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Isolators have been subjected to antimicrobial treatments to destroy environmental contamination since their introduction to the pharmaceutical industry in the 1980s. Initially, Peracetic acid/H2O2 was the sporicidal chemical used, but it was replaced by Vapor Phase Hydrogen Peroxide (VPHP) in the early 1990s. Currently, almost all isolators for product testing or production are decontaminated with VPHP. The majority of firms define success in decontamination as the complete kill of BIs with a population of 106 G. stearothermophilus spores. Occasionally, an unexpected BI positive occurs which is deemed a failure and this has been deemed the result of Rogue BIs.
In the early years after isolator introduction the sporicidal treatment of isolators was called sterilization.
Some regulators argued that sterilization of isolators was not a reasonable objective. Many surfaces inside an isolator are not easily treated with VPHP. Experience has shown that penetration of VPHP is limited. Although named decontamination, the process validation looks very much like sterilization.
It has become a standard expectation that biological indicators must always have a population of 106. This is most likely because inspectors or auditors perceived some kind of linkage between the SAL of 10-6 and a BI population of 106. Unfortunately, this idea is scientifically incorrect! It is not necessary to use a BI with a population of 10-6 to prove a SAL of 106. In fact neither 10-6 nor 106 have anything to do with the attribute of sterility. 106 spore population has even less significance in the decontamination of isolators.
Why SAL?
-6 10 as
a target
the risk based concept of SAL does not (or should not) matter.
To summarize this section 10-6 SAL is a measure of acceptable risk. 106 as a spore population on BIs has nothing to do with a 10-6 SAL . It has never been suggested that isolators SHOULD or MUST have a 10-6 SAL, which would be impossible to prove anyway!! Decontamination is not sterilization!! So, with these facts in mind why do we stipulate that BIs used in isolator decontamination must have a population of 106? Does the requirement for BIs with a population of 106 play a role in the so-called Rogue BI?
BIs are meant to challenge sterilization processes or to evaluate the efficacy of a sporicide.
BIs were never intended to provide the central design criterion for sterilization processes.
In sterilization process design the user should define how much lethality is necessary for their process BIs are biological evaluation that such lethality has been delivered. Unfortunately- in the case of VPHP there is no standard definition of lethality.
In steam sterilization we have defined BIER conditions and a standard measure of lethality the Fo. BIER=Biological Indicator Evaluation Resistometer.
The lethality value of Fo= 1 minute of heating at 121oC (which is the metric conversion of 250oF, the temp. of steam at 15 PSIG.)
The BIER standards allow us to define process D and Z values.
The D value is the time required to achieve a one spore log reduction at defined processing conditions.
Z value is a measure of rate of change of D value as kill conditions change (in moist heat this means temperature).
154.5 49 24.5
121
124
8.2
4.1
12.3
6.2
D is defined relative to process temperature i.e. D121. Process challenge is a product of D value x time. A process with a lethality of 12 minutes could be evaluated with: a BI with a population of 104 and a D121 of 3 minutes or a BI with a population of 106 and a D121 of 2 minutes. It both examples you would expect statistically 63% of the Bis to be positive. The challenge afforded by both of these BIs is exactly the same! Thus, both SHOULD be equally suitable for validation purposes! (reference USP <1211>) It is only regulatory and compliance convention that limits BI selection to those with a population of 106 scientifically 103 or 104 could for most processes work just as well.
of relative resistance.
D values are only roughly comparable only to other lots tested
There should be no observable change in D value with respect to increasing or decreasing spore population on a BI. This is true of BIs for dry heat and moist heat and is common to chemical and radiation sterilization as well.
Unusual BI performance
Is is common to use 30-100 BIs in VPHP decontamination
trails.
It is not uncommon to hear of reports of three such tests being
of a positive BI.
In many tests several BIs have been placed at a positive
G. stearothermophilus
105 population
Stainless Coupon SEM 8000X Typical Clump
106 BI populations.
This is probably a result of an over emphasis placed upon the
The typical bioburden is also much less resistant than G. stearothermophilus spores.
It is necessary to kill each organism only once! Extreme worst case conditions do not reduce risk and do not really accomplish enhanced product safety.
Summary
Super resistant BIs are not Rogues, the occur naturally for predictable reasons.
These super resistant BIs are not the result of a genetic phenomenon and have nothing to do with worst case. Super resistant BIs do not indicate high process or product risk. They are instead the result of over-aggressive and unnatural test conditions. There is nothing magical, scientifically special, or statistically meaningful about a 106 spore population. It is possible to use lower concentration BIs to reduce the likelihood of clumping. Decontamination is to eliminate bioburden which in cleaned isolators is very low.