AOAC 965.12

Tuberculocidal Activity of Disinfectants

Summary

The AOAC 965.12 Tuberculocidal Activity of Disinfectants test is a specialized version of the Use-Dilution Method (AOAC 955.14, 955.15, 964.02) to measure the tuberculocidal activity of disinfectants.

Tuberculosis (TB), caused by the organism Mycobacterium tuberculosis, is a disease which can infect the lungs and has the potential to be fatal. Therefore, it is highly important for facilities, households, and healthcare settings to protect their occupants against TB. 

Mycobacterium bovis, a related pathogen which causes tuberculosis disease in cows and other farm animals, is used in the Bacillus Calmette-Guérin (BCG) vaccine. The U.S. Environmental Protection Agency (EPA) considers Mycobacterium bovis as an alternative surrogate challenge organism for Mycobacterium tuberculosis. 

Test Overview

Modifications to the Use-Dilution Methods (AOAC 955.14, 955.15, 964.02)

• Culture from growth media slants is transferred to tissue grinders and homogenized to produce the test inoculum.
• Porcelain Penicylinders (carriers) are contaminated with the inoculum (15-20 mL inoculum/10 carriers) and remain in contact with the inoculum for 15 minutes.
• Carriers are placed in sterile Petri dishes matted with filter paper and dried.
• Carriers are submerged in the test substance and transferred to neutralizer with flame-sterilized wire hooks after the contact time.
• Secondary subculturing procedures to growth media of Proskauer Beck Broth (MPB), 7H9, and Kirchner’s Medium (KM).
• Products must achieve complete sanitization (no positive replicates out of ten total) to be registered as a tuberculocidal with the EPA.
• One carrier count replicate immediately before starting the efficacy test and two carrier count replicates immediately after the conclusion of the efficacy test.
• Additional controls including performance assessments of media.
• Incubation period of 60 days (if not passing) or 90 days (if passing). Confirmation streaks incubate for an additional 17-21 days (total incubation up to 111 days).  The long incubation time is due to the extended doubling time and growth of the organism.

The AOAC 965.12 Tuberculocidal Activity of Disinfectants test can be challenging to pass because the organism is difficult to kill, has special growth requirements, and has a prolonged incubation period.