A REPORT ON

STERILTY TEST FAILURE INVESTIGATION

OF

SYRINGE AND NEEDLE SETS BATCHES:

ABS221102, ABS221021, ABS221022, DBS221019, CCS221101, CC221102

AND

HYPODERMIC NEEDLE BATCH BFN221001

NOVEMBER, 2022
 TABLE OF CONTENTS
A. INTRODUCTION AND PREAMBLE
1. Failure scope
2. The Objective of the investigation
3. Investigation Procedure
4. Personnel Involved in the Investigation

B. IMMEDIATE ACTIONS TAKEN

1. TEST
i Culture media
ii Validation of equipment
iii History of Sterility test
iv Environmental Monitoring
v Sterility Test Analyst
vi Testing environment

2. PRODUCTION ENVIRONMENT
i Input materials and regrinds not hygienically kept
ii Container integrity
iii Process
iv After blistering
v Assembly Clean Room and Line
a) Equipment swab and personal hygiene check
b) HVAC
c) Environmental parameter check
d) Operators
e) Environmental Monitoring
f) Cleaning and disinfection record
g) Sterility test link with process Area

vi Sample resting
vii Pyrogen test

C.ANNEXURE I-XI

D. APPENDIX I-XXX
 A. INTRODUCTION AND PRE-AMBLE
Sterility test is used as a product release test where the sterility of a product is described
by the absence of viable and actively multiplying microorganisms when the product is
tested in specified culture media (compendia test). Detection of microbial growth in the
medical device product results in considering it as a non- sterile product, until an
investigation is performed establishing otherwise. An established failure of product
sterility leads to an adulterated product.
The method employed in conducting the sterility test is as clearly documented in the
company’s Standard operating Procedure reference HMA/QA/SOP030-00 in accordance
with the specifications of BP. 2022 and NIS 238:2020
1. Failure Scope
The scope of this investigation is to examine the laboratory test and the production
process to determine why the sterility test failures of seven (7) batches of finished
products could have occurred.

2. The Objective of the investigation

The objective of this investigation is to establish the root cause of the sterility test
failure, undertake corrective and preventive actions (CAPA) and to demonstrate that
the action taken is effective.

3. Investigation Procedure
The investigation was conducted in line with the Organisation’s Standard Operating
Procedure for Investigating Sterility Test Failure reference HMA/QA/SOP030-00 and
the Root Cause Analysis using Fish Bone Method.

4. Personnel Involved in the Investigation

The personnel involved included the Quality Control Manager, the Production
Manager, the Lead Microbiologist, Microbiology Analyst, Chemical Analyst, IPQA
Officer, Production and Shift Supervisors.
B. IMMEDIATE ACTIONS TAKEN
Due to the public health implication of a non-sterility findings, preliminary results of
the findings
was reported to the management. The affected batches on ground were immediately
quarantined and the investigation of the HACCP immediately commenced to
troubleshoot where
the problem emanated from. The areas of concentration included the test, test
environment and
production process, personnel, production environment, nature(weather), equipment
and
machines. Documented decision to increase the regime of dosage of gas sterilant was
taken by the
production crew authorized by the General Manager.

1. Tests
i. Culture media: Fluid Thioglycolate Medium, Tryptone Soya Broth
were prepared by the Microbiologist, who has more than 1year industrial
experience in doing this.
Growth promotion, positive and negative control tests performed were checked by
the Lead microbiologist to verify the growth supporting ability of the medium and
success of the medium sterilization process. The result obtained for the negative
control was as shown in ANNEXURE II indicating the media wasn’t the source
of contamination.
ii. Validation of equipment: Autoclave, Laminar Air Flow Cabinet were validated
by the Lead microbiologist. the result obtained for the autoclave validation is as
shown in ANNEXURE I indicating again the media wasn’t the source of the
failure as other batches of products analysed with the same batch of media passed
sterility test.
iii. History of Sterility Test: No failure of sterility test recorded for the past 2years.
Last occurrence was during the inception of dry season over two years ago.

iv. Environmental Monitoring: The environmental bioburden of the sterility room

as shown in ANNEXURE IIIa, IIIb & IIIc shows the sterility laboratory
complies with specification for the months of September, October, and November.
The environmental bioburden routine checks carried out over a period of over
3months for the assembly cleanrooms and operations shows noncompliance of
these units to specifications as high bioburden was recorded all through this
period. Microorganism such as Aspergillus sp. was isolated from the main
packaging area of the factory, which is a microorganism that poses a great threat in
the pharmaceutical industry
 v. The disinfectant and soap used in the cleaning of the production areas and
microbiology laboratory for over 6 months except from early November 2022 was
analysed for determination of its antimicrobial effectiveness and the result
obtained is as shown in ANNEXURE IV which indicated complete
ineffectiveness in antimicrobial activity. The use of particle shedding material for
cleaning in the production areas will also serve as a source of contamination.

vi. Sterility Test Analyst: The microbiology Analyst was validated by administering
Questionnaire to check if he carried out microbiology procedures according to
standard operating procedure (SOP). The response to this shows that the
microbiology analyst carries out analysis according to standard operating
procedures as shown in ANNEXURE V. His history of carrying out sterility test
was also examined and the result of this examination shows most of the sterility
test carried out by him over a period of one year passed sterility test.
vii. Testing environment: The laminar airflow chamber was validated to check if the
parameters that determines its effectiveness complies with specifications; the
results obtained were as shown in ANNEXURE VI showing the laminar flow
used in the analysis of the affected batches is working effectively as placed in an
ISO Class 5/Class B zone
viii. Sampling for Testing and Retention: The sampling for testing of sterile product
as obtained in this test was representative enough to give a reliable, representative
results as the sterilization chamber has been earlier on properly mapped out into 25
locations for the bigger chamber and 15 for the smaller chamber with the areas
taking into considerations being the remote areas where the sterilant gas are not
likely to reach.
An error was however discovered in the sampling of the sterilized product for
retention sample which were taken from a fixed location and therefore are biased
samples which cannot represent the entire population. An immediate CAPA has
been raised for this to be corrected. A retrain is also scheduled to engage the staff
in SOP for sampling for microbiological and physicochemical analysis of Medical
Device.
Collected samples in the microbiology lab also need to be more protected by
storing in the sterility lab and not anywhere else to protect it from extraneous
contamination.

2. Production Environment
i. Input materials and regrinds not hygienically kept
ii. Container integrity: physical examinations revealed perforated, open bags with
infestation of insects, rodents and their faeces. Microbial enumeration of input
materials, regrinds, intermediate components showed contaminated raw materials,
highly contaminated stored regrinds, components obtained immediately from
injection moulding with no growth, then a progressive build-up of microbial load
from barrel printing to pre-sterilization blistering and packaging unit as shown in
ANNEXURE VII

iii. Process: The investigation carried out indicated that majority of the affected
batches were sterilized by a particular E.O injection Operator. As shown in
ANNEXURE VIII, the result obtained from the administered questionnaire also
indicates the operator possess only peripheral knowledge of the sterilization process
and its implication.in any unusual events; checking sterilization records for EO gas
injection log, the records are unsatisfactory and suggestive of short charging
ethylene oxide gas quantity used in sterilization. Non-challant attitude towards
corrections is a common scene here. Communication gap between shifts is a
common occurrence which do affect the accuracy in the operation of this vital unit.
iv. After blistering, pre sterilization bioburden: The result of total viable counts of
pre-sterilized blistered products gave a high level of bioburden as shown in
ANNEXURE VIIa
v. Assembly Clean Room and Line:
a. the routine equipment swabs for hygiene check as shown in ANNEXURE IXa,
showed noncompliance of the hygiene of the equipment with specifications.
b. The HVAC checks for a period of over 3months (September to November)
shows non-compliance of the air delivery volume to specification as the air
delivery volume keeps depreciating since September till date as shown in
APPENDIX XIV & XV
c. Humidity, Temperature, Air particulate count and Pressure differential
shows noncompliance with specification of ISO Class 5/Class B zone as shown
in APPENDIX I-XXX
d. Operators: personnel hygiene and gowning check as tested by finger
dabbing, glove print as shown in ANNEXURE IXb, IXc shows the hygiene of
the production staff to be noncompliant with specification. The cultural and
microscopic characteristic of the isolated micro-organism from their glove and
finger was found to be Staphylococcus aureus which is the same microorganism
implicated in the sterility test failure.
e. Environmental Monitoring – Viable and Non-viable particulate including
identification of microbes from both critical and non-critical areas, with the
observation of trends which may indicate gradual degradation or deterioration in
operational handling
 f. Cleaning and disinfection records in assembly clean rooms. The
microbiological analysis of disinfectant used in cleaning was analysed for its
effectiveness and the result obtained was as shown in ANNEXURE IV proved
the disinfectant used till Early November, 2022 in cleaning to be ineffective
antimicrobially.
g. Sterility test link with process Area: Cultural and microscopic identification of
the microorganism isolated from failed sterility samples identified one of the
microorganism to be Staphylococcus aureus and the other to be Bacillus sp this
was confirmed by use of selective medium that allows the growth of
Staphylococcus aureus as shown in ANNEXURE X, this result further indicated
a link between the microbial contamination of the product and the poor hygiene
practices of the production staff and poor environmental hygiene since the same
microorganisms were isolated from the glove, finger dabbing and floor swab
tests of production floor and staff from different units while Bacillus is usually
of environmental origin .

ix. Retesting: The remnant sample of the failed batches was reanalysed by the Lead
microbiologist to confirm the sample’s failed sterility test. The result obtained
showed failed
sterility test for the second time in less than 24hrs. This outcome rules out
sample’s failure
of sterility as a result of microbiologist error as shown in ANNEXURE II.
x. Pyrogen Test: the result obtained from pyrogen complied with specifications.
As shown in
ANNEXURE XI.
C. ROOT CAUSE ANALYSIS: The root cause and failure analysis of the sterility test
failure was done using Fish bone method of analysis and Brain storming.
 FIGURE 1

Material Manpower Method

Contaminated
intermediate Poor Hygiene Sterilization Procedure
component
Production
Not followed
Storage Operators
Condition
Inadequate
Training
Sampling Error
Raw Materials
Dirty Inner
prone to
& Shipper EO Production
contamination Incorrect
Cartons Operators Operators
Sampling Method
Ineffective
Cleaning Agent

Sterility
Test
Poor Hygiene Failure
Humidity & Temperature
Underutilization of the (Inconsistent Compliance)
required quantity of
High Bioburden Ethylene Oxide for
complete sterilization
Non Compliance

Ineffective
Pressure Air
HVAC System
Differential Velocity
Hygiene
Test

Machine Measurement
Environment