Environmental Microbiology: An Interview with an
Microbiologics recently had
the opportunity to interview Dr. Edward Askew, the
founder of Askew Scientific Consulting, LLC. Edward has
been involved in the environmental industry for over 25
years. He became a laboratory supervisor for a
wastewater plant in 1995 for the city of Muscatine,
Iowa. Edward then became involved with the professional
organizations such as the Water Environment Federation (WEF)
and the American Water Works Association (AWWA). As a
committee member at the state and national level, Edward
has been involved with regulatory compliance with
federal regulation involving the Clean Water Act (CWA)
and Safe Drinking Water Act (SDWA). He is currently the
Part 4000 Coordinator for Standard Methods for the
Examination of Water and Wastewater and the chair of the
Water and Wastewater Community for AOAC International.
Why do we need regulatory
If we are to have clean water for drinking and a clean
environment, then there must be regulatory oversight.
Past history, such as the Cuyahoga River fires, caused
the public outrage that led to the formation of the
Environmental Protection Agency (EPA) and the CWA. But,
creation of federal regulations and agencies does not
assure protection for the public. Failure to have
meaningful regulatory oversight leads to complacency and
tragedies such as the recent case of E. coli
contamination of drinking water in Walkerton, Ontario.
Case History: The
Walkerton, Ontario O157:H7 strain of E. coli bacteria
In 2010, the public consumers of the drinking water were
exposed to E. coli O157:H7 and Campylobacter jejuni in
their drinking water for weeks. This tragedy was caused
by both the drinking water utility management's culture
of deceit and sloth and the regulatory agency's failure
to have meaningful oversight. Seven people died, and
more than 2,300 became ill. Some people, particularly
children, may be effected over their lifetime.
What controls needed to
be in place?
First of all, the regulators should have performed a
more in-depth audit of the sample collection practices
at the utility. Utility staff regularly falsified the
site the sample was collected due to the culture
promoted by the management. A review of utility records
and unannounced on-site observation with split samples
would have emphasized to the management that their
practices were under review.
Secondly, if the inspectors had looked into the chlorine
disinfection practices at the utility with emphasis on
the chlorine purchases, that would have led to the
discovery that the utility was not disinfecting the
drinking water as required. This discovery would have
lead regulators to discover that operators were
falsifying the chlorination/disinfection records.
What were the final outcomes
of the tragedy? The deaths and illnesses caused by the
utility management's actions have led to greater
emphasis on training of drinking water plant operators.
The regulators have reviewed their existing agencies and
have recommended increasing oversight with inspectors
that are technically proficient in the areas they
What are the major
federal and state regulatory rules that control
microbiological testing for water?
Current rules for Safe Drinking Water under either
Ground Water or Surface Water refer to 40 CFR Part 141
for the statutory analytical methods for chemical and
biological analytical methods. The Clean Water rules
refer to 40 CFR Part 136 for the statutory methods for
chemical and biological analytical methods.
How have they changed
over the last few years?
For microbiological methods, improvements such as the
sealable bacteria most probable number (MPN)
quantification trays used in enzyme substrate tests for
E. coli have allowed more laboratories to analyze their
water for this indicatory organism. Also, new EPA
methods for coliphages and rapid analysis utilizing
polymerase chain reaction (PCR) have expanded the
regulatory oversight of compliance with the SDWA and
What do you see as the major analytical concerns for
a water laboratory performing bacteria analysis?
Sample collection and preservation of microbiological
samples are key to producing accurate and defensible
results. The laboratory must have in place procedures
that meet the minimum requirements of the regulatory
method and they must actively audit these practices,
especially in the field.
Holding times must meet the
requirements of the regulations. The holding time begins
when the sample is collected and ends when the sample is
set-up for testing. Exceeding these holding times will
produce a biased result which most probably is an under
reported value. This result is termed to be a "false
negative" and places the laboratory in violation of
their regulatory requirements and can put the public at
What minimum quality
controls checks should a water laboratory have in place?
The dilution water should be checked regularly for
background bacteria that can produce false positive
results and chemical carryover from the laboratory
handling procedures that can produce false negative
The autoclave efficiency and the sterilization
procedures should be checked regularly. The use of
culture strips/ampules along with an autoclave
thermometer can determine the efficiency of the
autoclave. Also, samples of the equipment and media
sterilized should be checked for sterilization
Cleaning agents: Cross contamination by cleaning agents
used in the laboratory for glassware or countertops must
be monitored and eliminated. Chemical test such as the
Bromothymol Blue test will indicate cleaning agent
carry-over from the glassware washing procedures.
Media and Buffers:
All media and buffer preparation, storage and shelf-life
must be monitored and checked on a regular basis. The
use of positive bacteria controls for media and buffer
viability is key in preventing false negative results.
All temperatures should be monitored regularly and their
values recorded. Procedures need to be in place to
determine the accuracy of all temperature
Positive Controls and
All microbiological tests should have challenge
organisms to test whether the analysis will produce a
positive or negative result. These controls along with a
control blank should be run at a minimum with each
Initial and Ongoing
Demonstrations of Capability:
The analytical capabilities the analyst should be
determined on a continuous basis. The analyst's initial
demonstration of capability should be determined prior
to any results being reported and the analyst's ongoing
demonstration of capability will provide oversight of
Duplicate reproducibility measured by the Relative
Percent Difference will determine the procedure and
The dilution water or buffer solution used by the
analyst in an analysis batch is analyzed as a unique
sample. A positive result would indicate a cross
Both Single Blind Performance Tests and Laboratory
Internal Controls are needed to establish the analyst's
capabilities and determine if the analysis procedure is
performing correctly. Organisms that will produce
positive or negative results are used.
Root Cause Analysis:
All quality control checks must have limits that must be
met if the analysis is considered to be "In Control". If
the limits are exceeded, then the analysis is "Out of
Control" and a root cause analysis must be performed.
The laboratory must have a procedure in place outlining
the steps that must be taken to perform the analysis and
correct the problem.
What will the future hold
for microbiological analysis in a water laboratory?
In the near future, the laboratory will have to address
the quality control requirements for PCR analysis.
Different routes for cross contamination of water
samples need to be identified and prevented. Also, the
challenge of equating PCR results to the current
membrane filtration/most probable number tests will
require the laboratory to educate their clients. On the
horizon are the emerging pollutants such as coliphage
enumeration for the confirmation of four-log removal
under the EPA Ground Water Rule and the move from
indicator tests, such as the current E. coli analysis,
to pathogen-specific tests. All in all, the future will
be both interesting and demanding for the lab.
Thank you Mr. Askew
for sharing your expertise with the Microbiologics®
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