3.2.1 Lauryl Sulfate Tryptose (LST) broth

3.2.2 Brilliant Green Lactose 2% Bile (BGLB) broth

3.4.1 The medium used is LST broth, dispensed in 10 ml volumes into tubes containing gas vials (inverted Durham tubes).

3.4.2 Arrange LST broth tubes in rows of five, and mark them identifying the sample, the sample unit and the dilution to be inoculated.

3.4.3 Inoculate each of five tubes of double strength LST broth with 10 ml of the undiluted sample unit, and inoculate each of five tubes of single strength LST broth with 1 ml of the undiluted sample unit, and inoculate each of five tubes of LST broth with 0.1 ml of each undiluted sample unit.

3.4.4 Mix inoculum and medium by gently shaking or rotating the tubes, but avoid entrapping air in the gas vials.

3.4.5 Incubate the inoculated LST broth tubes at 35°C ± 0.5^o for 24 ± 2 hr. Examine for gas formation, record results, and on the same day begin the confirmed test for all gas-positive tubes (see section 3.5, below).

3.4.6 Incubate gas-negative tubes for an additional 24 ± 2 hr, examine, record the number of gas-positive tubes, add to the result obtained in step 3.4.5, above, and begin the confirmed test for the additional gas-positive tubes.

3.4.7 The absence of gas in all of the tubes at the end of 48 ± 2 hr of incubation constitutes a negative presumptive test.

3.4.8 Compute the "MPN" of presumptive coliforms per 100 ml of mineral water following the instructions in Part 5, to convert the number of gas-positive tubes to MPN values. Record results.

3.5.1 The confirmatory medium used is BGLB broth dispensed in 10 ml volumes in tubes containing gas vials.

3.5.2 Submit all gas-positive LST broth tubes to the confirmed test.

3.5.3 Shake or rotate the LST broth tubes to mix the contents and transfer one loopful from each positive LST broth tube to a tube of the BGLB broth. (Avoid transferring pellicle).