SANS 5484-2009 Resistance to fungal attack by mixed fungi《混合霉的抗真菌性》.pdf

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3、damage whatsoever than may result from the use of this material or the information contain therein, irrespective of the cause and quantum thereof. ISBN 978-0-626-22930-6 SANS 5484:2009Edition 3SOUTH AFRICAN NATIONAL STANDARD Resistance to fungal attack by mixed fungi Published by SABS Standards Divi

4、sion 1 Dr Lategan Road Groenkloof Private Bag X191 Pretoria 0001Tel: +27 12 428 7911 Fax: +27 12 344 1568 www.sabs.co.za SABS SANS 5484:2009 Edition 3 Table of changes Change No. Date Scope Foreword This South African standard was approved by National Committee SABS SC 1028G, Pesticides Resistance t

5、o fungal attack, in accordance with procedures of the SABS Standards Division, in compliance with annex 3 of the WTO/TBT agreement. This document was published in June 2009. This document supersedes SABS SM 484:1981 (first revision). SANS 5484:2009 Edition 3 1 Contents Page Foreword 1 Scope 3 2 Test

6、 fungi and culture medium 3 3 Maintenance of the test fungi 3 4 Apparatus and materials . 4 5 Preparation of test specimens 4 6 Procedure 4 7 Evaluation 5 SANS 5484:2009 Edition 3 2 This page is intentionally left blank SANS 5484:2009 Edition 3 3 Resistance to fungal attack by mixed fungi 1 Scope Th

7、is standard specifies a method for the determination of the resistance to fungal attack by mixed fungi. 2 Test fungi and culture medium 2.1 Test fungi Aspergillus niger, ATCC 16404, SABS TCC 355. Aureobasidium pullulans, ATCC 9348, SABS TCC 402. Chaetomium globosum, ATCC 16021, SABS TCC 15 or ATCC 3

8、6703, SABS TCC 381. Penicillium brevicompactum, ATCC 9056, SABS TCC 389. Trichoderma sp., ATCC 0043, SABS TCC 406. 2.2 Culture medium 2.2.1 Dissolve the following in enough distilled water to yield 1 L of the solution: Agar 15 g Ammonium nitrate 1,0 g Mono-potassium phosphate . 0,7 g Di-potassium ph

9、osphate . 0,7 g Magnesium sulfate, heptahydrate 0,7 g Sodium chloride 0,005 g Ferrous sulfate, heptahydrate 0,002 g Zinc sulfate, heptahydrate 0,002 g Manganous sulfate, heptahydrate 0,001 g 2.2.2 Add enough 0,1 M sodium hydroxide solution to ensure that after sterilization the pH value of the mediu

10、m is 6,3 0,2 at 25 C. 2.2.3 Sterilize the culture medium in bulk by autoclaving at a temperature of 121 C 2 C for 20 min. 3 Maintenance of the test fungi Subculture the test fungi at intervals of 14 d on strips of filter paper placed on slants of the culture medium in suitable tubes. SANS 5484:2009

11、Edition 3 4 4 Apparatus and materials 4.1 Distilled water, that has been distilled in an all-glass apparatus or de-mineralized water of equivalent purity. 4.2 Sterile distilled water. 4.3 Slant cultures of the test fungi. 4.4 Sterile inoculating wire (platinum or nichrome). 4.5 Sterile flasks. 4.6 P

12、etri dishes, of 150 mm in diameter. 4.7 Strips of sterile filter paper, approximately 10 mm 25 mm in size. 4.8 Sterile forceps. 4.9 Sterile pipettes, of capacity 10 mL and 1 mL (graduated in 0,1 mL divisions). 4.10 Wetting agent solution, 0,5 g/L solution in water of a non-ionic wetting agent of whi

13、ch a 0,1 g/L aqueous solution is non-toxic to the test fungus. 5 Preparation of test specimens 5.1 When relevant, subject the test specimens to the appropriate pre-treatment specified in the relevant standard for the required bacteria. 5.2 The quantity and dimensions of the test specimens shall be a

14、s specified in the relevant standard for the required bacteria. 6 Procedure 6.1 Treatment of test specimens 6.1.1 Place the test specimens in a suitable container and add distilled water that contains 0,05 % of the non-ionic wetting agent until they are completely covered. 6.1.2 Keep the specimens s

15、ubmerged for 1 h and then pour off the water. 6.2 Mixed fungus spore suspension 6.2.1 Prepare a spore suspension of each of the five fungi by adding 10 mL of sterile distilled water that contains 0,05 % of the non-ionic wetting agent to one slant culture of each fungus. Using a sterile inoculating w

16、ire, gently scrape the surface growth from the culture of the test fungus. 6.2.2 Filter the spore suspension through sterile coarse cotton gauze into a sterile flask that contains 50 mL of sterile distilled water, and shake the flask. Repeat this operation for each of the test fungi, filtering into

17、the same sterile flask. SANS 5484:2009 Edition 3 5 6.3 Inoculation 6.3.1 Small specimens 6.3.1.1 Using the sterile forceps, aseptically transfer each small test specimen (i.e. of size small enough to be accommodated in a Petri dish) to a Petri dish that contains 25 mL of sterile culture medium. Ensu

18、re that one side of each specimen is in firm contact with the culture medium. 6.3.1.2 If tensile strength tests have to be carried out subsequently, ensure that each end of the test specimen is bent up vertically (away from the agar surface) for a distance of at least 25 mm. Place a strip of sterile

19、 filter paper on the culture medium but not in contact with the test specimen. 6.3.1.3 Spray the mixed fungus spore suspension on to the surface of each test specimen and on to the surface of each strip of filter paper to the point of run-off. Ensure that the spore suspension is evenly distributed o

20、ver the surface of each test specimen. NOTE Frequent sterilization of the spraying area is necessary, and any acceptable method may be used. 6.3.2 Other specimens 6.3.2.1 Aseptically transfer each test specimen to a sterile covered glass jar that contains sterile distilled water and a sterile stand

21、that supports the specimen above the surface of the water. 6.3.2.2 Spray the mixed fungus spore suspension on to the surface(s) of each test specimen to the point of run-off, ensuring that the spore suspension is evenly distributed over the surface(s). NOTE Frequent sterilization of the spraying are

22、a is necessary, and any acceptable method may be used. 6.4 Incubation Incubate the containers at a temperature of 28 C 2 C and a relative humidity of 90 % 5 % for a period of at least 21 d. Record the growth each week. 6.5 Visual examination 6.5.1 After incubation, note whether the surface of each f

23、ilter paper strip in the Petri dishes is covered with the fruiting bodies of the fungi. Remove the test specimens from the Petri dishes and examine them for fungal growth. Where necessary, confirm fungal growth by microscopical examination. 6.5.2 In cases where fungal growth is not evident or where

24、there are signs of contamination, discard the specimens and repeat the test on a new set of test specimens. 7 Evaluation Report the extent of fungal growth on the control filter paper and on the test specimens as determined by visual examination (see 6.5). SABS This page has been left blank intentio

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