Some standard content:
National Standard of the People's Republic of China
Quarantine Rules for Tabacco Seed (hlue Mold) Quarantine1 Subject Content and Scope of Application
This standard specifies the quarantine objects, procedures and methods for tobacco seeds. This standard applies to the quarantine of all kinds of imported tobacco seeds. 2 Terminology
2.1 Quarantine: a measure taken to prevent the spread of dangerous tobacco pests and diseases. 2.2 Tobacco Blue MaldGB 15699--1995
Peromospora tabacina Adam and Peronospora nicotianae Speg, synonym Peronospora hyosymi f. sptubacina, also known as blue poison disease, is a devastating disease on tobacco. It has now spread to Europe, America and Australia. At present, this disease has not occurred in my country, so it is used as an object of external quarantine.
3 Quarantine Procedure
3.1 Sampling
Sampling shall be carried out in proportion to the total number of seeds in a batch. When sampling, five diagonal points shall be sampled in a chessboard or stratified manner, and 10 to 20 samples shall be taken at each point (for a small amount of tobacco seeds carried by passengers and sent by international parcels, the number of samples shall depend on the specific situation). The sampling shall be representative as much as possible, and the average sample shall be evenly drawn from the samples taken at each point. 3.2 Inspection of Sporangia and Sporangia
3. 2. 1 Inspection Method
Washing inspection method.
3.2.2 Principle
Use the vibration method to disperse the conidia and conidia attached to the seeds into the water. 3.2.3 Equipment
Centrifuge, microscope, slide, vibrator, triangular flask. 3.2.4 Reagents
Distilled water.
3.2.5 Steps
Take 5g of tobacco seeds and put them into a conical flask. Add a certain amount of distilled water and soak it for 0.5h. Then add distilled water and shake vigorously for 5min (you can also put it on an oscillator and shake it). Pour the washing liquid into a centrifuge tube and centrifuge at 1 000 r/min for 5 min. Remove most of the supernatant and add a small amount of distilled water to each centrifuge tube to re-float the precipitate. Take the suspension for preparation and examine it under a 400-500 times microscope. Check 10 sections on each slide (see Figure 1). Check 5 to 10 sections on each sample to see if there are dendritic sporangial stalks and opposite sporangia. The spore size is about (13-18) μm×16 μm×29 μm.
State Administration of Technical Supervision 1995-08-28 Approved 1996-03-01 Implementation
GB 156991995
Figure 110 Distribution of sections on the slide
Conidiophores and conidia of tobacco frost poisonous pathogen See Figure 2. Figure 2 Conidiophores and conidia sections of tobacco frost pathogens 3.3 Oospore test
3.3.1 Test method
Lactophenol oil transparency method.
3.3.2 Principle
Lactophenol oil plus aniline blue is a standard floating medium for fungi. Aniline blue is an alkaline dye that can stain the protoplasm of the fungus but not the cell wall, so it can distinguish the fungal blisters that are difficult to see clearly. 3.3.3 Instruments and equipment
Microscope, beaker, electric stove, water bath. 3.3.4 Reagents
Aniline blue, phenol, glycerol, lactophenol oil, distilled water. 3.3.5 Steps
GB 156991995
Take 5g of the mixed sample tobacco seeds, place them in a beaker and boil them with lactophenol oil for 15~30min until the tobacco seeds are transparent but not yet digested. After staining with aniline blue (cotton blue), use lactophenol oil as a floating medium and then examine under a microscope. Check 20 slides for each sample (the formula of lactophenol oil is phenol crystals (heated and dissolved in a water bath) 20mL, lactic acid 20mL, glycerol 40mL, and distilled water 20mL). 3.3.6 Statistics of test results
When counting the number of spores, use a hemocytometer to observe 80 small grids, and use the average number × 4000000, which is the number of spores per milliliter of suspension.
3.4 Seed test
3.4.1 Test method
Shadow bud test.
3.4.2 Instruments and reagents
Autoclave, incubator, culture blood, filter paper, sodium hypocyanate, sterile water. 3.4.3 Steps
Take 100 seeds of the mixed sample and repeat 4 times. Take 4 sterile 10cm diameter burettes or plastic burettes, place each burette on three layers of absorbent paper soaked with sterile water, evenly arrange 100 seeds on the soaked paper, cover with burettes, and keep humidity. Then place in a 15~~20℃ incubator for culture, and observe the seedlings for signs of tobacco burette mildew infection after 4~7 days. If the seeds are to be checked for pathogens, immerse the seeds in 1% sodium chlorate solution for 10 minutes, then place on sterile filter paper and culture for inspection. 3.4.4 Statistics of inspection results
Single disease incidence and average incidence rate are calculated according to formula (1) and (2): Single disease incidence rate (%)
Number of diseased seeds
Total number of diseased seeds tested
Average incidence rate (%) = 1
3.5 Isolation culture test
3.5.1 Inspection method
Resin culture test.
3.5.2 Steps
Use 2.5 g of nutrient salts suitable for hydroponics, 14 g of agar, and 1 000 mL of distilled water. Use a 300 mL wide-mouthed triangular bottle, fill each bottle with 0.01 mL of culture medium, plug it with cotton plugs, and make the culture medium into a slope after disinfection. After the culture medium solidifies, use surface-sterilized seeds, add 0.01 g of each bottle on the culture medium slope and place it at 15~20℃. After the seeds germinate, observe whether the seedlings have signs of infection by tobacco downy mildew. 3.5.3 Result Statistics
Statistic the percentage of infected frost disease, the statistical method is the same as 3.4.4. 3.6 Isolation trial planting
3.6.1 Inspection
Before the introduction of varieties into the country, the introducer should submit the introduction approval form, the quarantine certificate of the exporting country, and the trade contract to the entry phytosanitary authorities for inspection (including seeds from informal channels). 3.6.2 Registration of materials for inspection
The materials for inspection shall be sent to the quarantine nursery by the introducer (unit) and registered by the quarantine recipient. 3.6.3 Preliminary inspection
The quarantine personnel shall conduct a preliminary inspection of the materials for inspection in the preliminary inspection room. 3.6.4 Trial planting in the quarantine nursery
The materials for inspection that have passed the preliminary inspection can be placed in the quarantine nursery and planted in the sterilized soil and pots in the quarantine greenhouse. 431
3.6.5 Evaluation of the results of trial planting in isolation
GB 156991995
The materials for inspection shall be inspected in the greenhouse during the growth period, and the inspection and testing results shall be registered in detail, and an evaluation shall be made based on the test results.
4 Evaluation and treatment after quarantine inspection
4.1 Tobacco seeds imported into the country shall be deemed qualified if no inspected objects are found after quarantine inspection. 4.1.1 Issuance of quarantine release notice (Appendix A (supplement)) If the quarantine is deemed qualified, a quarantine release notice will be issued, or a quarantine release stamp will be affixed on the freight bill. After strict packaging, it will be sent to Qingzhou Tobacco Research Institute of China National Tobacco Corporation for isolation, trial planting and quarantine. 4.1.2 Domestic trial planting
If the inspected object is not detected after isolation and trial planting, it will be handed over to the inspector (unit) for domestic trial planting. 4.2 Failure of quarantinebZxz.net
If quarantine objects are found after quarantine, it is considered unqualified. 4.2.1 Issuance of quarantine treatment notice (Appendix B (supplement)) If quarantine is unqualified, the following treatment should be carried out and a quarantine treatment notice will be issued. 4.2.2 Destruction
If quarantine objects are found in the inspected tobacco varieties, and the quarantine objects are within 1% of the sampled tobacco varieties, they should be destroyed on the spot at the quarantine site.
4.2.3 Return to the original place
The quarantined objects among the inspected tobacco varieties can be returned to the original place under the following conditions: the materials are of great research and application value, and the quarantined objects of the inspected tobacco varieties shall not exceed 1%.
4.2.4. Isolate and test plant after treatment
For a small amount of valuable materials introduced, heat treatment can be used, and test plant in the quarantine greenhouse after treatment. Dry heat treatment method: Dry heat disinfection, ventilate the seeds at 60℃ for 2-3h in advance to make the seeds fully dry. Spread the seeds into a thickness of less than 2cm, and use a dry heat sterilizer at 70℃ for more than 2 hours. The treated seeds are tested in the quarantine greenhouse with sterile soil and pots to check whether there are quarantine elements. The propagated seeds (should be sterile) are handed over to the inspector (unit) and can be tested domestically. 5. Issuance of inspection certificate
When no quarantine object is found in the imported tobacco seeds, a plant quarantine certificate (Appendix C (Supplement)) is issued: When the quarantine does not meet the provisions of the trade contract, an inspection certificate [Appendix knife supplement) is issued, explaining the situation as a document for external claims, and preparing and preserving the specimens of the inspected tobacco seeds and the quarantine object as the basis for compensation. 6. Quarantine report
Includes the inspection date, batch number, quantity and weight. Quarantine result:
Handling opinion:
Quarantine officer:
Quarantine unit:
Inspection unit
Product name and quantity
Bill of lading/waybill number
Name and number of means of transport
CB 15699--1995
Quarantine release notice
(supplementary document)
Animal and Plant Quarantine Institute of the People's Republic of China
Release notice
Country/region of export
The above goods have passed the quarantine
No. of XX Animal and Plant Quarantine Institute of the People's Republic of China:
Marking mark
Inspection unit
Product name and quantity
Bill of lading/waybill number
Name and number of means of transport Number
GB156991995
Appendix B
Notification of quarantine treatment
(Supplement)
Animal and Plant Quarantine of the People's Republic of China
Notification of treatment
Exporting country/region
Quarantine situation and treatment requirements:
Animal and Plant Quarantine Institute of the People's Republic of China 434
No.:
Mark code
Issuer and address
GB 15699 - 1995
Appendix C
PHYTOSANITARY CERTIFICATE
MINISTRY OF AGRICULTURE OF PR CHINAName & Address of Cnnsignor
Namc &. Address of ConsigneeName & Quantity DeclaredPlace of Origin
Place uf Chrigin
Port uf Desparch
Date of Arrival
Tools for Pre-shipment
Meats f Canyeyance
Date of Completion of Discharg No
Mark & Number
This is to certify that the consignment described above was examined on by this service and the following disexsesandfor insect ests andfor weed seeds were Fourd:This batch of goods does not comply with the provisions of the trade contract/contract No.agrged between the contract Date of Issue
Name of Authorized Officer Signature
The Animal and Plant Quarantine Administration of the People's Republic of China and its officials or representatives do not assume any responsibility for issuing this certificate. The finencial lizbility will not be able to attend to this enimel and plant querantine service of PR China to any officer or representative of the service with tespect to this certificate.435
People's Republic of China
XX Animal and Plant Quarantine Service
GB 15699—1995
Appendix D
Inspection Certificate
(Supplement)
(.ANIMAL AND PLANT QUARAN-TINE SERVICE THE PEOPLE'S REPUBLIC OF CHINA)
Address:
(ADDRESS:
Registered Telegraph:
(CABLE:
Consignor
Consignee
(Commodity)
Weight &/or Quantity Declared
(Port or Station of Despatch)
(Means of Conveyance)
Mark & Number:
INSPECTION
CERTIFICATE)
(Mark & Number)
Date of Arrival (Date of Campletion of Discharge) This lot of commodity was examined on this service and the following diseases and/orinsect pests and/or weed seeds were found: This lot of commodity doesn't comply with the requirements stipulated in the contract No. signed between the buyer and the seller. (Inspector) NB In case of difference, the Chinese text shall be regarded as authentic,)436
Additional Notes:
This standard is proposed by the State Tobacco Monopoly Administration. CB156991995
This standard is under the jurisdiction of the National Tobacco Standardization Technical Committee: This standard was drafted by the Qingzhou Tobacco Research Institute of China National Tobacco Corporation, and the main drafters of this standard were Zhong Daoru, Shi Jinkai, Zhu En, Ai Shuli, and Liu Hongxiang. 437
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