1.1 This test method specifies the operational parameters required to grow and quantify a Bacillus subtilis biofilm comprised of vegetative cells and endospores (spores) using the colony biofilm method (CBM).2^,3 The resulting biofilm is representative of static environments that can develop a sporulating biofilm rather than being representative of one particular environment.

1.2 This test method utilizes a modified CBM to grow the biofilm. The CBM uses a semipermeable membrane on an agar plate as the biofilm growth surface and nutrient source.2^,3 In this test method, membranes are inoculated and incubated for a total of 8 days to promote sporulation within the biofilm.

1.3 This test method describes how to sample and analyze the biofilm for vegetative cells and spores. Biofilm population is expressed as total colony forming units (CFU) and spores per membrane.

1.4 Basic microbiology training is required to perform this test method.

1.5 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.