AOAC 966.04

Sporicidal Activity of Disinfectants

Summary and Test Overview

AOAC 966.04 Sporicidal Activity of Disinfectants is the test method of choice for registering sporicidal claims. 

Endospore-forming microorganisms are known for their tough nature, being able to survive on surfaces for extended periods of time, as well as being unaffected by many standard commercial disinfectants. They are also known for their proclivity in causing illness and diseases amongst humans. Therefore, products registered with sporicidal claims are necessary in situations, such as a healthcare setting, where it is important to combat the spread of these spore-forming bacteria. 

In this test method, the sporicidal efficacy of liquid agents is evaluated against carriers inoculated with Bacillus subtilis and Clostridium sporogenes. The preparation of the spore suspensions and the drying of the carriers are carefully done as to achieve 10⁵ – 10⁶ Colony Forming Units (CFU) per carrier. 

During the test, carriers are submerged in the liquid test substance for a pre-determined contact time. After completion of the contact time, carriers are then transferred (also known as subcultured) into a liquid recovery broth containing neutralizers, to ensure the complete cessation of sporicidal activity. After completion, the test is then incubated for a 21-day period, after which the recovery broth is qualitatively examined for evidence of growth of the challenge microorganisms.