Downloaded from https://www.everyspec.com .! ,, MIL-STD-81OC U. S. Department of Agriculture (SEA/FR) Northern Regional Research Center ARS Culture Collection 1815 N. University Street Peoria, Illinois 60604 (The fungi.may be distributed in a lyophilized state, or on agar slants), *American Type Culture collection 12301 Parklawn Drive Rockville, Maryland 20852 b. Maintain pure cultures of these such as potato dextrose agar except on strips of filter paper overlayed fungi separately on an appropriate medium that chaetomium globosum shall be cultured on the surface of mineral salts agar.. c.
Prepare mineral salts agar by dissolving 15.Og agar in a liter of the mineral salts solution described in 11.3.2.1.1. Jbt..e:Do not keep the stock cultures for more than 4 months at 6° z 4° C (43° 2 7° F); after that time, prepare subcultures and use them for the new stocks. d. Verify the purity of fungus cultures prior to the test. e. Incubate subcultures used for preparing new stock culttq’esor the spore suspension at 30° t 1.4° C (86° t 2.5° F) for 14 to 21 days. f. Prepare a s~re suspension of each of the five fungi by pouring into one subculture of each fungus 10 ml of an aqueous solution containing O.05g per liter of a nontoxic wetting Merit such as ,aodiumdioctyl sulfoauccinate or.
sodium lauryl sulfate. g. Use a rounded glass rod to gently scrape the surface growth from the culture of the test organiam. h. Pour the spore charge into a 125 ml capped Erlenme’yerflask containing 45 ml of water and 50 to 75 solid glass beads, 5 mm in diameter. i. Shake the flask vigorously to liberate the speres from the fruiting bodies and to break the spore clumps. .I. Filter the dispereed fungal spore suspension,into a flask through a 6 mm layer of glass wool contained in a glass funnel. w: This process should remove large mycelial fragments and clumps of agar. k. Centrifuge the filtered spore suspension and discard the supernatant liquid. 1.
Resuspend the residue in 50 ml of water and centrifuge. Wash the spores obtainsd from each of the fungi In this manner three times. m. Dilute the final washed residue with mineral-salts solution in such a manner that the resultant spore suspension shall contain 1,000,000 $ 200,000 spores per ml aa determined with a counting chamber. l@THOD 508.2 508.2-11 .-