Methods for Microbial Testing of Endoscopes

• Mark Hichens Cbiol, MRSB, MSc

Determination of Total Viable Count in final rinse waters by The Method of Membrane Filtration

Applicable standards

• Washer-disinfectors Part 2: Requirements and tests for washer-disinfectors employing thermal disinfection for surgical instruments, anaesthetic equipment, bowls, dishes, receivers, utensils, glassware, etc. (ISO 15883-2:2006)

• Washer-disinfectors - Part 4: Requirements and tests for washer-disinfectors employing chemical disinfection for thermolabile endoscopes (ISO 15883-4:2008)

• Health Technical Memorandum 2030 Part 3 Washer-disinfectors Validation and verification

• HSE Standards and Recommended Practices for Endoscope Reprocessing Units. QPSD-D-005-2. V 2.0

• HSE Standards and Recommended Practices for Central Decontamination Units, Version 2.0 – 2011

Validation and process control

• Critical limits

• Reproducibility

• Repeatability

• Linearity

• Reliability

• Accuracy and sensitivity

• Sample bottles

• Media

• Filters

• Filtration apparatus

• Incubators

• Scientists

Final test Quality Control

• Internal quality control

• External quality control

Sample numberSample matrix Date Count A Count B log10A Log10B A-B A+B RSD^2 RSDR

24179 27.09.16 6 4 2 0.77815125 0.602059991 0.176091 1.3802112 0.03255476 0.18043

24220 25.05.17 4 4 2 0.602059991 0.602059991 0 1.20412 0 0.00000

24295 14.11.17 2 7 2 0.301029996 0.84509804 -0.544068 1.146128 0.45068195 0.67133

24319 30.09.16 4 4 2 0.602059991 0.602059991 0 1.20412 0 0.00000

24409 26.09.16 7 10 2 0.84509804 1 -0.154902 1.845098 0.01409629 0.11873

24498 14.08.17 2 3 2 0.301029996 0.477121255 -0.176091 0.7781513 0.1024183 0.32003

24511 20.12.16 4 6 2 0.602059991 0.77815125 -0.176091 1.3802112 0.03255476 0.18043

24608 25.11.16 5 4 2 0.698970004 0.602059991 0.09691 1.30103 0.01109667 0.10534

24622 23.05.17 6 6 2 0.77815125 0.77815125 0 1.5563025 0 0.00000

24660 03.11.17 7 5 2 0.84509804 0.698970004 0.146128 1.544068 0.01791283 0.13384

24670 23.11.17 7 5 2 0.84509804 0.698970004 0.146128 1.544068 0.01791283 0.13384

24682 01.09.17 9 4 2 0.954242509 0.602059991 0.352183 1.5563025 0.1024183 0.32003

24692 03.08.17 3 10 2 0.477121255 1 -0.522879 1.4771213 0.25061074 0.50061

24738 15.08.17 3 5 2 0.477121255 0.698970004 -0.221849 1.1760913 0.07116426 0.26677

24756 04.08.17 2 4 2 0.301029996 0.602059991 -0.30103 0.90309 0.22222222 0.47140

Uncertainty of Measurement (RSD RC)

This sheet assists in providing an estimate of the uncertainty of measurement obtained from the paired counts.

The relative s tandard deviation of reproducibi l i ty (RSD R) for each pair i s ca lculated after a common logari thms (log 10) transformation of the

count. These are combined to obtain the "average" RSD RC expressed as a log10 va lue. Log transformation provides the nummeric RSDrc

Test portions with less than two results or for which one or more of the counts are zero are excluded.

Quality Control

Process Control

Ensure the component parts of the process are working effectively.

Ensure that the component parts are working consistently. Evaluate the component parts to ensure working consistently and improve the quality of the end product.

Product control

Ensure the product meets the demands of its intended purpose

FDA Cracks Down on Duodenoscope Manufacturers - Again

• FDA in 2015 ordered manufacturers to conduct a post-market surveillance study to determine whether healthcare facilities were able to properly clean and disinfect the device –Quality Control !

• Three companies were required to conduct a study to sample and culture reprocessed duodenoscopes

• Olympus failed to commence data collection, and Pentax and Fujifilm have failed to provide sufficient data.

• FDA threatened seizure, injunction, and civil money penalties.

Health Service Executive Standards and Recommended Practices for Commissioning, Validation and Testing in Endoscope Decontamination Units

The guidance is close to publication

It is comprehensive

Introduces new testing procedures, acceptable parameters

and recommends frequency of testing

Endoscope cleaning and disinfection process (Major components).

Cleaning Efficacy by residual soil detection Weekly Testing

Contamination of endoscopes will be proteinaceous in nature.

Cleaning efficacy tests undertaken using an artificial test soil and surrogate device accordance with EN ISO 15883 Part 5.

Qualification:- The chamber walls and load carrier should be free of residual soil

Process Challenge Device (PCD) tests (Hexalumen, Tosi, Flexicheck)

Qualification:- Process challenge devices test should be in accordance with the limits described by the manufacturer.

Frequency - weekly.

Residual Protein Test

• Determine the effectiveness of the cleaning process by determining residual protein

• Residual contamination can be made clearly visibly by swabbing the surfaces or lumens of the endoscope using residual protein tests capable of detecting >1µg of protein.

• Swabbing methods based on Bradford assay or Coomassie blue reagents are widely available

• If appropriately validated/quality controlled.

Residual Protein -frequency of testing

Ensuring testing of every product family in every chamber every quarter and each scope annually

DESCRIPTION

A

B

C

D

E

F

G

Endoscopes and

Product Families

Processed

Total Number of

Product

Families

Processed

Basins

(positions)

Required

Number of

Test for Machine Performance

per quarter

(A x B)

Required

Number of

Test for Machine Performance

per year

(C X 4)

Total number of

scopes

available to

the user

Required

number of

additional

tests to

establish

scope integrity.

E-D when E

is more than D

Total number of

tests per

year

D+F

Bronchoscope and

Cystoscopes, both

belonging to product family number 3

1 2 2 tests 8 5 0 8

Gastroscope product

family number 1

ERCP product family

number 2

Cystoscope product family number 3

3 8 24 tests 96 120 24 120

Disinfection efficacy

• Annual EWD Disinfection Efficacy testing

• Disinfection efficacy should be verified using the test method described in EN15883 (2009) part 4 section 4.4.2.4.

• Endoscope Lumen Sampling

Endoscope Lumen Sampling

• The Endoscope is brushed through with a sterile disposable long-stemmed cleaning brush after processing

• The brush end is aseptically transferred to 10 mL of sterile peptone water containing a neutraliser

• The eluate is filtered through a 47 mm diameter 0.45 m filter and then the filter placed onto a R2A agar plate and incubated at 30±2°C for up to five days

• The number of colony forming units (cfu) are counted

Guidance Levels

A result of less than 10 cfu per brush sample

should be obtained if the Endoscope

has been satisfactorily disinfected

Alternative technique - Lumen flushing

• Passing sterile water/neutralising buffer down the Endoscope lumens- suction, air/water and biopsy lumen

• Flush ~10 mL of water down the lumen and collect into a sterile sample bottle at the distal end.

• Laboratory analysis, examine for total viable count on R2A TSA or YEA media with incubation at 30±2°C for up to five days.

• The results should be below 10 cfu/100 mL.

• Good aseptic technique are essential to obtain good results

Endoscopes and Product Families Processed

Total Number of Product Families Processed

Scopes available to the user in each product family

Sampling Strategy

Cystoscopes, belonging to product family number 3

1 1 Cystoscope The single endoscope will be sampled every

quarter.

Bronchoscopes and Cystoscopes, both belonging to product family number 3

1 1 Bronchoscopes

1 Cystoscope

The two endoscopes will be sampled

alternatively every quarter as they are part of

the same product family. Therefore, each scope

will be sampled twice in the year.

ERCPs product family number 2 Gastroscope / Colonoscopes both belonging to product family number 1

2 1 ERCP endoscope

1 Gastroscope

1 Colonoscope

The ERCP endoscope will be sampled every

quarter.

The gastroscope and colonoscope will be

sampled alternatively every quarter as they are

part of the same product family.

ERCPs and EUS both belonging to product family number 2 Gastroscope product family number 1

2 1 ERCP endoscope

3 EUS endoscopes

1 Gastroscope

The ERCP and EUS endoscopes will be sampled

once each over the 12 month period as they are

part of the same product family and testing

should be rotated amongst them each quarter.

The gastroscope will be sampled every quarter

as it is the only member of that product family.

Quality of water

• Critical to the outcome of the decontamination process.

• Microbiologically

• Efficacy of cleaning agents and disinfectants

Supplied water quality Determinant and Unit Maximum Permitted Values

Final Rinse Other Stages

Appearance Clear, colourless

Degree of acidity (pH) 5.5 to 8.0

Conductivity at 25º (uS/cm) 30

Total dissolved solids (mg/100mL) 4

Total hardness, CaCO3 (mg/L) 50 200

Chloride, CI (mg/L) 10 120 10 120

Heavy metals, determined as Lead, Pb (mg/L)

10

Iron, Fe (mg/L) 2

Phosphate, P2O5 (mg/L) 0.2

Silicate, SiO2 (mg/L) 0.2

Total viable count (TVC) at 30ºC <10

Bacterial endotoxins (EU/mL) 0.25

Frequency – annually.

Final Rinse Water

• Microbiological quality of the water needs to be assessed regularly in accordance with EN ISO 15883 Part 1:2014 Annex D

• Total viable counts

• pH

• Conductivity

• Hardness

• Weekly

Aerobic colony count

(cfu) in 100mL

Interpretation Action and advice Colour grade

<1 cfu Satisfactory Use as normal Green

1 – 9 cfu on a regular

basis

Acceptable –indicates

that bacterial

numbers are under a

reasonable level of

control

Use as normal Green

10 – 100cfu Risk Assess

Request identification

of predominant

organisms and

resample

Carry out formal risk

assessment, based

upon clinical needs in

consultation with the

consultant

Microbiologist the IPC

team and the AE(D)

Orange

>100cfu Risk Assess

As above. Request

identification of

predominant

organisms and

resample

EWD should be taken

out of service until

water quality has

improved / issue

resolved.

RED

Pseudomonas aeruginosa

• Gram negative opportunistic pathogen. Ubiquitous in the environment

• Able to form biofilms and exhibit resistance to disinfectants

• In endoscopy is the most commonly reported micro-organism responsible for transmission of infection

Environmental mycobacteria (rapid growing)

• Opportunistic pathogens found widely in the environment. Commonly found in water distribution systems

• Exhibit resistance to high level disinfectants due to presence of hydrophobic lipid rich cell wall

• Infection of the patient may lead to miss-diagnosis of tuberculosis and implementation of un-necessary antibiotic therapy

Storage Cabinets for Heat Labile Endoscopes (CESC’s)

ISO EN 16442 Controlled environment storage cabinet for disinfected thermolabile endoscopes

The controlled environment provided by any CESC should ensure that there is no deterioration of the microbiological quality of the Endoscope

Routine Testing of CESC’s

Surface contamination

Indication of overall cleanliness

Indication of cleaning efficacy

Aero-contamination

Indication of the circulating air, and air that may be entering the cabinet

Annual re-validation

Placing of a sterile surrogate device in the cabinet for the validated time period confirmed at initial validation

Description of Test

Frequency

Result Required

Comments

Determine the contamination levels on the inside surfaces of the cabinet

Quarterly

The contamination levels identified shall be less than 25 cfu/25 cm² EN 16442 – Section 6.5

Reference should be ne made to EN 16442 – section 6.5

NB: Pathogenic organisms to be identified.

Check that the cabinet is capable of maintaining the quality of the Endoscopes

Annual The acceptable result for the test is

<10 cfu/lumen. Sterile surrogate device to be fitted to the Endoscope drying cabinet and left for a period of no less than what was established during commissioning.

Evaluate airborne microbial contamination.

Annual Reference should be made to EN

16442 Annex C

Environmental monitoring

• Alert level – CFU Levels that when exceeded, signal a possible deviation from that normally seen.

• Action Level – CFU levels that significantly exceed those normally seen.

Vacuum storage systems

• Validation of storage time

• Periodic validation of such systems is required to verify performance

• As specified by the manufacturer and/or by risk assessment

• Endoscope Lumen Sampling

• At the end of the defined storage period.

• There should be no degradation of the microbiological quality.

• Less than 10 cfu per sample

Final release testing

• Residual protein tests on reprocessed endoscopes as a measure of cleaning efficacy

• Endoscope Lumen Sampling as a measure of disinfection and storage efficacy

There is no such thing as chance or accident; the words merely signify our ignorance of some real and immediate cause. (Adam Clarke 1760-1832)