November 25, 2014 — The proper use of “BIs” – or biological indicators – in the healthcare setting is crucial to assess the effectiveness of the sterilization process. Failure to monitor sterilization processes using appropriately labeled and FDA-cleared BIs:
- violates infection-control standards;
- precludes the requisite assurance that the monitored sterilization process was performing effectively as labeled;
- poses an increased risk of healthcare-associated infections, or “HAIs,” due to the clinical use of contaminated surgical instruments;
- may prevent the prompt identification of a malfunctioning sterilizer;
- may result in the hospital (or another type of healthcare facility) having to “recall” and quarantine certain potentially contaminated surgical instruments; and
- may result in the hospital being cited and not receiving re-accreditation as required for Medicare and Medicaid reimbursements.
Bacterial endospores
Biological indicators (BIs) contain a standardized number of resistant, viable bacterial endospores, typically on the order of 10^5 (or 100,000).
Using a BI to demonstrate and document that the monitored sterilization process destroyed this high number of bacterial endospores — indicated by the BI’s “negative,” or “no growth,” result —provides a level of assurance (but not a guarantee) that the requisite conditions for the sterilization of the processed instruments were achieved.
Geobacillus stearothermophilus, Bacillus atrophaeus
Because the resistance of different species of bacterial endospores varies and depends on the mode of sterilization (e.g., dry heat, pressurized steam, a gas or a plasma), no single species or type of bacterial endospore is universally used to monitor every type of sterilization process.
Rather, each mode of sterilization is challenged using the species of spore-forming bacterium that is most resistant to that monitored mode, thereby providing the greatest likelihood that the conditions for sterilization were achieved and patient safety ensured.
For instance, endospores of Geobacillus stearothermophilus are used to monitor steam sterilizers, whereas endospores of Bacillus atrophaeus are used to monitor sterilization processes that use ethylene oxide (EtO) gas.
Endospores of B. atrophaeus are not used to monitor steam sterilizers because endospores of G. stearothermophilus are more resistant to pressurized steam.
Frequency of monitoring
Healthcare facilities use BIs to monitor sterilization processes daily, weekly, or with each load of instruments, depending on the mode of sterilization and the type of surgical instrument.
For example, most guidelines recommend that a BI be used to monitor:
- a traditional steam sterilization process at least once a week;
- each cycle of an EtO gas sterilization process; and
- each load of implantable or intravascular devices.
(Guidelines also recommend, for example, that a sterilization process be monitored after its installation within the medical facility, or after a major repair.)
Ensuring that the healthcare facility is monitoring each mode of sterilization as frequently as guidelines require is a quality measure important to the prevention of HAIs.
Failure to monitor a sterilization process using an appropriate and legally marketed BI may result in the ‘processed’ instruments remaining contaminated. — Lawrence F Muscarella, PhD
Sterility assurance level (SAL)
Although the policies and procedures of most healthcare facilities recommend that several other types of indicators (e.g., mechanical and chemical indicators, or “CIs”) additionally be used to monitor sterilization processes, BIs are an irreplaceable cornerstone of every infection-control and quality-assurance program.
Unlike CIs — whose use and results are limited and may merely indicate that a set of surgical instruments was exposed to a sterilizing agent (not that the instruments themselves were successfully sterilized — BIs, on the other hand, uniquely confirm (by way of a “no growth,” or “negative” result) that the conditions sufficiently lethal to achieve sterilization (i.e., to destroy all of the pathogens that might have contaminated the processed surgical instrument) were produced, at least at the location within the sterilizer where the BI was placed.
As a result, proper placement of the BI at the specific location within the sterilizer validated by the sterilizer’s manufacturer to be the most difficult to achieve the conditions for sterilization is necessary for the BI to yield meaningful results. Refer to the three articles labeled Box A, Box B, and Box C at the end of this article.
Indeed, sterilization is an inferred state that is associated with a probability, or sterility assurance level (SAL), typically of 10^-6 (or, 1/1,000,000).
This very small number confers the probability that no more than one resistant endospore on one instrument (or on a BI’s spore strip) would survive exposure of one million (1,000,000) instruments, each contaminated with one million endospores, to the sterilization process.
The smaller the SAL’s value, the greater the likelihood that the instrument will not be contaminated after exposure to the sterilization process. — Lawrence F Muscarella PhD
Biological indicators are necessarily designed with a specific number of resistant endospores (e.g., 100,000) so that their complete destruction by the sterilization process demonstrates a SAL of 10^-6.
Quality Services for Hospitals, Manufacturers, Patients: Click here to read about Dr. Muscarella’s quality services committed to reducing the risk of healthcare-associated infections, including CRE outbreaks linked to contaminated endoscopes and other reusable medical equipment.
Designs of biological indicators
Biological indicators (BIs) feature a number of different designs, some rudimentary, some more novel and unique.
The “spore-strip” design is a type of traditional BI that features a strip of paper on to which are impregnated (at least 100,000) bacterial endospores. This spore strip is enclosed within a sealed glassine envelop to protect its integrity and maintain both its functionality and the viability of the enclosed endospores.
This type of BI requires that the spore strip be removed from its sealed envelop or packaging after its exposure to the sterilization process, followed by its aseptic transference to the indicated growth medium for incubation.
Any endospores that survived the sterilization process would germinate during incubation, yielding a “positive” result for the BI that demonstrated the sterilization process’s failure.
In the past the FDA had cleared a spore-strip type of BI for monitoring the effectiveness of at least one LCS (refer to FDA clearance: K960570). Unlike traditional BIs, this one required that the spore strip be removed from its sealed envelop before its exposure to the sterilant (i.e., the LCS).
A sidebar update: As of 2011, the FDA now distinguishes a “BI” from a “spore strip test,” classifying them as two distinct types of regulated medical devices (even though, as noted above, one type of a BI features a spore strip).
Indeed, the FDA in the past used the terms BIs and spore strips interchangeably, but no longer.
Whereas the FDA has designated BIs for use with bona fide (traditional) sterilizers, in 2011 the Agency granted a de novo classification to a spore strip test labeled exclusively for use as an optional accessory with one type of liquid chemical sterilization process (i.e., the STERIS System 1E liquid processing system).
This discussion about spore strip tests is continued in Dr. Muscarella’s related article entitled, “The STERIS System 1E Liquid Chemical Sterilant Processing System: Looking Back and Forward” — click here to read it.
A second and more versatile type of BI design is “self-contained” and features both an inoculated spore strip (or other type of carrier) and the growth medium, each of which is enclosed in a single sealed glass ampoule.
A third type of BI design is both “self-contained” and enzyme-based, providing results in as few as 3 to 4 hours.
This discussion about the designs of BIs is continued in Dr. Muscarella’s related article entitled “Types and Characteristics of Biological Indicators for Monitoring Sterilization Processes,” which may be read by clicking here.
Incubation temperatures, times
After its exposure to a completed sterilization process, the spore strip of the first two types of BIs described above is cultured and incubated in a growth medium at a temperature that promotes the germination (or, growth) of any (viable) endospores that might have survived due to ineffective sterilization.
Biological indicators that contain G. stearothermophilus, for example, are incubated at 55–60o C (for as long as 7 days).
For comparison, the recommended incubation temperature required to promote the growth of waterborne bacteria—namely, of gram-negative bacteria (e.g., Pseudomonas spp.) and atypical mycobacteria—is 37o C (for 1 to 2 days).
Discussion: Liquid chemical sterilants
Virtually every infection control guideline requires that processes labeled to achieve sterilization in the healthcare setting be periodically monitored using an appropriate and validated BI.
Failure to comply with these requirements can call into doubt the effectiveness of the sterilization process, raising the specter that surgical instruments used on patients might have remained contaminated during their reuse.
To be sure, improperly sterilized surgical instruments are a documented cause of HAIs, with associated morbidity and mortality.
Which brings into focus the FDA’s current position on the validity of using BIs to monitor LCSs.
Indeed, in 1996 the FDA had cleared a BI for monitoring an automated processor for surgical instruments that used a LCS to achieve the claim of sterilization, a regulatory decision that at the time was controversial.
Moreover, this specific automated processor, itself cleared by the FDA in 1988, featured a water filtration system to improve the quality of the water it used to rinse the processed instruments terminally.
Quality assurance standards would therefore have seemingly required that, in addition to monitoring the LCS using a BI, the quality of the rinse water be monitored microbiologically, especially since the automated processor’s rinse water was associated with the specific claim of being “sterile.”
Only through such monitoring could the actual microbial quality of the rinse water and the proper functioning of the water filtration system have been verified.
But that was then, and this is now.
[Note: Refer to the FDA clearances: K960570, K060568, and K062269. Also, read the related article “The STERIS System 1E Liquid Chemical Sterilant Processing System: Looking Back and Forward” by clicking here.]
Today, the FDA’s course has been reversed, having no longer cleared any BIs for monitoring LCSs (or, for monitoring the microbial quality of filtered rinse water; see: Box A).
A revised position, the FDA now requires that a new type of device termed a “spore strip test,” recently classified via the FDA’s de novo process, be used to evaluate the performance of at least one automated surgical instrument processor that uses a LCS and was cleared in 2010.
Comment: The FDA’s revised regulatory stance
It is not entirely clear why the FDA originally cleared the use of a BI to monitor a LCS-based processor (despite this BI having no legally marked predicate device as otherwise required).
But in 2008, a time when the FDA sanctioned the use of traditional BIs to monitor the effectiveness of at least one LCS, Dr. Muscarella recommended, based solely on scientific grounds, that the FDA:
- re-evaluate the soundness of its clearance of a BI labeled to monitor the effectiveness of a LCS (note that the FDA had cleared the first spore-strip type of BI for monitoring a LCS-based processor in 1996; refer to the FDA’s clearance: K960570 for more details);
- re-evaluate the soundness of its clearance of a water filtration system labeled to produce “sterile” water from a healthcare facility’s tap; and
- consider re-labeling LCSs (i.e., high-level disinfectants/sterilants) with the specific intent to clarify their limitations and to specify their outcome as being “100% sporicidal,” as opposed to achieving “sterilization,” the latter claim of which is potentially suspect and confusing even though it is currently featured on the labeling of almost every LCS marketed in the U.S., including 2% glutaraldehyde.
The FDA appears to have adopted Dr. Muscarella’s recommendations, at least the first of these two. No longer cleared or marketed in the U.S. are a BI labeled for monitoring the effectiveness of a LCS and a water filtration system labeled to produce sterile water from a tap.
Summary
The FDA no longer considers it scientifically sound to use a BI to monitor the effectiveness of LCSs (see: Box A).
Namely, although in 1988 and 1996, respectively, the FDA had cleared for marketing in the U.S. both: (i) a LCS-based processor labeled to “sterilize” surgical instruments and (ii) a BI labeled for monitoring this processor’s effectiveness, the FDA has since changed its stance: Today no such types of medical devices are cleared by the FDA for use in the U.S.
Conclusion
Biological indicators (BIs) are integral to every infection-control and quality-assurance program, and their proper application and use is crucial to prevent the use of contaminated surgical instruments during surgery.
Whereas BIs may be used to monitor traditional sterilization processes, spore strip tests — which the FDA is now distinguishing from BIs — are required instead to evaluate the effectiveness of at least one LCS-based automated processor.
Recommendations
A number of recommendations for the safe use of BIs are provided.
1. Do not use BIs to monitor the effectiveness of LCSs or automated processors that use a LCS (or to monitor the effectiveness of a water filtration system). Review Box A and Box B, below, for a continuation of this discussion.
- Instead, the FDA recommends that healthcare facilities use newly classified spore strip tests that are specifically labeled for monitoring a LCS-based automated processor. The FDA now differentiates spore strip tests from BIs.
- Review Dr. Muscarella’s related article “The STERIS System 1E Liquid Chemical Sterilant Processing System: Looking Back and Forward” by clicking here.
2. Review the BI’s (or spore strip test’s) labeling to assure its proper use.
- Monitor each mode of sterilization using a BI that contains the most resistant species of endospores to that mode.
- For instance, use spores of G. stearothermophilus to monitor traditional steam sterilizers.
- Ensure that each sterilization mode is monitored as frequently as guidelines recommend and as the sterilizer’s label indicates (e.g., at least once a week).
- Similarly, use chemical indicators (CIs) as frequently as guidelines recommend (e.g., on the outside of every load).
- Perform periodic audits to ensure that the healthcare facility is using BIs, CIs, or spore strip tests properly and as labeled.
3. Because infections associated with surgical implants can be insidious and result in significant morbidity, it is recommended that a BI be used to monitor each load of implantable devices.
- Immediate-use steam sterilization, or flashing, especially of implants is not recommended.
- Read to Dr. Muscarella’s related article “Flash or Immediate Use Steam Sterilization: A Position Statement” — click here.
4. Use a “positive-control,” as required by the BI’s labeling, with every “test”
BI.
- Incubate both the test BI and the positive-control BI at the recommended temperature (e.g., 55-60o C) and for the required number of days (e.g., 7 days).
- Confirm a “no growth” result for the test BI and that the positive control BI’s endospores have germinated.
- Also, confirm that the use- or shelf-life of both the test BI and the positive-control BI (including the growth medium) has not expired.
5. Place the BI at the location inside the sterilizer that is most difficult to sterilize—for example, the sterilizer’s “cold spot” or the center of the load.
- Refer to the sterilizer’s labeling to identify this specific location.
6. As part of a complete infection-control and quality-assurance program, it is recommended that the rinse water used by automated processors and labeled to be “sterile” (or “bacteria-free”) be microbiologically monitored, to evaluate whether the water filtration system’s 0.2 (or 0.1) micron bacterial membrane
is failing and allowing bacteria to pass, warranting replacement.
- Do not, however, use a BI or spore strip test for this purpose.
- Instead, contact the manufacturer of the automated processor to receive instructions detailing the proper techniques for sampling and monitoring the rinse water’s quality.
- Contaminatedrinsewaterusedtoreprocessflexibleendoscopes has been causally linked to patient morbidity and mortality.
- Assurances that the rinse water used during endoscope reprocessing is not contaminated with microorganisms is important to patient safety.
- It can be reasonable argued that the rinse water produced by an automated instrument processor requires routine microbiological monitoring, and that the failure to monitor this rinse water enjoins the conclusion that the wet, processed instruments may be contaminated with potentially pathogenic waterborne bacteria.
- Contaminatedrinsewaterusedtoreprocessflexibleendoscopes has been causally linked to patient morbidity and mortality.
- Consider applying to instrument processors and their rinse water the same principles and techniques used to monitor the rinse water used during the reprocessing of hemodialyzers.
References: https://lfm-hcs.com/htmlsite/2008/refs070808.pdf
Article by: Lawrence F Muscarella, PhD; posted: 11/25/2014, Rev C.
Box A: Reasons why BIs may not be used to monitor filtration systems labeled to produce sterile water in the healthcare setting.
- Removing the spore strip or carrier from the BI’s sealed packaging for immersion in water to assess the water’s “sterility” and the filtration system’s performance violates the well-established principles of the proper use of a BI.
- No legally marketed BI in the U.S. is labeled to monitor the effectiveness of a water filtration system or the microbial quality of an automated processor’s rinse water.
- Nor today is any BI cleared by the FDA for monitoring the effectiveness of a LCS (the clearance and use of one in past years notwithstanding). Instead, the FDA now designates the use of spore strip tests for this purpose. The FDA now distinguishes spore strip tests from BIs (see: main article; also refer to FDA clearances: K960570, K060568, and K062269; and refer to the related article “The STERIS System 1E Liquid Chemical Sterilant Processing System: Looking Back and Forward” by clicking here).
- Validation and verification data have not been published
demonstrating: (i) that a water filtration system can reliably and consistently produce “sterile” filtered rinse water from a healthcare facility’s tap water supply (during worst-case testing conditions as the FDA requires); (ii) the proper sampling techniques for on-site microbiological monitoring of “sterile” filtered rinse water; and (iii) the location within the automated processor (or water filtration system) where the production of “sterile” rinse water is most difficult to achieve and where monitoring the rinse water would be necessary, among other limitations and constraints. - The incubation temperature of 55-60oC—which is used to promote the growth of surviving endospores of G. stearothermophilus—inhibits, if not altogether prevents, the growth of waterborne microorganisms for which an incubation temperature of 37o C is recommended.
Box B: Problems associated with using a BI to monitor a liquid chemical sterilant (LCS).
- Removing the spore strip or carrier from the BI’s sealed packaging for immersion into a LCS: (i) is inconsistent with the well-established principles and traditional labeling of BIs and, therefore, likely prevents the BI from yielding valid and reproducible results; (ii) is inconsistent with the methodology of long-adopted standardized sporicidal tests (i.e., the AOAC sporicidal test); (iii) may compromise the BI’s integrity, resulting in the inactivation of some of the spore strip’s viable endospores during handling (or, in the environmental contamination of the spore strip); and (iv) can result in viable endospores being washed- or rinsed-off of the BI’s spore strip, yielding a “false-negative” BI result (see: Box C).
- According to the FDA, LCSs cannot be routinely monitored biologically.
- Validation and verification data demonstrating where within a processor the conditions for sterilization are most difficult to achieve and, therefore, where specifically to place the BI to monitor the LCS have not been published.
- Data demonstrating that G. stearothermophilus is the species of bacterial endospores most resistant to LCSs have not been published.
Box C. Reasons for not removing a BI from its sealed packaging.
BIs are uniquely designed so that the sterilizing agent can penetrate their sealed packaging and contact the enclosed viable endospores. This design is necessary for the BI to yield valid and reproducible results.
Removing a spore strip (or carrier) from the BI’s sealed packaging to monitor the effectiveness of a LCS (or, to monitor the microbial quality of filtered rinse water) would be inconsistent with the well-established principles of BIs and
could cause the BI to yield erroneous results (see: Box A, B).
Among other concerns, manually removing a spore strip from the BI’s packaging prior to the strip’s immersion in a LCS may result in the inadvertent inactivation (or removal) of some of the BI’s viable endospores—due to the handling of the BI’s spore strip, not effective sterilization—yielding a “false-negative” BI result and the potential for the release of improperly sterilized surgical instruments for patient use.
Similarly, removing a spore strip from the BI’s packaging for immersion in a LCS may result in the washing- or rinsing-off (or otherwise dislodging) of some of the BI’s viable endospores—a scenario that would prevent the recovery and incubation of surviving endospores, possibly also yielding a “false-negative” BI result.
Further, handling and manually removing a spore strip from the BI’s packaging may result in environmental contamination of the spore strip, providing yet another opportunity for a “false-negative” result.