Hein Venter

Microbiological Quality Control

requirements in an ISO/IEC 17025
laboratory
Anatech Instruments
hein@anatech.co.za
 ISO/IEC 17025:2017
The international standard used to accredit competence of
testing and calibration laboratories worldwide

Laboratories assure that they have certain qualities:

- Ability technically to get a valid result. This involves

people, knowledge, equipment, supplies and processes
“getting it right”
- A system to ensure impartiality, consistency, reliability
“once right……always right”
 :2017 Risk Based Thinking

Decisions Based on Anticipated Risk and Opportunity

- Prescriptive requirements replaced by performance based
requirements
- Greater flexibility for processes, procedures, documented
information and organizational responsibilities
- Requires laboratory to plan and implement actions that
address risks
 The purpose of Risk Based Thinking
8.5.1 The Laboratory shall consider the risk and opportunities
associated with the laboratory activities in order to:
- Give assurance that the management system can achieve
its intended results
- Enhance opportunities to achieve the purpose and
objectives of the laboratory
- Prevent or reduce undesired impacts and potential failures
in the laboratory activities
- Achieve improvement
 Controls are the Foundation

Don’t risk your reputation

Microbiology controls are

a small portion of the
department budget, but
can prevent audit findings
and inaccurate results

Verify that risks are

managed through the use
of high quality RM
What can go Wrong?

TR28
 Why Do We Do QC?

- Protect our patients/customers QC’s checks:

- Protect our reputation - Reagents
- Ensure testing procedures, - Equipment
equipment and materials are - Test method
working properly - The technique
- Ensure personnel competence - Competence
- Improve daily workflow and
produce consistent and reliable
results
- Reduces risk of misdiagnosis
 Standards and Guidelines

ISO/IEC 17025:2017
General requirements for the competence of testing and calibration
laboratories
ILAC-G9:2005
Guidelines for the Selection and Use of Reference Materials
SANS 11133:2017 (ISO 11133:2014)
Microbiology of food, animal feed and water — Preparation,
production, storage and performance testing of culture media
SANAS TR 28-01
Criteria for the Validation and Quality Assurance in Microbiological
Testing
 What should we test the media for?

Low number recovery

Selective removal of unwanted
organisms
Enrichment of required organisms
Contamination
Durability
Antimicrobial resistance
Consistent results
 QC Strains – What Matters?

Should be reliable
Should be traceable
Should be dependable

Should be readily accessible

Should not occupy a lengthy
amount of time in the working day
 Changes to ISO11133
Describes performance testing for food and water
culture media
- No longer optional to implement
- Required when performing ISO methods
- Normative standard for all detection and enumeration
standards
- Recommends specific microorganism strains for quality
control of media
- Requirements for daily enumerated controls
- Tables of control strains are normative
- Dilution to extinction method added for liquid media
- Includes performance testing of diluents and transport
media
 SANS 11133:2017
Introduction
Sufficient testing should be carried out to demonstrate
a) the acceptability of each batch of medium,
b) that the medium is “fit for purpose”, and
c) that the medium can produce consistent results.
These three criteria are an essential part of internal quality control
procedures and, with appropriate documentation, will permit effective
monitoring of culture media and contribute to the production of both
accurate and reliable data.
Scope
This International Standard applies to producers such as:
a) commercial bodies producing and/or distributing ready-to-use or
semi-finished reconstituted or dehydrated media;
b) non-commercial bodies supplying media to third parties;
c) microbiological laboratories preparing culture media for their own
use.
 5 Test organisms for performance testing

5.3 Preservation and maintenance of test organisms

- lyophilization, storage on beads at –70 °C, or using liquid nitrogen
- number of transfers of test organisms should be documented
- Culture collections & Commercial suppliers - manufacturer’s
directions for their cultivation and use shall be followed
- Maintenance techniques should minimizes the opportunity for
cross contamination, mutation or alteration of typical
characteristics
Reference Strain From Culture Collection or Commercial Body
Reference stock cultures from Reference Strain
Stock cultures from Reference Stock Culture
Working cultures from Stock Culture
Maintenance of QC Strains
Reference Strain

Reference Stock Culture

Monthly Reference Stock Culture

Working Stock Culture

5.4.2.5.1 Inoculum level for productivity testing

5.4.2.5.1.1 Quantitative testing

- 80 cfu to 120 cfu per plate, minimum number of 50 cfu per plate
- Apply to Solid media, Filters, Dilutions and Transport media

5.4.2.5.1.2 Qualitative testing

- 103 to 104 cfu for qualitative tests of plate media
- ≤100 cfu productivity tests of pre-enrichment and enrichment media
- 104 to 106 cfu for qualitative tests of solid transport media

5.4.2.5.2 Inoculum level for selectivity testing

- 104 to 106 cfu is inoculated on to the plate or into the tube of medium

5.4.2.5.3 Inoculum level for specificity testing TR28

- inoculum level of 103 to 104 cfu is needed 6. The Use of Media
 Productivity Testing
6.4 General requirements for microbiological performance testing
When culture media are intended for enumeration purposes, quantitative testing
methods shall be performed.
7.2.1.1 Productivity
PR = NS / NO
NS = total colonies on test medium
NO = total colonies on ref medium - usually TSA

For Selective media

The Productivity Ratio ≥ 0.5
PR = Ns/No
Ns is the test medium (Baird Parker) and
NO is the reference culture medium.
TSA = 100, BPA = ≥ 50
 Acceptance criteria - solid media
7.2.2.1.2 Calculation and interpretation of results
Performance ratio should be:
- >0.70 for nonselective against ref
- >0.70 for all media tested against previous batch of the same
media
- >0.50 for selective media against non selective reference medium
- Investigate if >1.4
- For water media minimum PR =>0.80
These values are minimum requirements, not targets.

7.4 Methods for qualitative tests

Performance Testing Requirements
 8 Methods for performance testing of liquid culture media

8.2 Quantitative tube method for performance testing of liquid enrichment media
- dilution to extinction method

8.3 Qualitative tube method for performance testing of selective liquid media
- Inoculation of target organisms - ≤ 100 cfu
- Inoculation of non-target microorganisms - >1 000 cfu
- streak on a plate of a non-selective medium (e.g. TSA), selective medium (e.g. XLD)

8.4 Qualitative single tube method (turbidity) for performance testing of liquid media
8.4.2.1 Pre-enrichment media
- inoculum volume containing ≤ 100 cfu
8.4.2.2 Confirmation media
- Inoculum volume containing > 106 cfu/ml

subculture to a solid medium to demonstrate growth

 TR28 - 7. The Use of Reference Materials and Reference
Cultures
7.1 Reference materials shall, where possible, be traceable to SI units of
measurement, or to certified reference materials. Internal reference materials
shall be checked as far as is technically and economically practicable.

7.5.1 Reference materials

Reference materials and certified reference materials provide essential traceability
in measurements and are used, for example;
- to demonstrate the accuracy of results,
- to calibrate equipment,
- to monitor laboratory performance,
- to validate methods, and
- to enable comparison of methods.
If possible, reference materials should be used in appropriate matrices

Different from Daily Qualitative and Quantitative Quality

Controls
 What is Certified Reference Material?
Reference Materials:
- Homogenous
- Stable
- Fit for Use
- Quantitative or Qualitative
- ISO Guide 34

Certified Reference Material, CRM,

- Is metrologically traceable
- Each property value has an uncertainty assigned to it
- The material comes with a certificate providing traceability and
uncertainty

Metrological traceability means that there is a documented

unbroken chain of calibrations.
Requirements of a CRM COA
 Importance of Traceability
Traceability – verifies the organism is from an accredited and
reliable source, a license agreement verifies traceability
Security – verifies the organism is correct, which increase
confidence during testing and audits
Authenticity – verifies the correct identification and the
organism is not an “equivalent”
Certifications – verifies compliance with global standards

No traceability = no point!
Look for the ATCC Licensed Derivative® emblem for products derived
from ATCC® cultures.
Proficiency Testing Is Not a Quality Control Program

Good QC ensures that all process are in place and

working correctly

No need for concern when the PT sample arrives

“Royal treatment” will not be required

Because all systems are in place and working you can

relax!

You are assured that you will get correct results.

 Remember

QC is your responsibility and cannot be contracted out

Failure to QC correctly means you are risking:
- Failure of product
- Reputation of your company / institution
- Repercussions
- There are no short cuts only risks
QC is a test of the quality of the media not of the quality of the
quality control strain
PT’s are one-offs/ QC is on going
Organisms are living entities and therefore by nature they are
predictably unpredictable