(a) Absence of gas-formation.
(b) Formation of gas occupying less than ten per cent. (10%) of the closed arm.
(c) Formation of gas occupying more than ten per cent. (10%) of the closed arm.
More detailed records of the amount of gas formed, though desirable for purposes of study, are not necessary for carrying out the standard tests prescribed.
3. The formation within 24 hours of gas occupying more than ten per cent. (10%) of the closed arm of fermentation tube constitutes a positive presumptive test.
4. If no gas is formed in 24 hours, or if the gas formed is less than ten per cent. (10%), the incubation shall be continued to 48 hours. The presence of gas in any amount in such a tube at 48 hours constitutes a doubtful test, which in all cases requires confirmation.
5. The absence of gas formation after 48 hours’ incubation constitutes a negative test. (An arbitrary limit of 48 hours’ observation doubtless excludes from consideration occasional members of the B. coli group which form gas very slowly, but for the purposes of a standard test the exclusion of these occasional slow gas-forming organisms is considered immaterial.)
B. PARTIALLY CONFIRMED TEST.
1. Make one or more Endo’s medium or lactose-litmus-agar plates from the tube which, after 48 hours’ incubation, shows gas formation from the smallest amount of water tested. (For example, if the water has been tested in amounts of 10 cc., 1 cc., and 0.1 cc., and gas is formed in 10 cc., and 1 cc., not in 0.1 cc., the test need be confirmed only in the 1 cc. amount.)
2. Incubate the plates at 37° C., 18 to 24 hours.