As we have previously reviewed (here), we now know how sterilizers work. The next logical question therefore is not only how do they work but what can they kill?

As we reviewed sterilization can occur via a wide variety of methods including physical, heat or chemical sterilization. Sterilizers by their very nature are designed to kill all forms of life. A more specific definition can be taken from the HICPAC 2008 report which indicates sterilization is a process that “destroys or eliminates all forms of microbial life and is carried out in health-care facilities by physical or chemical methods” (1).

Those organisms of interest in the healthcare setting include;

Fungi-the group of eukaryotic organisms that includes microorganisms such as yeasts and molds.

Bacteria-a large group of unicellular microorganisms that have cell walls but lack organelles and an organized nucleus.

Viruses (not necessarily alive but biological pathogens nonetheless)-small infectious agent that replicates only inside the living cells of other organisms.

Spore forms-single-celled asexual or sexual reproductive body that is highly resistant to desiccation and heat and is capable of growing into a new organism.

Unicellular eukaryotic organisms– include protozoans, slime molds and some forms of algae.

All of the above examples can be killed in a sterilizer.

How can we quantify sterilization?

There are many ways that we can estimate sterilization but the standard method to calculate if we can achieve sterility is the OVERKILL method. In this method we assume that some level of contamination is on the product prior to sterilization. If we also assume that a spore log reduction (SLR) value of 12 is required in order to achieve a sterility assurance level (SAL-typically a 10-3 or 10-6 level SAL meaning chances are 1 in 1000, or 1 in 1,000,000 that sterility was not achieved), then we can calculate the D value, which is the decimal reduction time i.e. the time in minutes to kill 90% of organisms being tested.

To calculate the D value we can then use the Stumbo/Murphy/Cochran equation that uses the formula D = U/ log No – log Nu ; where D is the D-value, U = exposure time,  No = population of BIs used, Nu = ln (n/r) where n = total number of BIs used and r = number of negative BIs after exposure time.

Once we have the D value we can then use the overkill method which ensures that for instance if the bioburden on an article were one million cells, and all of that bioburden consisted of resistant spores then a 12 log (i..e twice what is needed) reduction could occur in 24 minutes of exposure with a SAL of  10–6 probability of a nonsterile unit. Hence the overkill nature as the length of sterilization is twice what theoretically is needed.

How can we prove sterilization?

There are two methods in order to prove sterilization

  1. Biological Indicators (BI)-typically a spore suspension of Geobacillus or Bacillus species (typically B. atrophaeus or B. sterothermophilus)  that are thermotolerant. These indicators are autoclaved with the sterilized load, and then removed and incubated. If sterilization has occurred all the spores will have been killed and the spores will not germinate into viable cells. If sterilization is incomplete the spores will germinate and viable cells will then interact with the media of the BI leading to a visibile color change.
  2. Post sterilization testing-the United States Pharmacopeia a detailed set or compendium of drug information defines a set of testing to be conducted to prove that a labelled sterile product is in fact sterile. This guidelines is USP 71

Thanks for reading and in our next post we will look to the role of disinfectants. As always please add your thoughts and feedback in the comment section.

 

References cited in this article

1)HICPAC 2008 Report