Biological Indicators and their role in validation.

Biological Indicators: As per ISO, test system containing viable microorganisms providing a defined resistance to a specific sterilization process is called biological indicator (BI's). Biological indicators are bacterial spore suspension used to check efficacy of any sterilization process. There are different types of biological indicators with different strain of microorganisms for different sterilization process.

  

Self contained Biological Indicator

         Spore Strip

Spore Strip

A complete biological indicator pack includes:

(1) Spores 
(2) Carrier material 
(c) Primary pack 

Spores are used in biological indicators because spores are the most resistance form. By using spores suspension we challenge the particular sterilization process.Its very difficult to kill the spores as compare to vegetative cells. Spores contain very little amount of water and contains minerals like calcium, magnesium etc. That's why they are more resistant and stable. Carrier material is that material on or in which spores are present. There are different types of carrier material available like paper, liquid, stainless steel etc. Carrier material should be non reactive and should not have any impact on the spores during processing. Then primary packing material is there. The primary packing material holds the carrier so it is also very important. Primary packing material must allow the sterilizing agent to penetrate and come in contact with the spores. During validation of particular sterilization process two types of studies are performed. 

(A) Heat distribution study (B) Heat penetration study

In heat distribution study, we check the temperature uniformity within the sterilization chamber and define the hot and cold spot. In that study we only use chemical indicator. But in case of heat penetration study biological indicators are used. Heat penetration means biological indicators should be placed with in the load to check the efficacy of sterilization process. For example if we sterilize garments load then these indicators should be placed with in the garments load to check the efficacy and if liquid loads are used then biological indicators should be placed with in the liquid load. One more important thing is that for porous load like garments, glassware etc. we should use biological indicator with paper carrier and if we sterilize liquid load like media then self contained liquid biological indicators ampules should be used. So choose your biological indicators wisely depending upon the sterilizing load. There are different sterilization process  used in pharmaceutical industries and different strains of microorganisms used as BI's. Perform the verification of spore count before using the spores in sterilization process. Chart is given below for use of particular strain for particular sterilization process.

Name of Sterilization Process	Microorganism Used
Steam sterilization (121°C)	Geobacillus stearothermophilus (ATCC 7953)
Steam sterilization (<121°C)	Bacillus subtilis (ATCC 5230)
Dry heat sterilization	Bacillus atrophaeus (ATCC 9372)
Gas sterilization	Bacillus atrophaeus (ATCC 9372)
Ionizing radiations sterilization	Bacillus pumilus (ATCC 27142)
Vapour phase hydrogen peroxide (VPHP) sterilization	Geobacillus stearothermophilus (ATCC 7953)

After completion of heat penetration study, these BI's collected and incubated at particular temperature for particular time as per certificate. If no contamination found during or completion of incubation period then it means sterilization process is effective to kill the known concentration of spores. But if contamination found during or completion of incubation period then sterilization process must be reviewed to check any failure. Discard biological indicators properly after use.

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