Microbiology OOS Flashcards
Microbiology Phase 1a
As described in the guidance this part of the investigation relates to elimination or confirmation of an obvious laboratory error. Examples include:
* Equipment failure e.g. autoclave or incubator impacting on the sterility of test media and giving a false positive result or failure to provide the correct incubation and growth temperature resulting in slower growth respectively
* Testing error e.g. negative or positive control failure, growth on a plate not in the test area.
In the latter case poor aseptic technique can result in a mould growth at the edge of the test petri-dish in an area away from the test sample
* Media failure (storage, use/re-melting). Growth media must be stored as per the manufacturer’s instructions to prevent deleterious effects on the chemical nature and subsequent impact on the ability of the microbes to recover and utilise the media for growth and detection
* Use and the re-melting of solid agar media can also affect the quality of the media and hence the testing reliability. Continued re-melting will denature some of the chemical growth elements and the media manufacturer must be consulted on the ability of the media to be re-melted
* A change in media supplier may also result in variability of the quality of the media e.g. peptone quality and will also influence the recovery and growth characteristics. Change control, impact assessment and revalidation need to be considered
* Correct method. Although possibly an obvious error, cases are still prevalent where the wrong method has been applied for the organisms being detected or a version update has not been incorporated. Equally the correct method may not have been followed and complied with e.g. use of the wrong neutraliser or dilution factor
Whatever the nature of the obvious error these issues should still be trended, so any recurrence is recorded and investigated.
Microbiology Phase 1b
If no obvious error is identified the investigation moves to Phase 1b.
At this point the analyst and supervisor work together to look at other possible causes for the unexpected result. Typically this will involve working through a checklist approach. The checklist should capture previous incidents and be updated as experience is gained from other testing issues.
The checklist should be part of the formal laboratory investigation process.
Examples included on the checklist related to microbiology testing include:
* Data review e.g. calculations related to dilution factors, examination of recorded sample storage times and conditions. Media batch numbers, recorded usage and expiry dates should be checked
* Equipment log-books for usage and conditions for the testing e.g. incubator temperatures calibration status and compliance with the test conditions. Examples have been recorded where the test media/sample were incubated at the wrong temperature. The reliability of a pH meter, if used for growth media checks, needs to be assessed e.g. damage to the pH probe.
* The supervisor checking with the technician the skill and competency of their aseptic technique. The manual manipulation of the sample is a critical part of the sampling and testing in microbiology. The application of the competency can be influenced by a variety of factors, including the number and type of samples being tested i.e. workload for the technician and the operating environment (insufficient workspace and cramped working conditions). Sample integrity and use of sterile sampling equipment reduces the risk of cross contamination
* Reviewing previous results for the test, product or technician to determine whether previous alerts (not action or above specification) have been recorded and captured in previous trends
Note that a specific challenge investigating microbiology results is the time taken since the analysis. Routine QC results including material, product and environmental monitoring data require from 3 to 7 days to be available and a sterility test needs 14 days. As a consequence a considerable time delay has occurred since the sample was tested. This has an impact on the ability to recall events or examine retained sampling and testing materials.
Procedures for the microbiologist should reflect this risk and particular attention given to accurate and detailed
record keeping and retention of items related to testing.
Microbiology Phase 2
If the outcome of Laboratory Phase 1a and 1b does not identify an assignable root cause or causes the investigation moves to Phase 2.
At this point a formal deviation is raised within the PQS to facilitate a wider investigation. The deviation should be communicated to the MAH, QP, QA and Production as they will have involvement in the Phase 2 actions.
In addition to further laboratory work there is a documented formal request for a thorough manufacturing investigation to take place:
* The laboratory investigation is directed towards developing a predefined testing plan often termed hypothesis testing. This requires identifying possible causes or eliminating influencing factors, such that an expected outcome is justified in advance of the testing taking place. This will reference what is being tested and why and how the results will be evaluated. The results generated do not replace the original data but provide additional information upon which decisions can be taken and justified. This reduces the risk of random actions or insufficient thought being given to the test. In contrast to analytical testing it is difficult to support the use of averages in microbiology results or the importance of outlier and other statistical tests. This is due to the inherent variation in microbiology analysis
* At this stage it is recommended that an additional at least equally experienced technician also undertakes some of the work to reduce the risk of technician variability. The work may involve the retesting of the same sample, or in rare cases, requesting another sample.
Where a new sample is requested it is recommended that quality assurance and a QP approve this step as it calls into question the sample integrity. It is also recommended that the hypothesis plan should be approved by QA/QP
* Phase 2 investigation also requires a detailed manufacturing investigation in parallel with detailed laboratory failure investigation. Prior reference to the time lapse in microbiology results being available also has significant impact on the manufacturing investigation. Since the taking of the sample there may have been significant changes to manufacturing environment:
○ Other materials/products may have been processed in the area
○ Cleaning and disinfection may have removed sources of contamination
○ Personnel activity in the area will have influenced the environment
Microbiology Phase 3
Phase 3 is described as the additional actions to be taken as part of the investigation of rejected batch(es). This captures the following:
* Impact assessment on other batches/equipment/activity. A small amount of contamination can quickly spread
* Identify and implement CAPA to prevent recurrence of the contamination
* In microbiological contamination incidents, particular consideration should be given to the deployment of QRM as root cause(s) may not have been identified
* Phase 3 concludes with a statement on the contamination impact of the rejection e.g. medical impact on the patient, deleterious product and spoilage effects
* Once rejected a batch can still be tested for root cause but the rejection cannot be reversed
Microbiology Test Results – Stability Results, OOT, OEE
Where microbiology results move to Phase 3 and arise from a stability test it requires prompt communication to the MAH, Regulator and QP as there is the possibility of product recall. Actions may include the testing of an earlier time point e.g. failure at 18 months and next planned testing at 24 months then testing may take place when the 18 months are confirmed OOS.
Where microbiology results are OOT but below action or alert limits e.g. environmental monitoring then statistics may be used for prediction of results at future time points, but caution has to be exercised given the previously stated dynamic nature of microbes e.g. the nature and population of the contamination will be affected by the use of the oxygen in the headspace of the product container.
A single atypical, OOE result still requires review, documenting and trending. Repeated, seemingly unrelated OOE results in a product, material or environment may be early indications of repeated contamination e.g. isolation of unexpected contaminant and possible repeat or change in levels of contaminants. There are significant differences in approach for sterile and non-sterile products and operations.
Microbiology Test Results – Stability Results, OOT, OEE
Where microbiology results move to Phase 3 and arise from a stability test it requires prompt communication to the MAH, Regulator and QP as there is the possibility of product recall. Actions may include the testing of an earlier time point e.g. failure at 18 months and next planned testing at 24 months then testing may take place when the 18 months are confirmed OOS.
Where microbiology results are OOT but below action or alert limits e.g. environmental monitoring
statistics may be used for prediction of results at future time points, but caution has to be exercised given the previously stated dynamic nature of microbes e.g. the nature and population of the contamination will be affected by the use of the oxygen in the headspace of the product container.
A single atypical, OOE result
still requires review, documenting and trending. Repeated, seemingly unrelated OOE results in a product, material or environment may be early indications of repeated contamination e.g. isolation of unexpected contaminant and possible repeat or change in levels of contaminants. There are significant differences in approach for sterile and non-sterile products and operations.
EM failure investigation example
Review:
personnel activity
Airflow patterns and HEPA testing
Aseptic technique
Gowning procedures
Trends and incidents
Maintenance
Pressure cascades
Room cleaning
Sanitisation procedures
training records
Interventions
Sterilisation cycles
Non-sterile products - key aspects of microbiological control
Good quality raw materials
Well controlled and maintained water systems
Well maintained plant
Effective cleaning and sanitisation
Protect product as much as possible
Understand and controlling any parts of the process at greater risk of contamination - wet gran, sugar coat, equipment cleaning
Well trained operators
Product storage controlled
