CN110029154A - The detection method of Mycoplasma synoviae in a kind of scale chicken house environment - Google Patents
The detection method of Mycoplasma synoviae in a kind of scale chicken house environment Download PDFInfo
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- CN110029154A CN110029154A CN201910405247.4A CN201910405247A CN110029154A CN 110029154 A CN110029154 A CN 110029154A CN 201910405247 A CN201910405247 A CN 201910405247A CN 110029154 A CN110029154 A CN 110029154A
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- 241000202942 Mycoplasma synoviae Species 0.000 title claims abstract description 58
- 238000001514 detection method Methods 0.000 title claims abstract description 34
- 241000287828 Gallus gallus Species 0.000 title claims abstract description 18
- 201000010099 disease Diseases 0.000 claims abstract description 43
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims abstract description 43
- 238000005070 sampling Methods 0.000 claims abstract description 34
- 239000007788 liquid Substances 0.000 claims abstract description 21
- 238000000034 method Methods 0.000 claims abstract description 20
- 229920000742 Cotton Polymers 0.000 claims abstract description 18
- 238000012360 testing method Methods 0.000 claims abstract description 16
- 150000007523 nucleic acids Chemical class 0.000 claims abstract description 14
- 102000039446 nucleic acids Human genes 0.000 claims abstract description 11
- 108020004707 nucleic acids Proteins 0.000 claims abstract description 11
- 238000004659 sterilization and disinfection Methods 0.000 claims abstract description 8
- 238000009736 wetting Methods 0.000 claims abstract description 8
- 239000000284 extract Substances 0.000 claims abstract description 5
- 238000007789 sealing Methods 0.000 claims description 14
- 238000004519 manufacturing process Methods 0.000 claims description 7
- 241000204031 Mycoplasma Species 0.000 claims description 5
- 241000204003 Mycoplasmatales Species 0.000 claims description 5
- 238000000246 agarose gel electrophoresis Methods 0.000 claims description 3
- 230000009514 concussion Effects 0.000 claims description 3
- 238000000605 extraction Methods 0.000 claims description 3
- 239000012634 fragment Substances 0.000 claims description 3
- 239000002775 capsule Substances 0.000 claims description 2
- 230000035479 physiological effects, processes and functions Effects 0.000 claims 1
- 150000003839 salts Chemical class 0.000 claims 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims 1
- 208000015181 infectious disease Diseases 0.000 abstract description 7
- 238000011156 evaluation Methods 0.000 abstract description 6
- 239000002504 physiological saline solution Substances 0.000 abstract description 6
- 230000002265 prevention Effects 0.000 abstract description 5
- 230000007613 environmental effect Effects 0.000 abstract description 3
- 244000052769 pathogen Species 0.000 abstract description 3
- 230000001717 pathogenic effect Effects 0.000 abstract description 3
- 241001260012 Bursa Species 0.000 description 6
- 230000000052 comparative effect Effects 0.000 description 3
- 238000012544 monitoring process Methods 0.000 description 3
- 230000002147 killing effect Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 208000035473 Communicable disease Diseases 0.000 description 1
- 206010017577 Gait disturbance Diseases 0.000 description 1
- 206010053759 Growth retardation Diseases 0.000 description 1
- 206010057190 Respiratory tract infections Diseases 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 230000003044 adaptive effect Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000002421 anti-septic effect Effects 0.000 description 1
- 238000009395 breeding Methods 0.000 description 1
- 230000001488 breeding effect Effects 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 231100000001 growth retardation Toxicity 0.000 description 1
- 230000005571 horizontal transmission Effects 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 238000004806 packaging method and process Methods 0.000 description 1
- 238000009304 pastoral farming Methods 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- 244000144977 poultry Species 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 230000005180 public health Effects 0.000 description 1
- 238000007430 reference method Methods 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6844—Nucleic acid amplification reactions
- C12Q1/6851—Quantitative amplification
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- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
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- Proteomics, Peptides & Aminoacids (AREA)
- Health & Medical Sciences (AREA)
- Biophysics (AREA)
- Chemical Kinetics & Catalysis (AREA)
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- Microbiology (AREA)
- Molecular Biology (AREA)
- Analytical Chemistry (AREA)
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- Biotechnology (AREA)
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- Bioinformatics & Cheminformatics (AREA)
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- Genetics & Genomics (AREA)
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Abstract
The application discloses a kind of detection method of Mycoplasma synoviae in scale chicken house environment.Sample collector needs band disposable plastic gloves and mask, immerses in the 10mL centrifuge tube equipped with 5mL physiological saline sterile cotton gauze with tweezers after disinfection and soaks；50cm long sampling area is determined in sampling location, is wiped 5 times back and forth with wetting cotton gauze, is filled in centrifuge tube and seal；It extracts liquid nucleic acid in centrifuge tube and carries out Mycoplasma synoviae Pathogen test.This method is by all kinds of body surface cotton swab in acquisition environment, and detecting it, whether there is or not Mycoplasma synoviae specific nucleic acid fragment and recall rates, analyzes and whether there is Mycoplasma synoviae cause of disease and environmental infection pressure in environment.The method of sampling has the characteristics that simple and easy, sample size is big, scheme is reproducible, and testing result can represent Mycoplasma synoviae pathogen capacity height in environment.Based on the identical method of sampling, evaluation criterion, it can be achieved that different field, not cause of disease content balance in same district, provide detection foundation for Mycoplasma synoviae prevention and control.
Description
Technical field
This application involves Mycoplasma synoviae detection technique field, in particular to bursa synovialis in a kind of scale chicken house environment
The detection method of mycoplasma.
Background technique
Mycoplasma synoviae mainly causes the infection of chicken and turkey, is poultry Important Infectious Diseases, is in recent years in severe infections
Trend causes the larger economic loss of farm.Mycoplasma synoviae mainly cause the respiratory tract infection of growth period chick, limping,
Growth retardation etc..Mycoplasma synoviae can be vertically spread, horizontal transmission causes chicken group's infection, so in monitoring provenance, environment
Mycoplasma synoviae cause of disease carrying rate is of great significance.
Existing method only provides the nucleic acid detection method of diagnostic sample (tissue sample, solution).Breeding enterprise is introduced a fine variety
Standard, effective evaluation method are needed to the Pathogen test of Mycoplasma synoviae, prevention early infection in environment in the process；Carry out ring
Lack strong operability, comparative high reference method for the evaluation of Mycoplasma synoviae cause of disease killing effect after the disinfection of border.
Summary of the invention
The detection method for being designed to provide Mycoplasma synoviae in a kind of scale chicken house environment of the application, to solve
After scale chicken house environment disinfected, lack strong operability, comparative senior staff officer for the evaluation of Mycoplasma synoviae cause of disease killing effect
The problem of test method.
According to an embodiment of the present application, a kind of detection method of Mycoplasma synoviae in scale chicken house environment is provided,
Include:
Sterile cotton gauze is immersed with the tweezers after disinfection and 5mL is housed by sample collector with disposable plastic gloves and mask
Wetting in the 10mL centrifuge tube of physiological saline；
The sampling area of 50cm long is determined in sampling location, is wiped 5 times back and forth with the cotton gauze of wetting, is filled in the centrifugation
It manages and seals；
Extract the detection of liquid nucleic acid progress Mycoplasma synoviae cause of disease in centrifuge tube after sealing.
Further, the sampling location includes metope, ground, stockline, waterline and egg rack, and the metope includes eminence wall
Face and lower metope, the ground include channel and coprodaecum.
Further, the method also includes: each sampling location at least acquires 6 parts of samples, and each sampling location is adopted
At least 50 square centimeters of sample region area.
Further, the centrifuge tube after the sealing at most saves 2 days under the conditions of 4 DEG C~10 DEG C.
Further, liquid nucleic acid carries out the detection packet of Mycoplasma synoviae cause of disease in centrifuge tube after the extraction sealing
It includes:
Centrifuge tube concussion mixes after sealing, and 0.25mL sample liquid is taken to extract sample liquid DNA；
Sample liquid DNA Mycoplasma synoviae universal primer is expanded into target fragment, obtains amplified production；
By the amplified production by 1.5% agarose gel electrophoresis, specific band at 206bp is observed, described in judgement
Sampling location whether there is Mycoplasma synoviae cause of disease.
Further, the detection of the Mycoplasma synoviae cause of disease further include:
It obtains test sample sum and there are the sample sizes of Mycoplasma synoviae cause of disease；
According to the test sample sum and described there are the sample size of Mycoplasma synoviae cause of disease, bursa synovialis branch is calculated
Substance cause of disease recall rate.
From the above technical scheme, the embodiment of the present application provides Mycoplasma synoviae in a kind of scale chicken house environment
Detection method, comprising: sample collector must band disposable plastic gloves and mask, sterile cotton gauze be immersed with tweezers after disinfection
It is soaked in 10mL centrifuge tube equipped with 5mL physiological saline；The sampling area for determining 50cm long wipes 5 with wetting cotton gauze back and forth
It is secondary, it fills in centrifuge tube and seals, extract the detection of liquid nucleic acid progress Mycoplasma synoviae cause of disease in centrifuge tube after sealing.This
Detection method, which passes through, acquires all kinds of body surface cotton swab in environment, and whether there is or not Mycoplasma synoviaes in detection cotton swab sample liquid
Specific nucleic acid fragment and recall rate number, analyze and whether there is Mycoplasma synoviae cause of disease and environmental infection pressure in environment.
The method of sampling has the characteristics that simple and easy, sample size is big, scheme is reproducible, and testing result can represent bursa synovialis in environment
The presence or absence of mycoplasma cause of disease and carrying capacity height.Based on the identical method of sampling, evaluation criterion, it can be achieved that different fields, difference
The comparison of cause of disease content in area provides detection foundation for Mycoplasma synoviae prevention and control.
Detailed description of the invention
In order to illustrate the technical solutions in the embodiments of the present application or in the prior art more clearly, below will be to institute in embodiment
Attached drawing to be used is needed to be briefly described, it should be apparent that, the accompanying drawings in the following description is only some implementations of the application
Example, for those of ordinary skill in the art, without creative efforts, can also obtain according to these attached drawings
Obtain other attached drawings.
Fig. 1 is to be preferably implemented to exemplify a kind of detection method stream of Mycoplasma synoviae in scale chicken house environment according to one
Cheng Tu.
Specific embodiment
Refering to fig. 1, the embodiment of the present application provides a kind of detection method of Mycoplasma synoviae in scale chicken house environment,
Include:
Step S1, sample collector is immersed sterile cotton gauze with the tweezers after disinfection with disposable plastic gloves and mask
It is soaked in 10mL centrifuge tube equipped with 5mL physiological saline；
Wherein, sterile cotton gauze is 6 layers of gauze of 2cm*2cm, is sterilized with suitable size newspaper independent packaging.
10mL plastic centrifuge tube, and pipe is provided with 5mL physiological saline, buckle closure after sterilizing are sealed with sealed membrane
Step S2, long 50cm sampling area is determined in sampling location, wiped 5 times back and forth with the cotton gauze of wetting, fill in institute
It states in centrifuge tube and seals；
Step S3, the detection of liquid nucleic acid progress Mycoplasma synoviae cause of disease in centrifuge tube after sealing is extracted.
The application utilizes PCR detection technique, can quickly and effectively evaluate Mycoplasma synoviae disease in scale livestock farming environment
Original is applicable in a large amount of sample acquisitions, laboratory testing analysis and environment disinfected on site and disappears to Mycoplasma synoviae there are situation
Malicious effect assessment.The application is equally applicable to the small-sized farm such as home farm, the important skill as Mycoplasma synoviae prevention and control
Art means.
Further, the sampling location includes metope, ground, stockline, waterline and egg rack, and the metope includes eminence wall
Face and lower metope, the ground include channel and coprodaecum.Chicken house environment includes each position that can be touched.The sampling of the application
Position is representative, including different direction, height, implements material, environment dead angle etc., has stronger Clinical significance of detecting, as a result has
There is persuasion.
Further, the method also includes: each sampling location at least acquires 6 parts of samples, and each sampling location is adopted
At least 50 square centimeters of sample region area.Each sampling location acquisition sample number is more, and sampling area is bigger, the knot of sampled data
Fruit is more accurate.
Further, the centrifuge tube after the sealing at most saves 2 days under the conditions of 4 DEG C~10 DEG C.What this method needed
Sampling article can prepare in advance on demand, be convenient for carrying transport.4 DEG C in transit~10 DEG C of test sample preservations, nucleic acid
The sample holding time was no more than 2 days before detecting.
Further, liquid nucleic acid carries out the detection packet of Mycoplasma synoviae cause of disease in centrifuge tube after the extraction sealing
It includes:
Centrifuge tube concussion mixes after sealing, and 0.25mL sample liquid is taken to extract sample liquid DNA；
Sample liquid DNA Mycoplasma synoviae universal primer is expanded into target fragment, obtains amplified production；
By the amplified production by 1.5% agarose gel electrophoresis, specific band at 206bp is observed, described in judgement
Sampling location whether there is Mycoplasma synoviae cause of disease.
The specific band of sample is judged to Sample Positive when consistent with positive control, shows to detect bursa synovialis branch original in sample
Body cause of disease.
Further, the detection of the Mycoplasma synoviae cause of disease further include:
It obtains test sample sum and there are the sample sizes of Mycoplasma synoviae cause of disease；
According to the test sample sum and described there are the sample size of Mycoplasma synoviae cause of disease, bursa synovialis branch is calculated
Substance cause of disease recall rate.
Recall rate is the ratio of positive sample quantity Yu test sample total amount.
The judgment criteria of chicken house environment is the height of positive rate: showing not detect synovia from environment when positive rate is zero
Capsule mycoplasma cause of disease, when detecting positive, having prompted corresponding sampling position, there are Mycoplasma synoviae cause of diseases.
Laboratory detection result is counted according to recall rate, and testing result feminine gender shows that Mycoplasma synoviae is not detected in environment
Cause of disease, and positive findings prompt corresponding sampled point there are Mycoplasma synoviae cause of disease, prompt to reinforce antiseptic hygiene dead angle.In addition it examines
The height of extracting rate may compare the Mycoplasma synoviae cause of disease relative loadings size of varying environment, have certain public health meaning.
Calculation: on absolute value, detection Mycoplasma synoviae, that is, exceeded；In relative value, there are tolerance intervals, can root
Depending on self-management target (disinfection control, routine monitoring early warning control etc.) difference.
The application specifies the sample collection method of monitoring chicken house Mycoplasma synoviae cause of disease, can be applied to different fields, no
With the scale chicken house in area etc., testing result is explicitly shown out Mycoplasma synoviae cause of disease, and there are situations.And apply this method energy
Compare the Mycoplasma synoviae recall rate of different places, have parallel comparative, has reference value to production management.
From the above technical scheme, the embodiment of the present application provides Mycoplasma synoviae in a kind of scale chicken house environment
Detection method, comprising: sample collector must band disposable plastic gloves and mask, sterile cotton gauze be immersed with tweezers after disinfection
It is soaked in 10mL centrifuge tube equipped with 5mL physiological saline；The sampling area for determining 50cm long wipes 5 with wetting cotton gauze back and forth
It is secondary, it fills in centrifuge tube and seals, extract the detection of liquid nucleic acid progress Mycoplasma synoviae cause of disease in centrifuge tube after sealing.This
Detection method, which passes through, acquires all kinds of body surface cotton swab in environment, and whether there is or not Mycoplasma synoviaes in detection cotton swab sample liquid
Specific nucleic acid fragment and recall rate number, analyze and whether there is Mycoplasma synoviae cause of disease and environmental infection pressure in environment.
The method of sampling has the characteristics that simple and easy, sample size is big, scheme is reproducible, and testing result can represent bursa synovialis in environment
The presence or absence of mycoplasma cause of disease and carrying capacity height.Based on the identical method of sampling, evaluation criterion, it can be achieved that different fields, difference
The comparison of cause of disease content in area provides detection foundation for Mycoplasma synoviae prevention and control.
Those skilled in the art will readily occur to its of the application after considering specification and practicing application disclosed herein
Its embodiment.This application is intended to cover any variations, uses, or adaptations of the application, these modifications, purposes or
Person's adaptive change follows the general principle of the application and including the undocumented common knowledge in the art of the application
Or conventional techniques.The description and examples are only to be considered as illustrative, and the true scope and spirit of the application are by following
Claim is pointed out.
It should be understood that the application is not limited to the precise structure that has been described above and shown in the drawings, and
And various modifications and changes may be made without departing from the scope thereof.Scope of the present application is only limited by the accompanying claims.
Claims (6)
1. the detection method of Mycoplasma synoviae in a kind of scale chicken house environment characterized by comprising
Sterile cotton gauze is immersed with the tweezers after disinfection and 5mL physiology is housed by sample collector with disposable plastic gloves and mask
Wetting in the 10mL centrifuge tube of salt water；
The sampling area of 50cm long is determined in sampling location, is wiped 5 times, is filled in the centrifuge tube back and forth with the cotton gauze of wetting
And it seals；
Extract the detection of liquid nucleic acid progress Mycoplasma synoviae cause of disease in centrifuge tube after sealing.
2. the method according to claim 1, wherein the sampling location includes metope, ground, stockline, waterline
And egg rack, the metope include eminence metope and lower metope, the ground includes channel and coprodaecum.
3. the method according to claim 1, wherein further include: each sampling location at least acquires 6 parts of samples,
At least 50 square centimeters of the sampling area area of each sampling location.
4. the method according to claim 1, wherein sealing after centrifuge tube under the conditions of 4 DEG C~10 DEG C at most
It saves 2 days.
5. the method according to claim 1, wherein liquid nucleic acid is slided in centrifuge tube after extraction sealing
The detection of liquid capsule mycoplasma cause of disease, comprising:
Centrifuge tube concussion mixes after sealing, and 0.25mL sample liquid is taken to extract sample liquid DNA；
Sample liquid DNA Mycoplasma synoviae universal primer is expanded into target fragment, obtains amplified production；
By the amplified production by 1.5% agarose gel electrophoresis, specific band at 206bp is observed, judges the sampling
Position whether there is Mycoplasma synoviae cause of disease.
6. according to the method described in claim 5, it is characterized in that, the detection of the Mycoplasma synoviae cause of disease further include:
It obtains test sample sum and there are the sample sizes of Mycoplasma synoviae cause of disease；
According to the test sample sum and described there are the sample size of Mycoplasma synoviae cause of disease, Mycoplasma synoviae is calculated
Cause of disease recall rate.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910405247.4A CN110029154A (en) | 2019-05-16 | 2019-05-16 | The detection method of Mycoplasma synoviae in a kind of scale chicken house environment |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201910405247.4A CN110029154A (en) | 2019-05-16 | 2019-05-16 | The detection method of Mycoplasma synoviae in a kind of scale chicken house environment |
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| CN110029154A true CN110029154A (en) | 2019-07-19 |
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109001450A (en) * | 2018-04-24 | 2018-12-14 | 中国农业科学院兰州兽医研究所 | A kind of kit and preparation method detecting chicken Mycoplasma synoviae antibody |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109001450A (en) * | 2018-04-24 | 2018-12-14 | 中国农业科学院兰州兽医研究所 | A kind of kit and preparation method detecting chicken Mycoplasma synoviae antibody |
Non-Patent Citations (2)
| Title |
|---|
| 周云锋: "一株鸡滑液囊支原体的分离鉴定", 《黑龙江畜牧兽医》 * |
| 韩雪娇: "某蛋种鸡场鸡滑液囊支原体传播规律及感染来源的研究", 《中国优秀硕士学位论文全文数据库》 * |
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Application publication date: 20190719 |