The Science of Speed: How Fast Biological Indicators have ......ISO 11138 Series 11138 –2: Ethylene oxide 11138 –3: Moist heat (steam) 11138 –4: Dry heat 11138 -5: Low temperature

3M Health Care AcademySM

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The Science of Speed:How Fast Biological Indicators have Changed Quality Control in Reprocessing

Janet PrustDirector – Global Standards


© 3M 2017. All Rights Reserved 2. All Rights Reserved.25 September 2019© 3M 3M Confidential.

Devil’s CascadeRainy Lake

Northwestern Ontario



Hexagenia Mayfly



Common Loon



Tullibee Cisco



Freshwater Mussels - Ontario


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What do these four species have in common?

Bioindicators (indicator species) • Species or a group of species whose function, population or status can reveal

the qualitative status of the environment in which it lives.

What environment are they bioindicators for? • Clean water. Clean water indicators are sensitive to pollution and are

a good indicator of water quality changes.



Viability is reflective of it’s environment

Common Biological Indicator



Learning Objectives

1. Review how biological indicators work

2. Understand how rapid readout biological indicators work

3. Discuss the biological indicators role in sterilization quality control.



Director – Global Standards

3M Health Care Employee

Medical Solutions Division

Janet Prust - Disclosure




© 3M 2017. All Rights Reserved© 3M 2016. All Rights Reserved3M Health Care AcademySM

12

Biological Indicator Basics



Biological Indicator – Definition:

A test system containing viable microorganisms providing a defined resistance to a specified sterilization process.

ISO/TS 11139: 2018. Sterilization of health care products — Vocabulary. International Organization for Standardization.



Types of Biological Indicators

Spore stripsInoculated carriers Liquid BIs



Self-contained Biological Indicators

Cap

Filter

Growth Media

with pH Indicator

Spore Strip Sleeve



The Microbiology of Biological Indicators



Bacterial Spores

Some types of bacteria can form spores to protect themselves from a hostile environment, such as lack of nutrients

• e.g. some Bacillus sp., Clostridium sp.

• Like nuts or seeds - no biological activity - dormant

Very tough and hard to kill – challenges the sterilization process



Bacterial Spores

Geobacillus stearothermophilus Bacillus atropheus

Too small to see . . . about 1-2 micrometers (.000001 meters)



Growth Media

• Mixture of sugars and other nutrients

• Formula optimized for the biological indicator microorganisms



Bacterial Spores – Activation and Germination

Google Images

A spore “sprouts”A spore “sprouts”Like a seed sprouts . . .



Bacterial cell

Spore

Biochemical and physical transformation steps

Bacterial Spores – Activation and Germination



Bacterial Growth

Bacterial cell



23

Biological Indicator Standards



ISO 11138 Series

11138 – 2: Ethylene oxide

11138 – 3: Moist heat (steam)

11138 – 4: Dry heat

11138 - 5: Low temperature steam formaldehyde

CAN/CSA-ISO 11138-1:17 Scope

This document specifies general requirements for production, labelling, test methods and performance characteristics of biological indicators, including inoculated carriers and suspensions, and their components, to be used in the validation and routine monitoring of sterilization processes.

This document specifies basic and common requirements that are applicable to all parts of ISO 11138. Requirements for biological indicators for particular specified processes are provided in the relevant parts of ISO 11138. If no specific subsequent part is provided, this document applies.



Biological Indicator Performance Requirements



Key BI Performance Parameters

Population (number of test organisms)• Number of organisms that must be killed to get a negative BI• The more organisms, the longer it takes to inactive the BI• Measured by distributing the spores on plates and counting• Expressed in exponents (e.g. 106 = 1,000,000)

D-value (resistance)• A measure of how resistant the organisms are to the process

(how hard to kill)• Measured as the time to reduce the population by 90%• Expressed in time units (usually minutes)



27

Biological Indicator Incubation Time



Biological Indicator IncubationProcess for providing the spores in the biological indicator with the perfect conditions to support the growth of the organisms

- Nutrients (food)- Optimal temperature- Optimal pH

The incubation conditions are designed to be able to recover a very small number of organisms that remain alive in the biological indicator after exposure to the sterilization process, if there is a sterilization process failure.



Biological Indicator Incubation

These biological processes happen during incubation . . .



Biological indicator incubation time is the time required to make a final determination that the BI is negative• If the biological indicator turns positive at any point, the test is

complete and appropriate action must be taken.





“An incubation period is commonly recognized to be 7 days for established sterilization processes . . .”

ISO 11138 (2017) Sterilization of health care products – Biological indicators –Part 1: General requirements



Reduced Incubation Time (RIT)

Reduced Incubation Time is any established biological indicator incubation time that is less than 7 days• For example: 48 hours, 24 hours, 3 hours, 1 hour or 24 minutes

• The biological indicator manufacturer compares (validates) the BI response at the RIT to the BI response after 7 days, against established criteria.

How is this validation accomplished?



RIT Test Methods

Pharmacopeias- No methodology defined in USP, EP

ISO Standards- No specific methodology defined in ISO 11138 series standards- No specific methodology defined in ISO 14161- Standard in Draft – ISO CD 16342

U.S. Food and Drug Administration (FDA)- Method defined in FDA 510k Guidance document



Fractional cycles and MPN

Page 6



Fractional cycles and MPN



U.S. FDA RIT VALIDATION

http://www.fda.gov/medicaldevices/deviceregulationandguidance/guidancedocuments/ucm071261.htm

Reduced incubation time validation is part of the legal requirements for biological

indicators used in hospitals in the United States.



http://www.fda.gov/medicaldevices/deviceregulationandguidance/guidancedocuments/ucm071261.htm#a2

U.S. FDA –RIT Methodology

30+’s → MPN = 0.357• Range 0-3 spores/BI

80+’s → MPN = 1.609• Range 0-7 spores/BI

AAMI WG 4 Presentation to U.S. FDA; May 26, 2016



Positive Biological Indicators• Any spores that survive the sterilization cycle (a sterilization cycle failure)

are exposed to the growth media, then germinate (change from spores to cells), and start to grow:

• As they germinate and grow, they create many chemical reactions, that can be detected as a “signal”, that the biological processes are underway.



Positive Biological Indicator “signals”

TurbidityBacterial cells scatter light

Media Color ChangepH shift caused by metabolic by-products



40

3M Rapid Readout Technology



3M Rapid Readout Technology

• Introduced in 1991

• Now used to monitor 3 different sterilization technologies (steam, ethylene oxide, hydrogen peroxide)

• Electronic detection of a fluorescent signal produced by the action of an enzyme system active in the germination and outgrowth phases of the test organism- Geobacillus stearothermophilus (steam, hydrogen peroxide): alpha-

glucosidase- Bacillus atropheus (ethylene oxide): beta-glucosidase



Glucosidase Enzyme Systems

Bacterial glucosidase enzymes play an important role in uptake and utilization of nutrients

“The enzyme activity observed by sterilization assurance assays appears likely to come from heat stable enzyme in the spore core and enzyme(s) synthesized in spore outgrowth.”

Setlow P, Korza G, Setlow B. Analysis of alpha-glucosidase enzyme activity used in a rapid test for steam sterilization assurance. J Appl Microbiol 2016; 120: 1326-1335.

Vihinen M, Mäntsälä P (1989) Microbial amylolytic enzymes. CritRev Biochem Mol Biol 24:329–418


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3M™ Attest™ Rapid Technology

Enzyme acts on a non-fluorescent substrate in the media, resulting in a fluorescent end product, which is

detected by the Auto-reader (“signal”)

Non-fluorescent substrate (MUG)

Fluorescent end-product (4-MU)


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Rapid Attest™ Technology

Enzyme acts on a non-fluorescent substrate in the media, resulting in a fluorescent end product, which is

detected by the Auto-reader (“signal”)

Non-fluorescent substrate (MUG)

Fluorescent end-product (4-MU)



Positive Biological Indicator “Signals”





Positive Biological Indicator “Signals”



FluorescenceFluorescent response generated by metabolic enzyme action



Optimized for Signal Production and Detection



Optimized for Signal Production and Detection

Media optimized to produce fluorescent signal



Optimized for Digital Production and Detection

Physical design to concentrate the signal

Advanced optics and software to detect the signal



Rapid Readout Biological Indicators Use Worldwide

Malaysia JapanChinaTaiwanAustraliaNew ZealandIndiaThailandIndonesiaKoreaSingaporePhilippinesVietnam

United StatesCanadaMexicoBrazilColombiaChilePeruArgentinaUruguayCaribbean Region

SpainItalyGreeceBelgiumFranceNetherlandsSaudi ArabiaTurkeyPolandSouth AfricaUKIrelandRussiaIsrael



51

Use of Biological Indicators in Sterilization Quality Control



Sterilization Process Monitoring

Goal of the sterilization process?• To kill all microorganisms!



Biological Indicators

Chemical Indicators

Physical Monitors

Sterilization Process Monitoring Tools



Physical Monitors

•Information from sensors in the sterilizer chamber walls or drain (e.g. temperature, pressure)

•Can confirm that the correct cycle was selected

•Provides a record of the cycle • Sensor placement



Chemical Indicators

Test system that reveals change in one or more predefined process variables based on a chemical or physical change resulting from exposure to a processISO/TS 11139:2005 Sterilization of health care products — Vocabulary

Bowie-Dick Tests

Pack indicatorsProcess indicators



Biological indicatorsBiological indicators are recognized by most authorities as being closest to the ideal monitors of the sterilization process because they measure the sterilization process directly by using the most resistant microorganisms (i.e., Bacillus spores), and not by merely testing the physical and chemical conditions necessary for sterilization. Since the Bacillus spores used in biological indicators are more resistant and present in greater numbers than are the common microbial contaminants found on patient-care equipment, the demonstration that the biological indicator has been inactivated strongly implies that other potential pathogens in the load have been killed.

CDC Guideline for Disinfection and Sterilization in Healthcare Facilities, 2008



Biological indicators

Biological indicators are recognized by most authorities as being closest to the ideal monitors of the sterilization process because they measure the sterilization process directly by using the most resistant microorganisms (i.e., Bacillus spores), and not by merely testing the physical and chemical conditions necessary for sterilization. Since the Bacillus spores used in biological indicators are more resistant and present in greater numbers than are the common microbial contaminants found on patient-care equipment, the demonstration that the biological indicator has been inactivated strongly implies that other potential pathogens in the load have been killed.




Biological indicators

Biological indicators are recognized by most authorities as being closest to the ideal monitors of the sterilization process because they measure the sterilization process directly by using the most resistant microorganisms (i.e., Bacillus spores), and not by merely testing the physical and chemical conditions necessary for sterilization. Since the Bacillus spores used in biological indicators are more resistant and present in greater numbers than are the common microbial contaminants found on patient-care equipment, the demonstration that the biological indicator has been inactivated strongly implies that other potential pathogens in the load have been killed.




Process Challenge Devicesprocess challenge device (PCD): Item designed to constitute a defined resistance to a sterilization process and used to assess performance of the process.

ISO/TS 11139: 2005. Sterilization of health care products — Vocabulary. International Organization for Standardization.



Use of PCDs

• Biological indicators should be inside of the load, but this is not feasible

• PCDs are easily retrieved without disturbing any of the load items

• PCDs make it more difficult for the sterilant to reach the BI, like the BI was located inside of the load



Frequency of use – biological indicators

Steam sterilization

Test CAN/CSA-Z314.-18 U.S.: ANSI/AAMI ST 79 (2017) Comprehensive guide to steam sterilization and sterility assurance in health care facilities

Routine Efficacy Monitoring BI PCD. Each day the sterilizer is used for each type of cycle

BI PCD. Weekly, preferably daily; Each cycle type used.

Load release – non-implant Physical parameters. BI PCD with CI provides increased sterility assurance

Optional. PCD with BI, Type 5, Type 6.

Load release - implants Every load. PCD with BI + Type 5 or 6 CI. Quarantine until BI result is available.

Every load. PCD with BI + Type 5 or 6 CI. Quarantine until BI result is available.



Considerations for monitoring every load with a BI/PCD:• Uniform standard of care for all patients

• Highest level of quality control

• Reduce the possibility of monitoring mistakes (implant vs. non-implant)

• Fast readout BIs facilitate frequent monitoring

Note: 53% of US Tier A hospitals monitor every load with BI PCD.

Source: 3M survey 2017, 2018.



Considerations for monitoring every load with a BI/PCD:• Uniform standard of care for all patients

• Highest level of quality control

• Reduce the possibility of monitoring mistakes (implant vs. non-implant)

• Fast readout BIs facilitate frequent monitoring

7 days 2 days 3 hours 1 hour Minutes!



Steam sterilization quality issues . . .

• Most steam sterilization failures are related to problems with the steam quality, rather than time or temperature (e.g. air, non-condensable gases, wet steam, dry steam)

• Physical monitors and most chemical indicators cannot detect these problems

• Poor steam quality reduces the killing power of the process. Biological indicators directly measure the effectiveness of the process, so they do detect steam quality problems.



SUMMARY1. Biological indicators are the direct measure of lethality of

the sterilization process, because they measure the effect of the process on spores.

2. Biological indicators can detect cycle quality problems.

3. Biological indicators incubation times have been reduced by detecting biological signals faster.

4. Fast incubation times make BIs easier to use, and enable better quality control by increased monitoring frequency.



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Thank you for your time and attention.

Contact information:

Janet Prust3M Medical Solutions DivisionSt. Paul, MN USA651 736 1546

[email protected]

Documents

The Science of Speed: How Fast Biological Indicators have ......ISO 11138 Series 11138 –2: Ethylene oxide 11138 –3: Moist heat (steam) 11138 –4: Dry heat 11138 -5: Low temperature