CN102936285A - Antibody for preventing and treating fish Edwardsiellaictaluri disease, preparation method and applications thereof - Google Patents
Antibody for preventing and treating fish Edwardsiellaictaluri disease, preparation method and applications thereof Download PDFInfo
- Publication number
- CN102936285A CN102936285A CN2012102692493A CN201210269249A CN102936285A CN 102936285 A CN102936285 A CN 102936285A CN 2012102692493 A CN2012102692493 A CN 2012102692493A CN 201210269249 A CN201210269249 A CN 201210269249A CN 102936285 A CN102936285 A CN 102936285A
- Authority
- CN
- China
- Prior art keywords
- antibody
- channel
- catfish
- disease
- preparation
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Abstract
The present invention discloses an antibody for preventing and treating a fish Edwardsiellaictaluri disease, a preparation method and applications thereof. The antibody preparation method comprises: inactivating Edwardsiellaictaluri having a preservation number of CCTCCNO:M2012024, and carrying out inoculation on egg laying avian in batches to obtain immunized eggs containing anti-Edwardsiellaictaluri antibody, wherein the antibody can be applicable for drugs for fish Edwardsiellaictaluri disease prevention and treatment. The antibody has characteristics of high specificity, no toxicity, no residue, no resistance, no biological safety hazard, and simple and rapid preparation method, and can be prepared into powder, wherein the powder can be adopted as a feed additive or a pre-material to be directly fed or sprinkled in the whole pool, such that the disadvantage that other antibody drugs can provide effects only by injection or soaking is overcome, use is convenient, and labor intensity is substantially reduced.
Description
Technical field
The present invention relates to a kind of antibody and its preparation method and application, be specifically related to a kind of antibody for prevention and treatment Yu Lei Channel-catfish tarda disease and its preparation method and application.
Background technology
The alepidotes such as Yellow catfish, Channel-catfish fish, catfish are the famous and precious freshwater fishes of China's high-quality, and these fish are not only stung between Fresh & Tender in Texture, few flesh, and are of high nutritive value, and deeply are subject to liking of consumers in general.Yet in recent years, a kind of prevailing disease is broken out in the culture zone of the Yellow catfish of various places, U.S. Cha Wei Channel-catfish, catfish, its cardinal symptom is: fin ray base portion, oral cavity, lower jaw, the gill cover, eye socket hyperemia, position, center, the crown is subcutaneous rubescent, when serious, skull splits, overhead a long and narrow hemorrhage band that festers appears in section, presents typical " red head disease of Pseudobagrus " symptom.This kind of disease has that infectivity is strong, the mortality ratio high, become the especially bottleneck problem of the sound development of Yellow catfish aquaculture of restriction alepidote.The countermeasure that at present take each culture zone is mainly to use broad-spectrum antibiotics, yet this kind of method not only can increase treatment cost, but also can bring the problems such as exceeding standard of residual antibiotic in the generation of resistance and fish body.Therefore, find a species specificity height again the anti-red head disease of Pseudobagrus Substitutes For Antibiotic of noresidue become the problem that the Yellow catfish aquaculture is needed solution badly.
From the immunology angle, prepare the antibody of specific anti-microbial pathogen, be the sustainable development of preventing and curing diseases.At present, the patent documentation that publication number is CN101829321A discloses the method for preparing anti-red head disease of Pseudobagrus fulvidraco vaccine: obtain safe and reliable vaccine with formalin inactivated vaccine, by the mode administration of soaking and injecting, immunoprotection reaches 70%-95%.Yet this vaccine is used certain limitation in large-scale cultivation: 1. labour intensity is large; 2. exempted from fish easily injured, caused secondary disease; 3. prevention work will be carried out in advance, for sick fish without therapeutic action.
Summary of the invention
For solving the deficiencies in the prior art, the object of the present invention is to provide a kind of easy to use, can not damage fish, not only can prevent but also can treat antibody of Yu Lei Channel-catfish tarda disease and preparation method thereof.
In order to realize above-mentioned target, the present invention adopts following technical scheme:
The preparation method of the antibody of prevention and treatment Yu Lei Channel-catfish tarda disease, it is characterized in that, with through inactivator deactivation De Channel-catfish tarda BRK-ADHSJ(Edwardsiella ictaluri BRK-ADHSJ) gradation inoculation egg fowl, the adaptive immune egg, the antibody that contains anti-tarda in aforementioned immune egg.
The preparation method of the antibody of aforesaid prevention and treatment Yu Lei Channel-catfish tarda disease is characterized in that Qian Shu Channel-catfish tarda BRK-ADHSJ is the bacterial strain that deposit number is CCTCC NO:M2012024.
The preparation method of the antibody of aforesaid prevention and treatment Yu Lei Channel-catfish tarda disease is characterized in that aforementioned inactivator is formaldehyde.
The antibody of prevention and treatment Yu Lei Channel-catfish tarda disease, it is characterized in that, utilize following method to make: use through inactivator deactivation De Channel-catfish tarda BRK-ADHSJ gradation inoculation egg fowl, the adaptive immune egg, contain the antibody of Kang Channel-catfish tarda BRK-ADHSJ in aforementioned immune egg.
The antibody of aforesaid prevention and treatment Yu Lei Channel-catfish tarda disease is characterized in that Qian Shu Channel-catfish tarda BRK-ADHSJ is the bacterial strain that deposit number is CCTCC NO:M2012024.
Aforesaid antibody is applied in the medicine of preparation prevention and treatment Yu Lei Channel-catfish tarda disease.
Aforesaid application, is characterized in that, by fresh immune egg spraying drying after the micro-coating of trehalose, makes the pulvis that contains aforementioned antibody, and aforementioned pulvis is is directly thrown something and fed as fodder additives or pre-batching.
Aforesaid application is characterized in that water is cooked auxiliary material, and aforementioned antibody is made to aqua or special-purpose dipping bath agent use.
Usefulness of the present invention is: the antibodies specific of prevention and treatment Yu Lei Channel-catfish tarda disease is high, nontoxic, noresidue, have no drug resistance, the lifeless matter potential safety hazard; The method industrialization degree for preparing this antibody is high, and efficiency is high; This antibody is applied in the medicine of preparation prevention and treatment Yu Lei Channel-catfish tarda disease, different from the use of Broad spectrum antibiotics, the medicine that use contains this antibody can not change the immunological status of disease fish, and after use antibody, the immunity system of sick fish still can be made effective reaction to other external sources infringements; This antibody not only can make aqua or special-purpose dipping bath agent is used, especially can make pulvis, as fodder additives or pre-batching, directly throw something and feed or full pool spilling head, overcome other antibody drugs and can only inject or soak effectively shortcoming, use very conveniently, greatly reduce labour intensity.
The accompanying drawing explanation
Fig. 1 is that the deposit number bacterial strain 16S rRNA gene order that is CCTCC NO:M2012024 is Yu the 16S rRNA gene order comparison chart of Channel-catfish tarda reference culture (AB453281.1, EU285522.1, EF015475.1).
Embodiment
Below in conjunction with specific embodiment, the present invention is done to concrete introduction.
The preparation method of the antibody of prevention and treatment Yu Lei Channel-catfish tarda disease, comprise the steps: 1. separation and purification pathogenic bacteria; 2. determine this separation and purification gained pathogenic bacteria Wei Channel-catfish tarda BRK-ADHSJ(Edwardsiella ictaluri BRK-ADHSJ); 3. obtain the immune egg of the antibody of Han Kang Channel-catfish tarda BRK-ADHSJ.
The first step: separation and purification Yellow catfish " red head disease of Pseudobagrus " pathogenic bacteria
1, material: testing the disease fish is the Yellow catfish of tool typical case " red head disease of Pseudobagrus " symptom, picks up from Huzhou, Zhejiang plant.
2, method: at first, get the sick fish of tool classical symptom alive, by stroke-physiological saline solution, rinse body surface 2 times, the cerebral tissue sampling with transfering loop in liver, kidney and the rubescent position of head.Then, on TSA, ordinary nutrient agar substratum, carry out plate streaking, after 28 ℃ of constant temperature culture 48h, the dominant colony that the picking morphological specificity is consistent carries out separation and Culture 3 times, is inoculated on the TSB liquid nutrient medium and continues enlarged culturing.Finally, bacterium liquid glycerol adding is in-80 ℃ of preservations, stand-by with bacterial strain as test.
Second step: determine the first step separation and purification gained pathogenic bacteria Wei Channel-catfish tarda BRK-ADHSJ
Embodiment mono-: Experimental infection
1, material: test is the healthy Yellow catfish fingerling of taking from Wujiang, Suzhou plant with healthy Yellow catfish, specification is 35 ± 5g, laboratory is transported in the plastics bag oxygenation back, in aquarium, support temporarily, the test water body is through continuous oxygenation circulating filtration, use automatic temperature control instrument Control experiment water temperature (28 ± 1) ℃, not bait throwing in during supporting temporarily.
2, method: Yellow catfish is supported and carries out Experimental infection after 5 days temporarily.Get the test bacterial strain of-80 ℃ of preservations, be seeded on the TSA flat board, cultivate 48 h for 28 ℃.The lower culture of stroke-physiological saline solution flushing doubling dilution become the bacteria suspension of different concns.The test fish after stablizing is supported in intramuscular injection temporarily, and every endnote is penetrated 0.2 mL, control group injection equivalent physiological saline, and every experimental concentration group arranges repeating groups.The test fish is positioned over the aquarium of 20 L, and every cylinder is put 10 tails in a suitable place to breed, and internal circulation filtering is processed ammonia nitrogen, nitrite and oxygenation, and temperature controller is temperature automatically controlled at 28 ℃, observes, records test fish death condition every day, takes out in time dead fish, Continuous Observation 14 days.The artificial challenge the results are shown in Table 1:
Test-results after the bacterial strain artificial challenge 120h that table 1 is separated to
| Bacterial concentration/CFUMl -1 | Attack toxic agent amount/mL | Test fish number/tail | Death toll/tail |
| 2.5×10 9 | 0.2 | 10 | 10 |
| 2.5×10 8 | 0.2 | 10 | 10 |
| 2.5×10 7 | 0.2 | 10 | 10 |
| 2.5×10 6 | 0.2 | 10 | 8 |
| 2.5×10 5 | 0.2 | 10 | 7 |
| Physiological saline | 0.2 | 10 | 1 |
| Blank | 0 | 10 | 0 |
Test-results shows, artificial challenge's method of intramuscular injection bacterial suspension can make healthy Yellow catfish morbidity, and artificial challenge's symptom is similar to " red head disease of Pseudobagrus " natural occurrence symptom, be that disease fin bar base portion, upper lower jaw, the gill cover, belly are hemorrhage, the skin of vertex color shoals, rubescent, anus and gonopore are congested, hemorrhage, evagination, and expand in abdominal cavity ascites etc.; When bacterial concentration reaches 2.0 * 10
7During CFU/ml, mortality ratio reaches 100%.
Embodiment mono-has confirmed: the pathogenic bacteria that the test obtained in the first step is Yellow catfish " red head disease of Pseudobagrus " with bacterial strain.
The fish body that " red head disease of Pseudobagrus " classical symptom infected group occurred to, method according to the first step narration is separated, the purifying pathogenic bacteria, the colony characteristics of the bacterial strain that the pathogenic bacteria colony characteristics obtained is separated to the first step is consistent, the bacterial strain that explanation is separated to from artificial challenge's sick fish is same bacterial strain with test with bacterial strain, is all pathogenic bacterias of Yellow catfish " red head disease of Pseudobagrus ".
Embodiment bis-: the thalli morphology microscopic examination of bacterial strain for test
1, material: the test bacterial strain that the first step separation and purification obtains.
2, method: get the test bacterial strain that the first step separation and purification obtains, streak inoculation, in ordinary nutrient agar, TSA flat board, is cultivated 48h for 28 ℃, the preparation bacterial suspension.Carrying out ne ar observes and gram staining liquid dyeing.Experimental result shows, this bacterial strain can form on ordinary nutrient agar, TSA plate culture medium that circle is smooth, protuberance, moistening and with small colonies glossy, the shape that is translucent.Gramstaining is negative, and rod-short is about 2 μ m, and diameter 0.5 μ m, without pod membrane, do not form gemma, between individuality, vertically connects in twos.
Embodiment tri-: test is identified with the Physiology and biochemistry of bacterial strain
1, material: the test bacterial strain that the first step separation and purification obtains.
2, mensuration project and result: referring to table 2
The biochemical characteristic of bacterial strain for table 2 test
| The mensuration project | Result | The mensuration project | Result |
| Oxidation/fermentation | F | H 2S | - |
| Ornithine decarboxylase ODC | - | Indoles produces IND | - |
| The bilingual enzyme ADH of arginine | - | N-acetyl-β glycoside β NAG | - |
| Lysine decarboxylase LDC | - | Β tilactase β GAL | - |
| Urase URE | - | D-Glucose GLU | - |
| L-arabitol LARL | - | D-sucrose SAC | - |
| Semi-lactosi hydrochlorate GAT | - | L-arabinose LARA | - |
| 5-ketone group-gluconic acid 5KG | - | D-arabitol DARL | - |
| Sodium.alpha.-ketopropionate RP | + | α tilactase α GAL | - |
| β glycoside β GLU | - | D-trehalose TRE | - |
| PEARLITOL 25C MAN | - | L-rhamnosyl RHA | - |
| D-Maltose MAL | - | Inositol INO | - |
| Pentitol ADO | - | D-cellobiose CEL | - |
| Ancient sugared PLE | - | D-glucitol SOR | - |
| β glucuroide β GUR | - | Maltoside enzyme α MAL | - |
The measurement result of table 2 shows: this test is the catalase positive by the principal character of bacterial strain, oxidase negative, fermented type.
Pathological change, morphological specificity, cultural characters and the physio-biochemical characteristics of bacterial strain for one, two, three shown tests in conjunction with the embodiments, can this separation and purification gained test of preliminary evaluation with bacterial strain Wei Channel-catfish tarda BRK-ADHSJ.
Embodiment tetra-: test is identified with the specific amplification of the 16S rRNA gene of bacterial strain
1, material: the test bacterial strain that the first step separation and purification obtains;
Primer, sequence is as follows:
A N1:5 ' ACAGTITGATCCTGGCTCAG 1 '
A N2:5 ' CGCTACCTrGTTACGACTT 1 '
Universal primer is given birth to work biotechnology company limited by Shanghai and is synthesized.
2, method: get the test that the first step separation and purification obtains and rule on the TSA flat board with bacterial strain, picking one ring culture after 28 ℃ of cultivation 48h, add 50 μ L sterilized waters, after pressure-vaccum mixes, centrifugal 1 min of 10000 r/min, remove supernatant, add again 50 μ L sterilized waters, after pressure-vaccum mixes, boiling water bath 10 min, centrifugal 5 min of 12000 r/min, get 1 μ L supernatant liquor for pcr amplification.
The amplified reaction volume is 60 μ L, adds respectively 10 * PCR damping fluid, 6 μ L, each 1 μ L of primer, and dNTP 2 μ L, Taq enzyme 0.5 μ L, sterilized water is supplied reaction volume.
The PCR reaction conditions is: 95 ℃ of denaturation 5 min, and 94 ℃ of sex change 1 min, 55 ℃ of annealing 45s, 72 ℃ are extended 45s, carry out altogether 30 circulations, and last 72 ℃ are extended 7 min end reactions.
After getting 6 μ L reaction solutions carry out electrophoresis detection on 1% sepharose, glue reclaims test kit and reclaims for product, carries out DNA sequencing.Examining order has been assisted by Shanghai Ying Jun Bioisystech Co., Ltd.
The test recorded is input in ncbi database and carries out the Blast comparison by bacterial strain 16S rRNA gene order, the homology analysis of its gene nucleotide series is found to the conservative property fragment of Yu Channel-catfish tarda reference culture (AB453281.1, EU285522.1, EF015475.1) has 100% homology, referring to Fig. 1.
Combining form is learned and physio-biochemical characteristics, can by separate of the test that obtains with the further Que of bacterial strain Ding Wei Channel-catfish tarda BRK-ADHSJ(Edwardsiella ictaluri BRK-ADHSJ).
Embodiment five: the drug sensitivity test of bacterial strain for test
1, material: the test bacterial strain that the first step separation and purification obtains.
2, method: bacterial strain enlarged culturing on the TSB liquid nutrient medium for the test that the first step separation and purification is obtained, after 28 ℃ of cultivation 48 h, liquid culture is coated on the TSA flat board, suitably after drying, paste drug sensitive test paper, measure antibacterial circle diameter after 28 ℃ of cultivation 24 h, press the susceptibility that the drug sensitive test paper product description is judged medicine.Drug sensitivity test the results are shown in Table 3
Table 3 medicament sensitivity test result
| The medicine name | Susceptibility | The medicine name | Susceptibility |
| Gentamicin Gentamycin | R | Rifampin Rifampicin | I |
| Ciprofloxacin Ciprofloxacin | S | Erythromycin E rythromycin | R |
| Trimethoprim-sulfamethoxazole Cotrimoxazole | S | Liu Suanyan NEOMYCIN SULPHATE Neomycin | S |
| Levofloxacin Ofloxacin | S | Streptomycin sulphate Streptomycin | R |
| Kantlex Kanamycin | R | Norxin Norfloxacin | I |
| Cephradine Ceftazidime | S | Lomefloxacin Ciprofloxzcin | S |
| Amoxycilline Trihydrate bp Amoxicillin | R | Cefoxitin Cefalothin | I |
| Penbritin Ampicillin | S | Pioneer IV Cefalexin | S |
| Lincomycin Lincomycin | R | Ceftriaxone Rocephin | S |
| Enoxacin Enoxacin | S | Tobramycin Tobramycin | R |
| Nifurazolidone Furazolidone | S | Doxycycline Doxycycline | S |
Annotate: the responsive sensitive of S; R resistance resistant; I medium sensitivity mid-sensitive.
Test-results shows, this bacterial strain is to Ciprofloxacin, trimethoprim-sulfamethoxazole, levofloxacin, Cephradine, penbritin, enoxacin, Nifurazolidone, Liu Suanyan NEOMYCIN SULPHATE, lomefloxacin, pioneer's IV, Ceftriaxone, doxycycline sensitivity; Insensitive to the gentamicin in trial drug, kantlex, amoxycilline Trihydrate bp, lincomycin, erythromycin, Streptomycin sulphate, tobramycin; To Rifampin, Norxin, cefoxitin medium sensitivity.The sensitive features of this strains is similar to Channel-catfish tarda reference culture.
The test-results of comprehensive above five embodiment, can determine the test bacterial strain Wei Channel-catfish tarda BRK-ADHSJ of separation and purification gained from " red head disease of Pseudobagrus " Yellow catfish of natural occurrence.Because of " red head disease of Pseudobagrus " Shi You Channel-catfish tarda, BRK-ADHSJ causes, so this disease is again be Channel-catfish tarda disease.
The applicant will the test of separation and purification gained submit the center preservation of Chinese Typical Representative culture collection to bacterium strain Channel-catfish tarda BRK-ADHSJ on February 19th, 2012 from " red head disease of Pseudobagrus " Yellow catfish of natural occurrence, and deposit number is CCTCC NO:M 2012024.
The 3rd step: the immune egg that obtains the antibody of Han Kang Channel-catfish tarda BRK-ADHSJ
At first, preparation deactivation De Channel-catfish tarda BRK-ADHSJ vaccine.Concrete operations are as follows:
Under aseptic condition, get under-80 ℃ of conditions and preserve De Channel-catfish tarda BRK-ADHSJ bacterial classification, in 10 mlTSB meat soups, postvaccinal meat soup is put into to water bath chader, 28 ℃ of shaking culture 24 h with a small amount of microbionation of transfering loop picking from strain tube.
Bacterium liquid 10 ml that recovery is cultivated are seeded in 500 mlTSB meat soups again, and postvaccinal meat soup is 28 ℃ of continuation shaking culture 48h in water bath chader.The TSB meat soup of secondary cultivation is uniformly coated on the TSA flat board prepared, coated TSA flat board is put into to 28 ℃ of constant incubators, cultivate 48h, until the bacterium lawn covers with whole flat board.Lawn with growing on sterilized physiological saline aseptic technique washing plate, be placed in sterilized infusion bottle by the bacteria suspension under washing.
To from flat board, be inoculated into the ratio of 200 mlTSB broth cultures in every 10 ml bacterium liquid by the bacterium liquid of wash-out, carry out the enlarged culturing of bacterium liquid.Shaking culture 48 h under 28 ℃ of conditions, thus the high density bacterial suspension obtained.The high density bacterial suspension that enlarged culturing is obtained, by volume ratio is added the formaldehyde of 5-6 ‰, puts into water bath chader, 28 ℃ of vibration 48 h deactivations.
By deactivation vaccine completely, aseptic technique is sub-packed in 500 ml sterilizing high temperature bottles, and carries out the paraffin sealing.Divide the inactivated vaccine installed to be placed in 4 ℃ of preservations, standby.
The inactivated vaccine of preparation is inoculated on sheep blood TSA agar plate, after 28 ℃ of constant temperature culture 24-48h, observes on flat board whether the cause of disease bacteria growing is arranged, to determine in vaccine whether contain not deactivation pathogenic bacteria completely.
Then, the immune egg of the antibody of preparation Han Kang Channel-catfish tarda BRK-ADHSJ.Concrete operations are as follows:
Selection has the healthy egg fowl that the no-special pathogen of laying eggs of high immunne response ability carries, and adopts the deactivation Hou De Channel-catfish tarda BRK-ADHSJ vaccine of above-mentioned preparation and suitable adjuvant inoculation egg fowl.Interval 7 d after inoculation for the first time, then inoculate for the second time with same dosage, method, for the second time after the inoculation again interval 60 d with same dose, inoculate the 3rd time.Choose and get high immune egg since the 20th d after the inoculation for the first time, contain the antibody of Kang Channel-catfish tarda BRK-ADHSJ in this immunity egg.
The antibody obtained by above-mentioned preparation method, can be applied in the medicine of prevention and treatment fish class Channel-catfish tarda disease.This antibody is as effective constituent, then adds pharmaceutically acceptable auxiliary material or carrier, makes various preparations and uses.
As a kind of preferred scheme, water is cooked auxiliary material, and water can be that physiological saline can be also distilled water, and antibody is made to aqua or special-purpose dipping bath agent use.Can among a small circle, directly soak healthy fish or sick fish, also can full pool spilling head, conveniently use.
As a kind of preferred scheme, adopt trehalose as carrier, antibody is made to pulvis, this pulvis is is directly thrown something and fed as fodder additives or pre-batching.Concrete making method is as follows: at first use 0.5% benzalkonium bromide solution or other similar thimerosal by immune egg soaking disinfection, then with aseptic water washing, dry.Yolk or whole egg are collected and homogenate to form immune egg liquid, and by spraying drying after the micro-coating of trehalose, make the immune powdered egg of the antibody of Han Kang Channel-catfish tarda BRK-ADHSJ, wherein, the 10%-20% that the weight of trehalose is immune egg liquid weight.
The antibody of prevention and treatment Yu Lei Channel-catfish tarda disease is made to pulvis, as fodder additives or pre-batching, directly throw something and feed, also can full pool spilling head, overcome other antibody drugs and can only inject or soak effectively shortcoming, use very conveniently, and greatly reduce labour intensity.
By the following examples the present invention is further detailed.
Embodiment six: the content detection of antibody in the immune powdered egg of prevention and Zhi Liao Channel-catfish tarda disease
1, material: the immune powdered egg of producing by above technique
2, method: application SDS-PAGE(sodium lauryl sulphate-polyacrylate hydrogel) cataphoretic determination method, the immune powdered egg of producing by above technique is detected, in the antibody that detected result is: Kang Channel-catfish tarda BRK-ADHSJ, pure IgY content is 85.98%.
Embodiment seven: the antibody titer that detects the immune powdered egg of Kang Channel-catfish tarda BRK-ADHSJ
1, material: detect with the antigen-scheme of the present invention of usining prepare separation and purification and through deactivation De Channel-catfish tarda BRK-ADHSJ vaccine as detectable antigens.
2, method: detect the antibody titer of the immune powdered egg of prepared De Kang Channel-catfish tarda BRK-ADHSJ by " ELSA " method, with the powdered egg prepared without the common birds, beasts and eggs of immunity in contrast, result is as table 4:
Table 4 antibody titer detected result
Above detected result shows that the immune powdered egg of Kang Channel-catfish tarda BRK-ADHSJ can be combined with the deactivation Hou of separation and purification gained De Channel-catfish tarda BRK-ADHSJ vaccine protein-specific, and there is very high antibodies and tire, and not immune egg fowl lay eggs the powdered egg of preparation only showed with this vaccine albumen very low in conjunction with tiring.Experiment results proved, immune powdered egg prepared by scheme of the present invention can specificity Yu Channel-catfish tarda BRK-ADHSJ vaccine protein binding, and there is very high binding ability.
The immune powdered egg of the antibody of embodiment eight: Han Kang Channel-catfish tarda BRK-ADHSJ is suffered from the preventive effect of " red head disease of Pseudobagrus " to Yellow catfish.
1, material: healthy Yellow catfish, take from Wujiang, Suzhou plant; The immune powdered egg of producing by above technique; Not immune egg gained powdered egg.
2, method: healthy Yellow catfish is divided into experimental group and control group, wherein experimental group is thrown something and fed 2 weeks with 0.2% addition with immune powdered egg of the present invention or the 2mg/L aquaculture water is splashed 3 days, after control group takes same means to process, Yong Channel-catfish tarda BRK-ADHSJ carries out artificial challenge's experiment to two groups of Yellow catfishs: test is the healthy Yellow catfish fingerling of taking from Wujiang, Suzhou plant with healthy Yellow catfish, specification is 35 ± 5g, laboratory is transported in the plastics bag oxygenation back, in aquarium, support temporarily, not bait throwing in during supporting temporarily, the test water body is through continuous oxygenation circulating filtration, guarantee that dissolved oxygen is more than 5mg/L, use automatic temperature control instrument Control experiment water temperature (28 ± 1) ℃, fish intramuscular injection 2.0 * 10 for test
7CFU/ml concentration De Channel-catfish tarda BRK-ADHSJ bacterium liquid 0.2ml/ tail.Test-results after 14 d is as table 5:
The prevention situation of the immune powdered egg of the antibody of table 5 Han Kang Channel-catfish tarda BRK-ADHSJ to Yellow catfish " red head disease of Pseudobagrus "
Above-mentioned test shows: through the present invention, prepared immune powdered egg is thrown something and fed or the experimental group Yellow catfish of splashing can produce the significantly ability of opposing " red head disease of Pseudobagrus ", and the sickness rate of control group approaches 100%.
The immune powdered egg of the antibody of embodiment nine: Han Kang Channel-catfish tarda BRK-ADHSJ is to infecting the result for the treatment of of " red head disease of Pseudobagrus " Yellow catfish.
1, material: healthy Yellow catfish, take from Wujiang, Suzhou plant; The immune powdered egg of producing by above technique; Not immune egg gained powdered egg.
2, method: at first Yong Channel-catfish tarda BRK-ADHSJ carries out Experimental infection to healthy Yellow catfish: test is the healthy Yellow catfish fingerling of taking from Wujiang, Suzhou plant with healthy Yellow catfish, specification is 35 ± 5g, laboratory is transported in the plastics bag oxygenation back, in aquarium, support temporarily, not bait throwing in during supporting temporarily, the test water body, through continuous oxygenation circulating filtration, guarantees that dissolved oxygen is more than 5mg/L; Use automatic temperature control instrument Control experiment water temperature (28 ± 1) ℃, fish intramuscular injection 2.0 * 10 for test
7CFU/ml concentration De Channel-catfish tarda BRK-ADHSJ bacterium liquid 0.2ml/ tail.
Then, in metainfective the 2nd day, the Yellow catfish of morbidity is divided into to two groups at random, and the amount that wherein experimental group is got feed 0.2% is added the immune powdered egg of the present invention 2 weeks or, with the amount of the 2mg/L 5d that splashes continuously, control group takes same method to not immune powdered egg of throwing something and feeding.The treatment situation is as table 6:
The treatment situation of the immune powdered egg of the antibody of table 6 Han Kang Channel-catfish tarda BRK-ADHSJ to Yellow catfish " red head disease of Pseudobagrus "
Test-results shows: immune powdered egg of the present invention is thrown something and fed or the enforcement of splashing can be good at treating experimental group and infects " red head disease of Pseudobagrus " Yellow catfish, and its curative ratio can reach 70%, and control group infects " red head disease of Pseudobagrus " Yellow catfish state of an illness and almost do not obtain any improvement.
Antibody of the present invention can be applied to known any method in the cultivation of the alepidotes such as Yellow catfish, Channel-catfish fish, catfish it as fodder additives by those of ordinary skills.
It should be noted that, above-described embodiment does not limit the present invention in any form, and all employings are equal to replaces or technical scheme that the mode of equivalent transformation obtains, all drops in protection scope of the present invention.
Claims (8)
1. the preparation method of the antibody of prevention and treatment Yu Lei Channel-catfish tarda disease, is characterized in that, use through inactivator deactivation De Channel-catfish tarda (
Edwardsiella ictaluri) gradation inoculation egg fowl, adaptive immune egg, the antibody that contains Kang Channel-catfish tarda in above-mentioned immune egg.
2. the preparation method of the antibody of prevention according to claim 1 and treatment Yu Lei Channel-catfish tarda disease, is characterized in that, Shang Shu Channel-catfish tarda is the bacterial strain that deposit number is CCTCC NO:M2012024.
3. the preparation method of the antibody of prevention according to claim 2 and treatment Yu Lei Channel-catfish tarda disease, is characterized in that, above-mentioned inactivator is formaldehyde.
4. the antibody of prevention and treatment fish class Channel-catfish tarda disease, is characterized in that, utilizes method claimed in claim 1 to make.
5. the antibody of prevention according to claim 4 and treatment Yu Lei Channel-catfish tarda disease, is characterized in that, above-mentioned tarda is the bacterial strain that deposit number is CCTCC NO:M2012024.
6. the application of antibody claimed in claim 4 in the medicine of preparation prevention and treatment Yu Lei Channel-catfish tarda disease.
7. application according to claim 6, is characterized in that, by fresh immune egg spraying drying after the micro-coating of trehalose, makes the pulvis that contains above-mentioned antibody, and above-mentioned pulvis is is directly thrown something and fed as fodder additives.
8. application according to claim 6, is characterized in that, water is cooked auxiliary material, and above-mentioned antibody is made to the aqua full pool spilling head.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012102692493A CN102936285A (en) | 2012-08-01 | 2012-08-01 | Antibody for preventing and treating fish Edwardsiellaictaluri disease, preparation method and applications thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012102692493A CN102936285A (en) | 2012-08-01 | 2012-08-01 | Antibody for preventing and treating fish Edwardsiellaictaluri disease, preparation method and applications thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN102936285A true CN102936285A (en) | 2013-02-20 |
Family
ID=47695216
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2012102692493A Pending CN102936285A (en) | 2012-08-01 | 2012-08-01 | Antibody for preventing and treating fish Edwardsiellaictaluri disease, preparation method and applications thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102936285A (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106620683A (en) * | 2015-10-29 | 2017-05-10 | 四川农业大学 | Bicombinant oral microsphere vaccine used for fish and purpose thereof |
| CN111100833A (en) * | 2019-12-31 | 2020-05-05 | 浙江省淡水水产研究所 | Recombinant strain for expressing outer membrane protein of Edwardsiella ictaluri, preparation method and application |
| CN111273007A (en) * | 2020-03-13 | 2020-06-12 | 内江师范学院 | Kit and detection method for rapidly detecting fish Edwardsiella ictaluri |
| CN116036101A (en) * | 2023-01-29 | 2023-05-02 | 广东省农业科学院动物科学研究所 | Application of trehalose in medicines for improving drug-resistant bacteria sensitivity and assisting chicken disease-resistant breeding |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101829321A (en) * | 2009-03-12 | 2010-09-15 | 通威股份有限公司 | Vaccine for preventing red head disease of Pseudobagrus fulvidraco |
-
2012
- 2012-08-01 CN CN2012102692493A patent/CN102936285A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101829321A (en) * | 2009-03-12 | 2010-09-15 | 通威股份有限公司 | Vaccine for preventing red head disease of Pseudobagrus fulvidraco |
Non-Patent Citations (2)
| Title |
|---|
| YONGPING XU: "Application of chicken egg yolk immunoglobulins in the control of terrestrial and", 《BIOTECHNOLOGY ADVANCES》 * |
| 王斌: "鸡抗迟缓爱德华氏菌卵黄抗体的应用研究", 《大连水产学院学报》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106620683A (en) * | 2015-10-29 | 2017-05-10 | 四川农业大学 | Bicombinant oral microsphere vaccine used for fish and purpose thereof |
| CN111100833A (en) * | 2019-12-31 | 2020-05-05 | 浙江省淡水水产研究所 | Recombinant strain for expressing outer membrane protein of Edwardsiella ictaluri, preparation method and application |
| CN111273007A (en) * | 2020-03-13 | 2020-06-12 | 内江师范学院 | Kit and detection method for rapidly detecting fish Edwardsiella ictaluri |
| CN116036101A (en) * | 2023-01-29 | 2023-05-02 | 广东省农业科学院动物科学研究所 | Application of trehalose in medicines for improving drug-resistant bacteria sensitivity and assisting chicken disease-resistant breeding |
| CN116036101B (en) * | 2023-01-29 | 2023-10-03 | 广东省农业科学院动物科学研究所 | Application of trehalose in improving drug-resistant bacteria sensitivity and assisting chicken disease-resistant breeding |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102154176B (en) | Turbot pathogenic strain and inactivated vaccine for ascites disease | |
| CN104498441B (en) | Duck hepatitis A virus (HAV) type III low virulent strain and live vaccine prepared therefrom and application | |
| CN102847146B (en) | Vaccine used for preventing tilapia streptococcal disease | |
| CN103834596A (en) | Bacillus subtilis shou003, anti-vibrio protein and preparation method and applications of bacillus subtilis shou003 and anti-vibrio protein | |
| CN109929772B (en) | Riemerella anatipestifer disease region dominant serotype strain and application thereof | |
| CN113583972A (en) | Escherichia coli bacteriophage capable of reducing antibiotic resistance and application thereof | |
| CN104232535B (en) | Nocardia seriolae induced low virulent strain and application thereof | |
| CN113430173A (en) | Wide-cracking-spectrum high-temperature-resistant salmonella bacteriophage and application thereof | |
| CN111254121A (en) | Salmonella bacteriophage and application thereof in medicine for preventing and treating salmonella infection diseases | |
| CN102936285A (en) | Antibody for preventing and treating fish Edwardsiellaictaluri disease, preparation method and applications thereof | |
| CN105039474A (en) | Preparation method of recombinant chicken interferon-alpha standard substance | |
| CN110468069A (en) | A kind of Lactobacillus casei YFI-5 and its application in anti-carp herpesviral II type | |
| CN105132310B (en) | One plant of cold water fish probiotics Bacillus strain and application thereof | |
| CN104611274A (en) | Haemophilus parasuis and application thereof | |
| Mohamed | Some studies on Pseudomonas species in chicken embryos and broilers in Assiut governorate | |
| CN105039241B (en) | Shelled Turtle Trionyx Sinensis heart cell continuous cell line and its construction method and cryopreservation method | |
| CN109207436A (en) | One plant of 4 type aviadenovirus strain of I group and its application | |
| CN110423714A (en) | A kind of lactic acid bacteria composite fungicide and its application in anti-carp herpesviral II type | |
| CN101829321B (en) | Vaccine for preventing red head disease of Pseudobagrus fulvidraco | |
| CN110075289A (en) | A kind of haemophilus parasuis, Streptococcus suis and Actinobacillus pleuropneumoniae triple inactivated vaccine and its application | |
| CN101531967B (en) | Method for producing alpha interferon and dedicated bacteria therefor | |
| CN100443502C (en) | Vitellus immune globulin for preventing prawn virus, tis preparing method and use thereof | |
| CN109266621A (en) | One plant of new light green color aerococcus bacteriophage AVP and application thereof | |
| CN105886429B (en) | A kind of Aeromonas hydrophila attenuation bacterium of antibiotic-free label and application | |
| CN110408573B (en) | Lactobacillus rhamnosus YFI-6 and application thereof in resisting giant salamander iridovirus |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C12 | Rejection of a patent application after its publication | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20130220 |