC No.13044), the viable bacteria content of the clostridium butyricum GBW-N1 is 10 hundred million CFU/g (the clostridium butyricum preservation number is CGMCC No.14499), and the enzyme activity of the glucose oxidase is 1000U/g.

The collection unit of the bacillus coagulans E21: china general microbiological culture Collection center (CGMCC); address: western road No.1, north west city of township, beijing, institute of microbiology, china academy of sciences; the preservation date is as follows: 2016, 9, 26 months; the preservation number of the Bacillus coagulans is CGMCC NO. 13044.

The collection of Clostridium butyricum GBW-N1: china general microbiological culture Collection center (CGMCC); address: western road No.1, north west city of township, beijing, institute of microbiology, china academy of sciences; the preservation date is as follows: year 2017, month 08, day 07; the preservation number of the Clostridium butyricum is CGMCC No. 14499.

Substitution resistance test 1: feed additive for later stage of broiler chicken breeding

The experimental design is shown in table 6, the control group 1 (antibiotic group) starts to add 20% colistin sulfate according to the conventional addition amount at the age of 30 days, the control group 2 starts to add the compound traditional Chinese medicine preparation according to the conventional addition amount at the age of 30 days, and the experimental group starts to add the antibacterial agent at the age of 30 days according to the feed intake of 500 g/ton per day.

Table 6: on-site 'substitution resistance' test design in later stage of broiler chicken breeding

The test results are shown in fig. 4: compared with the control group 1, the test group has better forming degree and no water excrement phenomenon in the excrement form of the broiler chicken; control 2 was dark brown in color relative to the feces of the other two groups, with a small amount of visible feed particles. In the aspect of daily gain, the daily gain of the broiler chickens in the test group is obviously higher than that of the broiler chickens in the control group, which shows that the antibacterial agent can completely replace the antibiotic product, and the using effect is obviously better than that of the antibiotic and the compound traditional Chinese medicine preparation.

Substitution resistance test 2: water drinking additive for later stage of meat duck breeding

The experimental design is shown in Table 7, and the control group was started to add 20% at 28 days of ageColistin sulfate, the test group started to add the alternative antibacterial agent to drinking water at the age of 28 days, and the addition amount of the alternative antibacterial agent is 250g/m³And (3) water.

Table 7: on-site 'resistance replacing' test design for meat duck breeding later stage

The results of the meat duck breeding test are shown in table 8: compared with a control group, the survival rate of the meat duck in the test group is improved by 2.59 percent, the feed-meat ratio is reduced by 0.025 percent, and the slaughter weight is increased by 0.14 jin, which shows that the alternative antibacterial agent taking the lactobacillus plantarum GBW-LP001 as the main component can completely replace the use of antibiotics, and meanwhile, positive results are obtained in the production performance of the meat duck.

Table 8: on-site 'substitute resistance' test breeding data in later breeding period of meat ducks

The total data of the survival rate, the feed-meat ratio and the slaughter weight are calculated according to a weighted average method.

The above examples are only intended to illustrate the technical solution of the present invention, but not to limit it; although the present invention has been described in detail with reference to the foregoing embodiments, it will be apparent to those skilled in the art that various changes may be made and equivalents may be substituted for elements thereof; and such modifications or substitutions do not depart from the spirit and scope of the corresponding technical solutions.

Sequence listing

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tggacaagca tattacgatg gctgatttga agggcacctt aattctggtt gccaagactt 480

tgtttggcga tcaattcgat gttcggctac ggccaagctt ctttccattc acggaaccat 540

ccgtagaagc tgatgtaact tgctttaatt gcaatggcaa gggctgtgca atctgtaagc 600

aaacgggttg gatcgaagta ctgggtgccg gcatggttca cccccacgtg ttagaaatgt 660

ctggcattga tccagaagaa tatggtggtt ttgcttt 697

<210> 3

<211> 673

<212> DNA

<213> Lactobacillus plantarum (Lactobacillus plantarum)

<400> 3

gagttgattt taagttccga caatgttttc tatatccata ccacattgat cccatattta 60

cgggcagctg gtgaaatgaa gacgaagcca acgcaacatt ctgttaagga attgcggagt 120

tatgggattc agccgaacat gttagttgtc cggactgaac aaccaattac gcgggaaatg 180

cggaataaga ttgcgtcctt ctgtgacgtg gaaccagaag cagtcattga atccttagac 240

gttaagacga tttattcaat tccgttgaat gttcagaaac aaaacatgga ccaaatcgtc 300

cttgaccatt ttgatgtaca ggcacctaag gccgacatga gtgaatggat tgacttagaa 360

catcatgttc agaacttatc acggaccatc aagattgcgc tagtcggaaa atacgtcgct 420

ttacaggatg cttatatctc agtgacggaa gcattgaagc atgctggcta tacggatgat 480

gccgacattg atttgaagaa gatttctgct gaagatgtta cgccagaaaa tgtcgaagaa 540

ctactcggcg atgctgacgg aatcttagtt cctggtggct ttggtgatcg gggaattgaa 600

ggtaagatta cggcaatcaa gtatgcccgt gaaaacgacg tgccattctt aggtatctgc 660

ttggggaatg caa 673

<210> 4

<211> 770

<212> DNA

<213> Lactobacillus plantarum (Lactobacillus plantarum)

<400> 4

gatccatttt ttaatgggta aggccaagaa cttgaaaaat cgggtgcgtt catatttcaa 60

aagcagccat actggtaaga ccgcgcggtt agtatcagag attgcggact ttgagtttat 120

tgtgacctca acggataagg aagccttcct gctagaaatt acgttaattc aaaaacatca 180

accgtacttt aatattaaat taaaaaaggg aaccggttat ccatacatta agattaccaa 240

tgaacgagat ccccagatcc tgatcgtcag tgatgttcgt aaagacggtg gctactactt 300

tgggccatat cccaacgttt acgcggctca ggaaacggtc aattttattc agaaggtcta 360

tccgttgcgg cgttgccatg gtttccagaa gcggccgtgc ctctattatc atatgggcca 420

atgcttggga gcttgcttta aaacggtgcc ggtggcagaa tatgacgctc agatcaagcg 480

gattaaatcc tttttgaatg gtcacgttga aacggtcaag aagcagttaa ctaaacgaat 540

ggatcaagcg gccgctgacc tagaatttga acgagccgct gaattacgtg atcaactgaa 600

ttatattgaa atgacggttg aaaagcagaa aattatttca aacgataata cgccccgcga 660

cctgtttaat ttctatcttg ataaaggctg gctttcgatt caggtcttct ttattcggca 720

agcgcggttg atgaagcgcg agaaacgctt gttccgatta aatggttcaa 770

<210> 5

<211> 819

<212> DNA

<213> Lactobacillus plantarum (Lactobacillus plantarum)

<400> 5

ttatgattga aatttgaaaa acctaacatt cataaaattg atgaaaacga caactatggt 60

aagtttgttg tagaaccgct tgaacgcggt tatggtacaa ctttagggaa ttcacttcgt 120

cggattcttc tttcttcttt acctggcgct gctgttacta gtattcaaat tgatggtgtt 180

cttcatgaat tttcaacgat tgagggcgta acggaagacg ttacagcaat tatcttgaat 240

gttaagaaga ttgcacttaa gttggaatca gacgaaacca agacgttgga aatcgacgtt 300

aagggtcctg ctaacgttac tgccagtgat atcattggcg atgcggacgt agaagtcttg 360

aatccagact taccaatttg taccgtagca gacggggcac acttccatat gcgtatgacc 420

gcaaatactg gtcgtggtta tgtttccgct gaggataaca aacatcgtga agatgacatg 480

ccaattggcg ttttagctgt tgattcattg tattctccaa tcgaacgtgt caactaccaa 540

gttgaaaaca cgcgggttgg tcaacgtgat gatttcgata agttaacctt agacgtttgg 600

acaaatggtt caatcactcc aagtgaagcc attagtctat cagccaaaat cctgactgat 660

cacctttcaa tcttcgtaaa tctcactgat gaagctaaaa acactgatgt gatggtcgag 720

aaggaagaaa cgcataagga aaagatgtta gaaatgacga ttgaagagtt agatctctcc 780

gtccgttcat acaattgctt gaagcgtgca ggtactcac 819

<210> 6

<211> 18

<212> DNA