Some standard content:
【GB168821997】
Standard for monitoring plague in animals
Plague is a typical natural epidemic source disease. By the end of 1995, the natural epidemic sources that had been determined were distributed in 234 counties (cities, banners) in 17 provinces (autonomous regions) in my country. In order to implement the "Law of the People's Republic of China on the Prevention and Control of Infectious Diseases", through the monitoring of plague animal diseases, grasp the dynamics of the epidemic situation, evaluate the control effect, and provide a scientific basis for epidemic prediction and formulation of prevention and control countermeasures, this standard is specially formulated.
During the development process of this standard, my country's theoretical research results and field practical experience in plague monitoring were fully utilized and expressed in relevant chapters. This standard was proposed by the Ministry of Health of the People's Republic of China. The responsible drafting unit of this standard: National Plague and Brucellosis Prevention and Control Base; participating drafting units: Inner Mongolia Autonomous Region Epidemiology Prevention and Control Institute, Qinghai Provincial Endemic Disease Prevention and Control Institute, Yunnan Provincial Epidemiology Prevention and Control Institute.
Drafters of this standard: Li Shubao, Liu Jiyou, Li Tiehua, Li Chao, Ma Yongkang. This standard is interpreted by the Chinese Academy of Preventive Sciences, the technical unit entrusted by the Ministry of Health. 1 Scope
This standard specifies the main hosts, vectors, pathogens and serological monitoring standards of various plague foci in my country, as well as the quality control and technical methods of monitoring. This standard is applicable to the monitoring of plague foci of Himalayan marmot, gray marmot, long-tailed marmot, Mongolian marmot, Daurian ground squirrel, Alashan ground squirrel, long-tailed ground squirrel, long-clawed gerbil, Brandt's vole, large woolly mouse and house mouse (yellow-breasted mouse). Monitoring is based on counties (cities, banners) 2 Referenced standards
The provisions contained in the following standards constitute the provisions of this standard through reference in this standard. The versions shown at the time of publication of this standard are valid. All standards will be revised, and the parties using this standard should explore the possibility of using the latest versions of the following standards. GB159781995 Standards and principles for the treatment of human plague epidemic areas GB15991-1995 Plague diagnostic standards
3 Definitions
This standard adopts the following definitions.
3.1 Natural plague foci: During the epidemic of animal plague, plague bacteria parasitize specific hosts and are mainly transmitted between hosts and other animals through vector fleas. They are independent of humans and continue to circulate in nature for a long time, and can cause human plague epidemics. This phenomenon is called the natural plague foci. Places with natural foci are called natural plague foci. 3.2 Animal plague monitoring: Regularly and quantitatively monitor the epidemic dynamics of animal plague in the plague foci, observe the ecology of host animals and vector insects, and study the infection, transmission, preservation laws and geographical distribution characteristics of animal plague.
3.3 Main host: Species that can ensure the long-term continuation of plague bacteria in a specific ecosystem.
3.4 Epidemic area: Area where plague occurs or spreads among humans or animals. 3.5
Epidemic point: Local area where human or animal plague occurs. 3.6
Infected rat: Rodents that are naturally infected and have plague bacteria isolated from their bodies are called infected rats.
3.7 Clustering: Animal plague appears in groups in time, space, or both time and space.
3.8 Optimal habitat: The natural environment that is most suitable for the host animal to live and survive. 3.9 Mobile monitoring points: The monitoring range is not less than 2500ha, mainly conducting self-dead rodent inspection bacteria and serum epidemiological monitoring, and focusing on mastering the number of major hosts and vector insects. The monitoring time is generally 20 days.
3.10 Fixed monitoring points: The monitoring range is 10,000ha, and long-term systematic observations are made on the changes in geographical habitats, the number of host animals, vector insects, and plague bacteria to master the dynamics and study the epidemic and preservation laws of plague animal diseases. The monitoring time for each point is 3 to 5 years. 4 Animal plague monitoring standards
4.1 The monitoring period of the source of yellow rats (including Daurian, Alxa, and long-tailed yellow rats, the same below) is from April to September: the scope of mobile monitoring points is 2500ha, and the scope of fixed monitoring points is 10000ha; the habitat is divided by three indicators: landform, vegetation, and number of yellow rats, and a habitat distribution map with a scale of 1:10000 is drawn. Stratified sampling is carried out according to the proportion of 0.5% of the area of each type of habitat. The number of yellow rats is monitored once a day in April and July by the bow-clip method of one hectare; at least 20 live yellow rats are sampled every ten days for flea sampling; 300 to 500 animals are inspected at fixed points for animal bacteria testing, and more than 90 groups of fleas are tested for bacteria; serum for serological testing accounts for no less than 8% of the predicted number of rats.
4.2. The monitoring period of the marmot (including Himalayan, gray, long-tailed, Mongolian marmot, the same below) epidemic source is from April to early October; the mobile point is 5,000ha, and the fixed point is 22,500ha; the geographical habitat is divided by three indicators: landform vegetation and the number of marmots, and a habitat map with a scale of 1:10,000 is drawn. The number of marmots is monitored by stratified sampling at a ratio of 0.5% of the area of each habitat. The route method is used for mobile monitoring points: 5 representative routes are selected, each route is 5km long, the field of vision is 50m wide, walking is 3km per hour, and riding is 5km per hour. The survey area is calculated by multiplying the route length by the field of vision width, and finally the density of marmots per hectare is calculated. Surveys are conducted once a month or in May and July. The sampling of marmot fleas is conducted at least 30 marmots every ten days; the detection of marmot bacteria is mainly based on sick and dead marmots. When the density of marmots is 0.1/ha, 10% of the number of marmots will be sampled, and when the density is above 0.2/ha, 5% of the number of marmots will be sampled. The number of fleas should be no less than 250 groups, and the serological test should be based on the pastoral points of natural villages, with 5% of the dog serum sampled in pastoral areas and 20% of the dog serum sampled in agricultural areas. The focus of the Mongolian marmot epidemic source is mainly on the number of marmots and serological monitoring. 4.3 Long-clawed gerbil epidemic source
Monitoring time: April to May, October to November. The mobile monitoring points are 2500ha, and the fixed monitoring points are 10000ha. The habitat is divided by four indicators: landform, vegetation, soil, and the number of long-clawed gerbils, and a topographic map with a scale of 1:10000 is drawn. Stratified sampling is carried out at a rate of 0.5% for mobile points and 0.2% to 0.5% for fixed points. The number of long-clawed gerbils is investigated by the overlapping night bow-trap method with one hectare as the unit. At least 20 live fleas are sampled at each point every ten days, and more than 100 fleas are sampled throughout the year: the host bacteria test focuses on finding dead rats. No less than 500 captured rats are tested for bacteria at each point throughout the year, and all fleas obtained are tested for bacteria; 200 to 500 rat serum samples are tested at each point. 4.4 Brandt's vole epidemic source
Monitoring time: April to May, August to September. The monitoring range is not less than 10,000ha, and the search control range is 100,000ha. The habitat division refers to the long-clawed gerbil. Stratified sampling is carried out at a rate of 0.2% of the area of each type of habitat, and the number of Brandt's vole is monitored by the one-day-one-male-skull cloth-trap method. For flea sampling, 100 Brandt's voles should be combed and checked at each point; host bacteria testing should focus on finding dead mice, and more than 300 rodents should be tested at each point throughout the year, and all fleas obtained should be tested for bacteria; more than 10C copies of Brandt's voles serum should be tested at each point.
4.5 Large woolly plague source
Monitoring time: January to December, with March to August and December as the focus. Fixed and mobile monitoring points cover an area of 2000ha, with one fixed point and 2 to 4 mobile points established each year in each county. For host number monitoring, fixed points are sampled once a month, and mobile points are sampled once a quarter. Each time, 100 mouse cages are placed using the cage night method for three consecutive nights to calculate the capture rate. For flea sampling, more than 20 large woolly rats and field mice are recommended for each habitat; no less than 500 hosts are tested at each fixed point throughout the year, and more than 100 groups of fleas are tested throughout the year; pay attention to collecting the serum of hunting and herding dogs and the serum of wild animals (fox, catfish, weasel, etc.) in the epidemic source, and test more than 150 serum samples of various rodents and indicator animals.
4.6 Domestic rat (yellow-breasted rat) epidemic source
Monitoring time: January to December. Guangdong March to May, Fujian April to October, and western Yunnan May to September are the focus. The fixed point monitoring range is 2000ha, and the mobile point monitoring range is 500-1000ha. Within this range, 100-200 households in villages (villages) are selected for fixed points, and 50-100 households in villages (villages) are selected for mobile points for monitoring. Each county has one fixed point and 2 to 4 mobile points; for the survey of domestic rodents, 100 representative rooms are selected each month, and a mousetrap (cage) is set in each room for three consecutive days; for wild nocturnal mice, 100 traps are set with a 5m trap line method for two consecutive nights, and the capture rates indoors and outdoors are calculated respectively. For body flea sampling, 20 domestic mice (yellow-breasted mice) are caught alive each month; 300 to 500 animals are tested for bacteria at fixed points, and flea tests are conducted for no less than 100 groups; 50 to 100 serum samples are tested monthly for serology, and 500 to 1000 samples are tested throughout the year.
Appendix A
A1 Method for surveying the density of yellow rats
(Standard Appendix)
Method for surveying the density of major host animals
A1.1 Survey time: Two surveys are conducted in April and July each year. Sampling method: Sample plots are drawn in layers according to the proportion of 0.5% of the area of each habitat in the monitoring area. A1.2
The spacing between sample plots is greater than 200m.
Sample plot measurement: Each sample plot is a unit (100m×100m), and the four corners are marked.
A1.4: Check holes: Check holes at 5m intervals in the sample plot, and mark them after finding them A1.5 Traps: Before the yellow squirrels come out of their holes, use the pit trap method and patrol once every 2 hours during the trapping time.
A1.6 Calculation: Calculate according to formula (A1).
Number of rats
Yellow squirrel densityOne-way square area (ha)
Marmot density survey method
And route.
Survey time: Survey once in May and July each yearSampling method: Stratified sampling of sampling plots at a ratio of 0.5% of the area of each habitat in the monitoring areaSurvey method:
Visual method
Select 5 to 10 representative habitat plots in the monitoring area, each plot is 5 to 10 hectares. First, mark the area around the plot to determine the scope of the plot, select a place 100 to 200 meters away from the plot, use geographical barriers or shelters as cover, and use a telescope to observe the number of marmots in the plot when the marmots are active every day. Observe for 2 consecutive days, twice a day. The density is calculated by the highest value at one time.
Early climbing density is the same as the ground distance (ha)
Early climbing number
A2.3.2 Route method
（A2)
In the monitoring area, select 5 to 10 representative monitoring routes, each line is 10ha. The route field width is 50 to 100m, the route distance is based on walking 3km/h, riding 5km/h, early climbing
Early climbing density = exposure line length × standard field width/1000mA3 Density survey of long-clawed gerbils and Brandt's voles.（A3)
A3.1 Survey time: Long-clawed gerbils and Brandt's voles are monitored once in spring and autumn. A3.2 Sampling area: In the monitoring area, stratified sampling is carried out at a ratio of 0.5% of the habitat area, and the sample plot is 1ha (100m×100m) as a unit. A3.3 Sample plot measurement and hole checking: Same as yellow rat (A1.3, A1.4). A3.4 All rat holes found during the hole checking with traps shall be blocked. At 7-8 am the next day, a plate of traps shall be placed at each hole entrance that was stolen. The trapping time is 24 hours. Patrol once every 2 hours during the day, once at dusk and dawn. During the patrol, remove the rats caught and re-trap the rats at the original hole entrance, and continue to hit.
Rat density: Calculate according to formula (A4).
Number of traps
Rat densityArea of a group (h)
A4 Survey of night-walking rat density in the wild
（A4)
A4.1 Number of traps (cages) to be investigated: In units of months, select 1 to 3 representative habitats, and set up 100 traps (cages)/time in each habitat. In the source of large woolly plague, the traps (cages) are set up for 3 consecutive days each time, and in the source of domestic plague, the traps (cages) are set up for 2 consecutive days each time. A4.2 Rat trapping tools: The models of traps (cages) should be unifiedA4.3 Bait: The plate traps are uniformly made of white flour (vegetable oil) oil cakes, and the rat cages are made of fried dough sticks or sweet potatoes, each piece is 3 to 5 mg.
A4.4 Deployment method: Use the 5m trap (cage) line method for deployment, the spacing between the traps (cages) should exceed 200m, and the direction of the trap mouth and the cage mouth should be consistent. A4.5 Laying time: Lay the trap after sunset and retrieve it before sunrise. If there is heavy wind and rain during the time when the trap is in place, the capture rate cannot be calculated. A4.6 Capture rate: Calculate according to formula (A5).
Number of rats caught
Capture rate (%) = (number of days × 100A5 Survey on density of domestic rodents
(A5)
A5.1 Number of surveys: In the monitoring area, select a representative village (village) every month, select 100 houses (including warehouses) in the village (village), and place a plate (piece) of traps (cages) in each house. A5.2 Rat-catching tools: the same as night-walking rats in the wild. A5.3 Bait: the same as night-walking rats in the wild.
A5.4 Time and method of placement: Continuously place for 48 to 72 hours/time, patrol once in the morning and evening every day, remove the captured rats, and replace the bait. A5.5 Capture rate: calculated according to formula (A6).
Catch* (%) = (number of days × 100
Number of rats caught
Appendix B||t t||(Standard Appendix)
B1 Survey Quantity
Survey Method for Rat Flea Index
During the monitoring period, 20 live main host animals are randomly selected for flea inspection every ten days in various plague natural foci.
B2 Survey Method
The captured live main host animals are individually bagged and anesthetized with ether in the flea inspection room. The fleas are combed with a comb or brush to obtain fleas for identification and classification. The fleas are grouped and sent for inspection according to the same host, the same direction, and the same species.
B3 Calculation
Total number of eggs
Rat total egg index No. 1
Total number of rats
Base flea number
Fruit egg index No. 1
Total number of rats
Limited egg infection rate (%)
Infected rats
Total number of mice
C1 Operation method
Appendix C
(B2)
(B3)
(Standard Appendix)
Indirect hemagglutination test method
Arrange the small test tubes on the test tube rack, mark the tube numbers, add 0.9mL of diluent to the first tube, and then add 0.5mL to each tube. Take 0.1mL of the test serum, add it to the first tube, mix it and dilute it in multiples, and take 0.5mL from the last tube and discard it. Then use a 1mL graduated pipette to add a drop of 2.5% F antigen-sensitized blood cells (0.05mL) to each tube, oscillate to mix the blood cells, place it in a 37℃ incubator or at room temperature for 2 to 3 hours to observe the results. For suspected materials, make 6 tubes, and each batch of tests should add 0.5mL of diluent plus a drop of 2.5% F antigen-sensitized blood cells as a control. C2 Result Judgment The positive result is defined as the uneven distribution of red blood cells at the bottom of the tube, with umbrella-shaped or irregular rough edges. The positive results are further divided according to the red blood cells at the bottom of the tube and the degree of roughness of the edges:
C2.1++++: Tight agglutination, agglutination covers the bottom of the tube, with obvious folded edges. When the antibody is excessive, it agglutinates into a sparse flower cluster. C2.2+++: Relatively tight agglutination, agglutination covers the bottom of the tube, in a disc shape without folded edges.
C2.3+++: Incomplete agglutination of blood cells, the bottom of the tube is a neat circle, and there is obvious agglutination of blood cells inside and outside the circle.
C2.4+: There are more non-agglutinated blood cells, which sink to the bottom of the tube and present a neat circle, and there are very few agglutinated blood cells outside the circle.
C2.5-: The blood cells do not agglutinate, sink to the bottom of the tube, and present a neat circle. The test tube method uses "10+10" as the positive limit of the indirect hemagglutination test. Red blood cells are densely packed in the shape of buttons at the bottom of the tube, with smooth and neat edges, or appear as a narrow circle, which is a negative result
Appendix D
(Standard Appendix)
Quality Control Standard for Plague Indirect Hemagglutination Test
D1 Inspection Material Standard
The glassware used is cleaned, dried and sterilized, the measuring instrument is calibrated and qualified, the diagnostic Fi antigen, F sensitized blood cells, diluent pH 6.9-7.1), ion concentration 0.15mol/LNaCl and plague diagnostic serum are prepared by a unified method, qualified in quality inspection, and stored at 4°C. D2 Serum sample collection and storage standards
Pay attention to sterility when collecting blood and separating serum to prevent hemolysis and contamination. The serum should be no less than 0.5mL. The serum of the test subject does not need to be inactivated and absorbed. The serum should be used for the test as soon as possible. If it cannot be tested immediately, sodium azide or thimerosal (final concentration of the former is 0.1% and the latter is 0.01%) should be added or frozen for storage, but it cannot exceed half a month. D3 Preparation standard of quality control serum (internal control sample) Take 10μL plague diagnosis serum with a micro syringe and add 9.9mL homologous normal serum, mix thoroughly, then take 100μL accurately with a micro syringe, and inject each into An Yan, deep freeze or freeze vacuum dry for storage
D4 "True value" determination standard of quality control serum
Add 1mL diluent to the quality control serum An Yan, mix well, take 0.5mL and put it into the second tube for continuous multiple dilution, put the whole amount of An Shao into the first tube, which is regarded as 1:10, shake and mix well, add 1 drop of 2.5%Fi blood cell to each tube with a 1mL pipette, mix well at room temperature for 2-3 hours and observe the results. Test the quality control serum once a day for 20 consecutive days, and the titer with the highest frequency (at least 17 times) is regarded as the "true value". The batch is considered qualified when the ratio of the result to the "true value" titer does not exceed ±1 dilution. In actual work, one internal control sample is tested for each batch of test samples. D5 Participation in inter-laboratory quality control standards
Inter-laboratory quality control standards: The external control serum samples issued by the quality control reference laboratory are qualified if the titer is ±1 of the standard. The test samples in actual work should be treated the same as the control samples. When the tested serum shows a positive reaction of more than 1:20, it must be inactivated and absorbed, and three rows of retests must be performed to determine the titer. The plague hemagglutination positive reaction standard and diagnostic standard shall be implemented in accordance with the "Plague Serological Diagnostic Standards".6 Calculation: Calculate according to formula (A1).
Number of swatters
Density of yellow squirrels
Survey method of marmot density
And route.
Survey time: Survey once in May and July each year Sampling method: Stratified sampling of sampling plots at a ratio of 0.5% of the area of each habitat in the monitoring area Survey method:
Visual method
Select 5 to 10 representative habitat plots in the monitoring area, each plot is 5 to 10 ha. First, mark the area around the plot to determine the scope of the plot, select a place 100 to 200 m away from the plot, use a geographical barrier or shelter as cover, and use a telescope to observe the number of marmots in the plot during the most active days of the marmots, and observe for 2 consecutive days, twice a day. Calculate the density based on the highest value of one time.
Early climbing density is the same as the ground length (ha)
Early climbing number
A2.3.2 Route method
（A2)
In the monitoring area, select 5 to 10 representative monitoring routes, each line is 10ha. The route field width is 50 to 100m, the route distance is based on walking 3km/h, riding 5km/h, early climbing
Early climbing density = line length × standard field width/1000mA3 Density survey of long-clawed gerbils and Brandt's voles.（A3)
A3.1 Survey time: Long-clawed gerbils and Brandt's voles are monitored once in spring and autumn. A3.2 Sampling area: In the monitoring area, stratified sampling is carried out at a ratio of 0.5% of the habitat area, and the sample plot is 1ha (100m×100m) as a unit. A3.3 Sample plot measurement and hole checking: Same as yellow rat (A1.3, A1.4). A3.4 All rat holes found during the hole checking with traps shall be blocked. At 7-8 am the next day, a plate of traps shall be placed at each hole entrance that was stolen. The trapping time is 24 hours. Patrol once every 2 hours during the day, once at dusk and dawn. During the patrol, remove the rats caught and re-trap the rats at the original hole entrance, and continue to hit.
Rat density: Calculate according to formula (A4).
Number of traps
Rat densityArea of a group (h)
A4 Survey of night-walking rat density in the wild
（A4)
A4.1 Number of traps (cages) to be investigated: In units of months, select 1 to 3 representative habitats, and set up 100 traps (cages)/time in each habitat. In the source of large woolly plague, the traps (cages) are set up for 3 consecutive days each time, and in the source of domestic plague, the traps (cages) are set up for 2 consecutive days each time. A4.2 Rat trapping tools: The models of traps (cages) should be unifiedA4.3 Bait: The plate traps are uniformly made of white flour (vegetable oil) oil cakes, and the rat cages are made of fried dough sticks or sweet potatoes, each piece is 3 to 5 mg.
A4.4 Deployment method: Use the 5m trap (cage) line method for deployment, the spacing between the traps (cages) should exceed 200m, and the direction of the trap mouth and the cage mouth should be consistent. A4.5 Laying time: Lay the trap after sunset and retrieve it before sunrise. If there is heavy wind and rain during the time when the trap is in place, the capture rate cannot be calculated. A4.6 Capture rate: Calculate according to formula (A5).
Number of rats caught
Capture rate (%) = (number of days × 100A5 Survey on density of domestic rodents
(A5)
A5.1 Number of surveys: In the monitoring area, select a representative village (village) every month, select 100 houses (including warehouses) in the village (village), and place a plate (piece) of traps (cages) in each house. A5.2 Rat-catching tools: the same as night-walking rats in the wild. A5.3 Bait: the same as night-walking rats in the wild.
A5.4 Time and method of placement: Continuously place for 48 to 72 hours/time, patrol once in the morning and evening every day, remove the captured rats, and replace the bait. A5.5 Capture rate: calculated according to formula (A6).
Catch* (%) = (number of days × 100
Number of rats caught
Appendix B||t t||(Standard Appendix)
B1 Survey Quantity
Survey Method for Rat Flea Index
During the monitoring period, 20 live main host animals are randomly selected for flea inspection every ten days in various plague natural foci.
B2 Survey Method
The captured live main host animals are individually bagged and anesthetized with ether in the flea inspection room. The fleas are combed with a comb or brush to obtain fleas for identification and classification. The fleas are grouped and sent for inspection according to the same host, the same direction, and the same species.
B3 Calculation
Total number of eggs
Rat total egg index No. 1
Total number of rats
Base flea number
Fruit egg index No. 1
Total number of rats
Limited egg infection rate (%)
Infected rats
Total number of mice
C1 Operation method
Appendix C
(B2)
(B3)
(Standard Appendix)
Indirect hemagglutination test method
Arrange the small test tubes on the test tube rack, mark the tube numbers, add 0.9mL of diluent to the first tube, and then add 0.5mL to each tube. Take 0.1mL of the test serum, add it to the first tube, mix it and dilute it in multiples, and take 0.5mL from the last tube and discard it. Then use a 1mL graduated pipette to add a drop of 2.5% F antigen-sensitized blood cells (0.05mL) to each tube, oscillate to mix the blood cells, place it in a 37℃ incubator or at room temperature for 2 to 3 hours to observe the results. For suspected materials, make 6 tubes, and each batch of tests should add 0.5mL of diluent plus a drop of 2.5% F antigen-sensitized blood cells as a control. C2 Result Judgment The positive result is defined as the uneven distribution of red blood cells at the bottom of the tube, with umbrella-shaped or irregular rough edges. The positive results are further divided according to the red blood cells at the bottom of the tube and the degree of roughness of the edges:
C2.1++++: Tight agglutination, agglutination covers the bottom of the tube, with obvious folded edges. When the antibody is excessive, it agglutinates into a sparse flower cluster. C2.2+++: Relatively tight agglutination, agglutination covers the bottom of the tube, in a disc shape without folded edges.
C2.3+++: Incomplete agglutination of blood cells, the bottom of the tube is a neat circle, and there is obvious agglutination of blood cells inside and outside the circle.
C2.4+: There are more non-agglutinated blood cells, which sink to the bottom of the tube and present a neat circle, and there are very few agglutinated blood cells outside the circle.
C2.5-: The blood cells do not agglutinate, sink to the bottom of the tube, and present a neat circle. The test tube method uses "10+10" as the positive limit of the indirect hemagglutination test. Red blood cells are densely packed in the shape of buttons at the bottom of the tube, with smooth and neat edges, or appear as a narrow circle, which is a negative result
Appendix D
(Standard Appendix)
Quality Control Standard for Plague Indirect Hemagglutination Test
D1 Inspection Material Standard
The glassware used is cleaned, dried and sterilized, the measuring instrument is calibrated and qualified, the diagnostic Fi antigen, F sensitized blood cells, diluent pH 6.9-7.1), ion concentration 0.15mol/LNaCl and plague diagnostic serum are prepared by a unified method, qualified in quality inspection, and stored at 4°C. D2 Serum sample collection and storage standards
Pay attention to sterility when collecting blood and separating serum to prevent hemolysis and contamination. The serum should be no less than 0.5mL. The serum of the test subject does not need to be inactivated and absorbed. The serum should be used for the test as soon as possible. If it cannot be tested immediately, sodium azide or thimerosal (final concentration of the former is 0.1% and the latter is 0.01%) should be added or frozen for storage, but it cannot exceed half a month. D3 Preparation standard of quality control serum (internal control sample) Take 10μL plague diagnosis serum with a micro syringe and add 9.9mL homologous normal serum, mix thoroughly, then take 100μL accurately with a micro syringe, and inject each into An Yan, deep freeze or freeze vacuum dry for storage
D4 "True value" determination standard of quality control serum
Add 1mL diluent to the quality control serum An Yan, mix well, take 0.5mL and put it into the second tube for continuous multiple dilution, put the whole amount of An Shao into the first tube, which is regarded as 1:10, shake and mix well, add 1 drop of 2.5%Fi blood cell to each tube with a 1mL pipette, mix well at room temperature for 2-3 hours and observe the results. Test the quality control serum once a day for 20 consecutive days, and the titer with the highest frequency (at least 17 times) is regarded as the "true value". The batch is considered qualified when the ratio of the result to the "true value" titer does not exceed ±1 dilution. In actual work, one internal control sample is tested for each batch of test samples. D5 Participation in inter-laboratory quality control standards
Inter-laboratory quality control standards: The external control serum samples issued by the quality control reference laboratory are qualified if the titer is ±1 of the standard. The test samples in actual work should be treated the same as the control samples. When the tested serum shows a positive reaction of more than 1:20, it must be inactivated and absorbed, and three rows of retests must be performed to determine the titer. The plague hemagglutination positive reaction standard and diagnostic standard shall be implemented in accordance with the "Plague Serological Diagnostic Standards".6 Calculation: Calculate according to formula (A1).
Number of swatters
Density of yellow squirrels
Survey method of marmot density
And route.
Survey time: Survey once in May and July each year Sampling method: Stratified sampling of sampling plots at a ratio of 0.5% of the area of each habitat in the monitoring area Survey method:
Visual method
Select 5 to 10 representative habitat plots in the monitoring area, each plot is 5 to 10 ha. First, mark the area around the plot to determine the scope of the plot, select a place 100 to 200 m away from the plot, use a geographical barrier or shelter as cover, and use a telescope to observe the number of marmots in the plot during the most active days of the marmots, and observe for 2 consecutive days, twice a day. Calculate the density based on the highest value of one time.
Early climbing density is the same as the ground length (ha)
Early climbing number
A2.3.2 Route method
（A2)
In the monitoring area, select 5 to 10 representative monitoring routes, each line is 10ha. The route field width is 50 to 100m, the route distance is based on walking 3km/h, riding 5km/h, early climbing
Early climbing density = line length × standard field width/1000mA3 Density survey of long-clawed gerbils and Brandt's voles.（A3)
A3.1 Survey time: Long-clawed gerbils and Brandt's voles are monitored once in spring and autumn. A3.2 Sampling area: In the monitoring area, stratified sampling is carried out at a ratio of 0.5% of the habitat area, and the sample plot is 1ha (100m×100m) as a unit. A3.3 Sample plot measurement and hole checking: Same as yellow rat (A1.3, A1.4). A3.4 All rat holes found during the hole checking with traps shall be blocked. At 7-8 am the next day, a plate of traps shall be placed at each hole entrance that was stolen. The trapping time is 24 hours. Patrol once every 2 hours during the day, once at dusk and dawn. During the patrol, remove the rats caught and re-trap the rats at the original hole entrance, and continue to hit.
Rat density: Calculate according to formula (A4).
Number of traps
Rat densityArea of a group (h)
A4 Survey of night-walking rat density in the wild
（A4)
A4.1 Number of traps (cages) to be investigated: In units of months, select 1 to 3 representative habitats, and set up 100 traps (cages)/time in each habitat. In the source of large woolly plague, the traps (cages) are set up for 3 consecutive days each time, and in the source of domestic plague, the traps (cages) are set up for 2 consecutive days each time. A4.2 Rat trapping tools: The models of traps (cages) should be unifiedA4.3 Bait: The plate traps are uniformly made of white flour (vegetable oil) oil cakes, and the rat cages are made of fried dough sticks or sweet potatoes, each piece is 3 to 5 mg.
A4.4 Deployment method: Use the 5m trap (cage) line method for deployment, the spacing between the traps (cages) should exceed 200m, and the direction of the trap mouth and the cage mouth should be consistent. A4.5 Laying time: Lay the trap after sunset and retrieve it before sunrise. If there is heavy wind and rain during the time when the trap is in place, the capture rate cannot be calculated. A4.6 Capture rate: Calculate according to formula (A5).
Number of rats caught
Capture rate (%) = (number of days × 100A5 Survey on density of domestic rodents
(A5)
A5.1 Number of surveys: In the monitoring area, select a representative village (village) every month, select 100 houses (including warehouses) in the village (village), and place a plate (piece) of traps (cages) in each house. A5.2 Rat-catching tools: the same as night-walking rats in the wild. A5.3 Bait: the same as night-walking rats in the wild.
A5.4 Time and method of placement: Continuously place for 48 to 72 hours/time, patrol once in the morning and evening every day, remove the captured rats, and replace the bait. A5.5 Capture rate: calculated according to formula (A6).
Catch* (%) = (number of days × 100
Number of rats caught
Appendix B||t t||(Standard Appendix)
B1 Survey Quantity
Survey Method for Rat Flea Index
During the monitoring period, 20 live main host animals are randomly selected for flea inspection every ten days in various plague natural foci.
B2 Survey Method
The captured live main host animals are individually bagged and anesthetized with ether in the flea inspection room. The fleas are combed with a comb or brush to obtain fleas for identification and classification. The fleas are grouped and sent for inspection according to the same host, the same direction, and the same species.
B3 Calculation
Total number of eggs
Rat total egg index No. 1
Total number of rats
Base flea number
Fruit egg index No. 1
Total number of rats
Limited egg infection rate (%)
Infected rats
Total number of mice
C1 Operation method
Appendix C
(B2)
(B3)
(Standard Appendix)
Indirect hemagglutination test method
Arrange the small test tubes on the test tube rack, mark the tube numbers, add 0.9mL of diluent to the first tube, and then add 0.5mL to each tube. Take 0.1mL of the test serum, add it to the first tube, mix it and dilute it in multiples, and take 0.5mL from the last tube and discard it. Then use a 1mL graduated pipette to add a drop of 2.5% F antigen-sensitized blood cells (0.05mL) to each tube, oscillate to mix the blood cells, place it in a 37℃ incubator or at room temperature for 2 to 3 hours to observe the results. For suspected materials, make 6 tubes, and each batch of tests should add 0.5mL of diluent plus a drop of 2.5% F antigen-sensitized blood cells as a control. C2 Result Judgment The positive result is defined as the uneven distribution of red blood cells at the bottom of the tube, with umbrella-shaped or irregular rough edges. The positive results are further divided according to the red blood cells at the bottom of the tube and the degree of roughness of the edges:
C2.1++++: Tight agglutination, agglutination covers the bottom of the tube, with obvious folded edges. When the antibody is excessive, it agglutinates into a sparse flower cluster. C2.2+++: Relatively tight agglutination, agglutination covers the bottom of the tube, in a disc shape without folded edges.
C2.3+++: Incomplete agglutination of blood cells, the bottom of the tube is a neat circle, and there is obvious agglutination of blood cells inside and outside the circle.
C2.4+: There are more non-agglutinated blood cells, which sink to the bottom of the tube and present a neat circle, and there are very few agglutinated blood cells outside the circle.
C2.5-: The blood cells do not agglutinate, sink to the bottom of the tube, and present a neat circle. The test tube method uses "10+10" as the positive limit of the indirect hemagglutination test. Red blood cells are densely packed in the shape of buttons at the bottom of the tube, with smooth and neat edges, or appear as a narrow circle, which is a negative result
Appendix D
(Standard Appendix)
Quality Control Standard for Plague Indirect Hemagglutination Test
D1 Inspection Material Standard
The glassware used is cleaned, dried and sterilized, the measuring instrument is calibrated and qualified, the diagnostic Fi antigen, F sensitized blood cells, diluent pH 6.9-7.1), ion concentration 0.15mol/LNaCl and plague diagnostic serum are prepared by a unified method, qualified in quality inspection, and stored at 4°C. D2 Serum sample collection and storage standards
Pay attention to sterility when collecting blood and separating serum to prevent hemolysis and contamination. The serum should be no less than 0.5mL. The serum of the test subject does not need to be inactivated and absorbed. The serum should be used for the test as soon as possible. If it cannot be tested immediately, sodium azide or thimerosal (final concentration of the former is 0.1% and the latter is 0.01%) should be added or frozen for storage, but it cannot exceed half a month. D3 Preparation standard of quality control serum (internal control sample) Take 10μL plague diagnosis serum with a micro syringe and add 9.9mL homologous normal serum, mix thoroughly, then take 100μL accurately with a micro syringe, and inject each into An Yan, deep freeze or freeze vacuum dry for storage
D4 "True value" determination standard of quality control serum
Add 1mL diluent to the quality control serum An Yan, mix well, take 0.5mL and put it into the second tube for continuous multiple dilution, put the whole amount of An Shao into the first tube, which is regarded as 1:10, shake and mix well, add 1 drop of 2.5%Fi blood cell to each tube with a 1mL pipette, mix well at room temperature for 2-3 hours and observe the results. Test the quality control serum once a day for 20 consecutive days, and the titer with the highest frequency (at least 17 times) is regarded as the "true value". The batch is considered qualified when the ratio of the result to the "true value" titer does not exceed ±1 dilution. In actual work, one internal control sample is tested for each batch of test samples. D5 Participation in inter-laboratory quality control standards
Inter-laboratory quality control standards: The external control serum samples issued by the quality control reference laboratory are qualified if the titer is ±1 of the standard. The test samples in actual work should be treated the same as the control samples. When the tested serum shows a positive reaction of more than 1:20, it must be inactivated and absorbed, and three rows of retests must be performed to determine the titer. The plague hemagglutination positive reaction standard and diagnostic standard shall be implemented in accordance with the "Plague Serological Diagnostic Standards".5% stratified sampling survey method:
Visual method
Select 5 to 10 representative habitat plots in the monitoring area, each plot is 5 to 10ha. First, mark the area around the plot to determine the scope of the plot. Select a place 100 to 200m away from the plot, use a geographical barrier or shelter as cover, and use a telescope to observe the number of early otters in the plot during the most active time of the day. Observe for 2 consecutive days, twice a day. Calculate the density based on the highest value of one time.
Early otters density same ground distance (ha)
Early otters number
A2.3.2 Route method
（A2)
In the monitoring area, select 5 to 10 representative monitoring routes, each line is 10ha. The width of the route is 50-100m, the route distance is based on walking at 3km/h, riding at 5km/h, early climbing
Early climbing density = length of exposed line × width of standard field/1000mA3 Density survey of long-clawed gerbils and Brandt's voles. (A3)
A3.1 Survey time: Long-clawed gerbils and Brandt's voles are monitored once in spring and autumn. A3.2 Sampling area: Stratified sampling is carried out in the monitoring area at a ratio of 0.5% of the habitat area, and the sample plot is 1ha (100m×100m) as a unit. A3.3 Sample plot measurement and hole checking: Same as yellow rat (A1.3, A1.4). A3.4 All rat holes found during the hole checking with cloth clips are blocked. At 7-8 am the next day, a plate of cloth clips is placed on each hole entrance that was stolen, and the clamping time is 24h. Patrol once every 2 hours during the day, once at dusk and dawn, remove the rats caught during the patrol and re-set the rat traps at the original hole entrance, and continue to swat.
Rat density: calculated according to formula (A4).
Number of swatting
Rat density A group area (h)
A4 Survey of night-walking rat density in the wild
（A4）
A4.1 Number of traps (cages) surveyed: monthly, select 1 to 3 representative habitats, and set 100 traps (cages) per time for each habitat. The traps (cages) are set for 3 consecutive days each time in the source of large woolly plague, and for 2 consecutive days in the source of domestic plague. A4.2 Rat swatting tools: The models of traps (cages) should be unified A4.3 Bait: The board traps are uniformly made of white flour (vegetable oil) oil cakes, and the rat cages are made of fried dough sticks or sweet potatoes, each piece is 3 to 5 mg.
A4.4 Laying method: Use 5m clamp (cage) line method, the spacing between clamps (cages) should be more than 200m, and the direction of the clamp mouth and cage mouth should be consistent. A4.5 Laying time: Lay the clamps after sunset and retrieve them before sunrise. If there is heavy wind and rain during the time of laying the clamps and cages, the capture rate cannot be calculated. A4.6 Capture rate: Calculate according to formula (A5).
Number of rats caught
Capture rate (%) = (number of days × 100A5 Survey on density of domestic rodents
(A5)
A5.1 Number of surveys: In the monitoring area, select a representative village (village) every month, select 100 houses (including warehouses) in the village (village), and place a plate (piece) of traps (cages) in each house. A5.2 Rat-catching tools: the same as night-walking rats in the wild. A5.3 Bait: the same as night-walking rats in the wild.
A5.4 Time and method of placement: Continuously place for 48 to 72 hours/time, patrol once in the morning and evening every day, remove the captured rats, and replace the bait. A5.5 Capture rate: calculated according to formula (A6).
Catch* (%) = (number of days × 100
Number of rats caught
Appendix B||t t||(Standard Appendix)
B1 Survey Quantity
Survey Method for Rat Flea Index
During the monitoring period, 20 live main host animals are randomly selected for flea inspection every ten days in various plague natural foci.
B2 Survey Method
The captured live main host animals are individually bagged and anesthetized with ether in the flea inspection room. The fleas are combed with a comb or brush to obtain fleas for identification and classification. The fleas are grouped and sent for inspection according to the same host, the same direction, and the same species.
B3 Calculation
Total number of eggsbzxz.net
Rat total egg index No. 1
Total number of rats
Base flea number
Fruit egg index No. 1
Total number of rats
Limited egg infection rate (%)
Infected rats
Total number of mice
C1 Operation method
Appendix C
(B2)
(B3)
(Standard Appendix)
Indirect hemagglutination test method
Arrange the small test tubes on the test tube rack, mark the tube numbers, add 0.9mL of diluent to the first tube, and then add 0.5mL to each tube. Take 0.1mL of the test serum, add it to the first tube, mix it and dilute it in multiples, and take 0.5mL from the last tube and discard it. Then use a 1mL graduated pipette to add a drop of 2.5% F antigen-sensitized blood cells (0.05mL) to each tube, oscillate to mix the blood cells, place it in a 37℃ incubator or at room temperature for 2 to 3 hours to observe the results. For suspected materials, make 6 tubes, and each batch of tests should add 0.5mL of diluent plus a drop of 2.5% F antigen-sensitized blood cells as a control. C2 Result Judgment The positive result is defined as the uneven distribution of red blood cells at the bottom of the tube, with umbrella-shaped or irregular rough edges. The positive results are further divided according to the red blood cells at the bottom of the tube and the degree of roughness of the edges:
C2.1++++: Tight agglutination, agglutination covers the bottom of the tube, with obvious folded edges. When the antibody is excessive, it agglutinates into a sparse flower cluster. C2.2+++: Relatively tight agglutination, agglutination covers the bottom of the tube, in a disc shape without folded edges.
C2.3+++: Incomplete agglutination of blood cells, the bottom of the tube is a neat circle, and there is obvious agglutination of blood cells inside and outside the circle.
C2.4+: There are more non-agglutinated blood cells, which sink to the bottom of the tube and present a neat circle, and there are very few agglutinated blood cells outside the circle.
C2.5-: The blood cells do not agglutinate, sink to the bottom of the tube, and present a neat circle. The test tube method uses "10+10" as the positive limit of the indirect hemagglutination test. Red blood cells are densely packed in the shape of buttons at the bottom of the tube, with smooth and neat edges, or appear as a narrow circle, which is a negative result
Appendix D
(Standard Appendix)
Quality Control Standard for Plague Indirect Hemagglutination Test
D1 Inspection Material Standard
The glassware used is cleaned, dried and sterilized, the measuring instrument is calibrated and qualified, the diagnostic Fi antigen, F sensitized blood cells, diluent pH 6.9-7.1), ion concentration 0.15mol/LNaCl and plague diagnostic serum are prepared by a unified method, qualified in quality inspection, and stored at 4°C. D2 Serum sample collection and storage standards
Pay attention to sterility when collecting blood and separating serum to prevent hemolysis and contamination. The serum should be no less than 0.5mL. The serum of the test subject does not need to be inactivated and absorbed. The serum should be used for the test as soon as possible. If it cannot be tested immediately, sodium azide or thimerosal (final concentration of the former is 0.1% and the latter is 0.01%) should be added or frozen for storage, but it cannot exceed half a month. D3 Preparation standard of quality control serum (internal control sample) Take 10μL plague diagnosis serum with a micro syringe and add 9.9mL homologous normal serum, mix thoroughly, then take 100μL accurately with a micro syringe, and inject each into An Yan, deep freeze or freeze vacuum dry for storage
D4 "True value" determination standard of quality control serum
Add 1mL diluent to the quality control serum An Yan, mix well, take 0.5mL and put it into the second tube for continuous multiple dilution, put the whole amount of An Shao into the first tube, which is regarded as 1:10, shake and mix well, add 1 drop of 2.5%Fi blood cell to each tube with a 1mL pipette, mix well at room temperature for 2-3 hours and observe the results. Test the quality control serum once a day for 20 consecutive days, and the titer with the highest frequency (at least 17 times) is regarded as the "true value". The batch is considered qualified when the ratio of the result to the "true value" titer does not exceed ±1 dilution. In actual work, one internal control sample is tested for each batch of test samples. D5 Participation in inter-laboratory quality control standards
Inter-laboratory quality control standards: The external control serum samples issued by the quality control reference laboratory are qualified if the titer is ±1 of the standard. The test samples in actual work should be treated the same as the control samples. When the tested serum shows a positive reaction of more than 1:20, it must be inactivated and absorbed, and three rows of retests must be performed to determine the titer. The plague hemagglutination positive reaction standard and diagnostic standard shall be implemented in accordance with the "Plague Serological Diagnostic Standards".5% stratified sampling survey method:
Visual method
Select 5 to 10 representative habitat plots in the monitoring area, each plot is 5 to 10ha. First, mark the area around the plot to determine the scope of the plot. Select a place 100 to 200m away from the plot, use a geographical barrier or shelter as cover, and use a telescope to observe the number of early otters in the plot during the most active time of the day. Observe for 2 consecutive days, twice a day. Calculate the density based on the highest value of one time.
Early otters density same ground distance (ha)
Early otters number
A2.3.2 Route method
（A2)
In the monitoring area, select 5 to 10 representative monitoring routes, each line is 10ha. The width of the route is 50-100m, the route distance is based on walking at 3km/h, riding at 5km/h, early climbing
Early climbing density = length of exposed line × width of standard field/1000mA3 Density survey of long-clawed gerbils and Brandt's voles. (A3)
A3.1 Survey time: Long-clawed gerbils and Brandt's voles are monitored once in spring and autumn. A3.2 Sampling area: Stratified sampling is carried out in the monitoring area at a ratio of 0.5% of the habitat area, and the sample plot is 1ha (100m×100m) as a unit. A3.3 Sample plot measurement and hole checking: Same as yellow rat (A1.3, A1.4). A3.4 All rat holes found during the hole checking with cloth clips are blocked. At 7-8 am the next day, a plate of cloth clips is placed on each hole entrance that was stolen, and the clamping time is 24h. Patrol once every 2 hours during the day, once at dusk and dawn, remove the rats caught during the patrol and re-set the rat traps at the original hole entrance, and continue to swat.
Rat density: calculated according to formula (A4).
Number of swatting
Rat density A group area (h)
A4 Survey of night-walking rat density in the wild
（A4）
A4.1 Number of traps (cages) surveyed: monthly, select 1 to 3 representative habitats, and set 100 traps (cages) per time for each habitat. The traps (cages) are set for 3 consecutive days each time in the source of large woolly plague, and for 2 consecutive days in the source of domestic plague. A4.2 Rat swatting tools: The models of traps (cages) should be unified A4.3 Bait: The board traps are uniformly made of white flour (vegetable oil) oil cakes, and the rat cages are made of fried dough sticks or sweet potatoes, each piece is 3 to 5 mg.
A4.4 Laying method: Use 5m clamp (cage) line method, the spacing between clamps (cages) should be more than 200m, and the direction of the clamp mouth and cage mouth should be consistent. A4.5 Laying time: Lay the clamps after sunset and retrieve them before sunrise. If there is heavy wind and rain during the time of laying the clamps and cages, the capture rate cannot be calculated. A4.6 Capture rate: Calculate according to formula (A5).
Number of rats caught
Capture rate (%) = (number of days × 100A5 Survey on density of domestic rodents
(A5)
A5.1 Number of surveys: In the monitoring area, select a representative village (village) every month, select 100 houses (including warehouses) in the village (village), and place a plate (piece) of traps (cages) in each house. A5.2 Rat-catching tools: the same as night-walking rats in the wild. A5.3 Bait: the same as night-walking rats in the wild.
A5.4 Time and method of placement: Continuously place for 48 to 72 hours/time, patrol once in the morning and evening every day, remove the captured rats, and replace the bait. A5.5 Capture rate: calculated according to formula (A6).
Catch* (%) = (number of days × 100
Number of rats caught
Appendix B||t t||(Standard Appendix)
B1 Survey Quantity
Survey Method for Rat Flea Index
During the monitoring period, 20 live main host animals are randomly selected for flea inspection every ten days in various plague natural foci.
B2 Survey Method
The captured live main host animals are individually bagged and anesthetized with ether in the flea inspection room. The fleas are combed with a comb or brush to obtain fleas for identification and classification. The fleas are grouped and sent for inspection according to the same host, the same direction, and the same species.
B3 Calculation
Total number of eggs
Rat total egg index No. 1
Total number of rats
Base flea number
Fruit egg index No. 1
Total number of rats
Limited egg infection rate (%)
Infected rats
Total number of mice
C1 Operation method
Appendix C
(B2)
(B3)
(Standard Appendix)
Indirect hemagglutination test method
Arrange the small test tubes on the test tube rack, mark the tube numbers, add 0.9mL of diluent to the first tube, and then add 0.5mL to each tube. Take 0.1mL of the test serum, add it to the first tube, mix it and dilute it in multiples, and take 0.5mL from the last tube and discard it. Then use a 1mL graduated pipette to add a drop of 2.5% F antigen-sensitized blood cells (0.05mL) to each tube, oscillate to mix the blood cells, place it in a 37℃ incubator or at room temperature for 2 to 3 hours to observe the results. For suspected materials, make 6 tubes, and each batch of tests should add 0.5mL of diluent plus a drop of 2.5% F antigen-sensitized blood cells as a control. C2 Result Judgment The positive result is defined as the uneven distribution of red blood cells at the bottom of the tube, with umbrella-shaped or irregular rough edges. The positive results are further divided according to the red blood cells at the bottom of the tube and the degree of roughness of the edges:
C2.1++++: Tight agglutination, agglutination covers the bottom of the tube, with obvious folded edges. When the antibody is excessive, it agglutinates into a sparse flower cluster. C2.2+++: Relatively tight agglutination, agglutination covers the bottom of the tube, in a disc shape without folded edges.
C2.3+++: Incomplete agglutination of blood cells, the bottom of the tube is a neat circle, and there is obvious agglutination of blood cells inside and outside the circle.
C2.4+: There are more non-agglutinated blood cells, which sink to the bottom of the tube and present a neat circle, and there are very few agglutinated blood cells outside the circle.
C2.5-: The blood cells do not agglutinate, sink to the bottom of the tube, and present a neat circle. The test tube method uses "10+10" as the positive limit of the indirect hemagglutination test. Red blood cells are densely packed in the shape of buttons at the bottom of the tube, with smooth and neat edges, or appear as a narrow circle, which is a negative result
Appendix D
(Standard Appendix)
Quality Control Standard for Plague Indirect Hemagglutination Test
D1 Inspection Material Standard
The glassware used is cleaned, dried and sterilized, the measuring instrument is calibrated and qualified, the diagnostic Fi antigen, F sensitized blood cells, diluent pH 6.9-7.1), ion concentration 0.15mol/LNaCl and plague diagnostic serum are prepared by a unified method, qualified in quality inspection, and stored at 4°C. D2 Serum sample collection and storage standards
Pay attention to sterility when collecting blood and separating serum to prevent hemolysis and contamination. The serum should be no less than 0.5mL. The serum of the test subject does not need to be inactivated and absorbed. The serum should be used for the test as soon as possible. If it cannot be tested immediately, sodium azide or thimerosal (final concentration of the former is 0.1% and the latter is 0.01%) should be added or frozen for storage, but it cannot exceed half a month. D3 Preparation standard of quality control serum (internal control sample) Take 10μL plague diagnosis serum with a micro syringe and add 9.9mL homologous normal serum, mix thoroughly, then take 100μL accurately with a micro syringe, and inject each into An Yan, deep freeze or freeze vacuum dry for storage
D4 "True value" determination standard of quality control serum
Add 1mL diluent to the quality control serum An Yan, mix well, take 0.5mL and put it into the second tube for continuous multiple dilution, put the whole amount of An Shao into the first tube, which is regarded as 1:10, shake and mix well, add 1 drop of 2.5%Fi blood cell to each tube with a 1mL pipette, mix well at room temperature for 2-3 hours and observe the results. Test the quality control serum once a day for 20 consecutive days, and the titer with the highest frequency (at least 17 times) is regarded as the "true value". The batch is considered qualified when the ratio of the result to the "true value" titer does not exceed ±1 dilution. In actual work, one internal control sample is tested for each batch of test samples. D5 Participation in inter-laboratory quality control standards
Inter-laboratory quality control standards: The external control serum samples issued by the quality control reference laboratory are qualified if the titer is ±1 of the standard. The test samples in actual work should be treated the same as the control samples. When the tested serum shows a positive reaction of more than 1:20, it must be inactivated and absorbed, and three rows of retests must be performed to determine the titer. The plague hemagglutination positive reaction standard and diagnostic standard shall be implemented in accordance with the "Plague Serological Diagnostic Standards".5% stratified sampling, with 1ha (100m×100m) as the unit. A3.3 Sample measurement and hole checking: Same as yellow rat (A1.3, A1.4). A3.4 All rat holes found during the hole checking with traps shall be blocked. At 7-8 am the next day, a plate of traps shall be placed at each hole entrance that was stolen. The trapping time is 24 hours. Patrol once every 2 hours during the day, once at dusk and dawn. During the patrol, remove the rats caught and re-trap the rats at the original hole entrance, and continuously hit them.
Rat density: calculated according to formula (A4).
Number of traps
Rat densityArea of a group (h)
A4 Survey of night-walking rat density in the wild
（A4)
A4.1 Number of traps (cages) to be investigated: In units of months, select 1 to 3 representative habitats, and set up 100 traps (cages)/time in each habitat. In the source of large woolly plague, the traps (cages) are set up for 3 consecutive days each time, and in the source of domestic plague, the traps (cages) are set up for 2 consecutive days each time. A4.2 Rat trapping tools: The models of traps (cages) should be unifiedA4.3 Bait: The plate traps are uniformly made of white flour (vegetable oil) oil cakes, and the rat cages are made of fried dough sticks or sweet potatoes, each piece is 3 to 5 mg.
A4.4 Deployment method: Use the 5m trap (cage) line method for deployment, the spacing between the traps (cages) should exceed 200m, and the direction of the trap mouth and the cage mouth should be consistent. A4.5 Laying time: Lay the trap after sunset and take it back before sunrise. If there is heavy wind and rain during the time when the trap is in place, the capture rate cannot be calculated. A4.6 Capture rate: Calculate according to formula (A5).
Number of rats caught
Capture rate (%) = (number of days × 100A5 Survey on density of domestic rodents
(A5)
A5.1 Number of surveys: In the monitoring area, select a representative village (village) every month, select 100 houses (including warehouses) in the village (village), and place a plate (piece) of traps (cages) in each house. A5.2 Rat-catching tools: the same as night-walking rats in the wild. A5.3 Bait: the same as night-walking rats in the wild.
A5.4 Time and method of placement: Continuously place for 48 to 72 hours/time, patrol once in the morning and evening every day, remove the captured rats, and replace the bait. A5.5 Capture rate: calculated according to formula (A6).
Catch* (%) = (number of days × 100
Number of rats caught
Appendix B||t t||(Standard Appendix)
B1 Survey Quantity
Survey Method for Rat Flea Index
During the monitoring period, 20 live main host animals are randomly selected for flea inspection every ten days in various plague natural foci.
B2 Survey Method
The captured live main host animals are individually bagged and anesthetized with ether in the flea inspection room. The fleas are combed with a comb or brush to obtain fleas for identification and classification. The fleas are grouped and sent for inspection according to the same host, the same direction, and the same species.
B3 Calculation
Total number of eggs
Rat total egg index No. 1
Total number of rats
Base flea number
Fruit egg index No. 1
Total number of rats
Limited egg infection rate (%)
Infected rats
Total number of mice
C1 Operation method
Appendix C
(B2)
(B3)
(Standard Appendix)
Indirect hemagglutination test method
Arrange the small test tubes on the test tube rack, mark the tube numbers, add 0.9mL of diluent to the first tube, and then add 0.5mL to each tube. Take 0.1mL of the test serum, add it to the first tube, mix it and dilute it in multiples, and take 0.5mL from the last tube and discard it. Then use a 1mL graduated pipette to add a drop of 2.5% F antigen-sensitized blood cells (0.05mL) to each tube, oscillate to mix the blood cells, place it in a 37℃ incubator or at room temperature for 2 to 3 hours to observe the results. For suspected materials, make 6 tubes, and each batch of tests should add 0.5mL of diluent plus a drop of 2.5% F antigen-sensitized blood cells as a control. C2 Result Judgment The positive result is defined as the unev
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