Public Health
Inspector’s
Guide
To the Principles and Practices
of Environmental Microbiology
FEBRUARY 2013
Public Health Ontario
Public Health Ontario (PHO) is a Crown corporation
dedicated to protecting and promoting the health of all
Ontarians and reducing inequities in health. As a hub
organization, PHO links public health practitioners, frontline health workers and researchers to the best scientific
intelligence and knowledge from around the world.
Our mission is to support health care providers, the public
health system and partner ministries in making informed
decisions and taking informed action. PHO provides
transparent and timely expert scientific advice, technical
support and practical tools related to infection prevention
and control; surveillance and epidemiology; health
promotion, chronic disease and injury prevention;
environmental and occupational health; health emergency
preparedness; public health laboratory services; research;
professional development; and knowledge services.
For more information, visit www.oahpp.ca
Ontario Agency for Health Protection and Promotion (Public Health
Ontario). Public health inspector’s guide to the principles and practices of
environmental microbiology. 4th ed. Toronto, ON: Queen’s Printer for
Ontario; 2013.
ISBN 978-1-4606-0678-0 [PDF]
ISBN 978-1-4606-0677-3 [Print]
All or part of this document may be reproduced for educational purposes
only without permission.
© Queen’s Printer for Ontario, 2013
Preface
This revised edition of the Public Health Inspector's Guide to
the Principles and Practices of Environmental Microbiology
was developed by the Public Health Ontario Laboratories
(PHOL) to assist Ontario’s local boards of health staff in the
informed use of public health laboratory services for
environmental microbiological testing. The last edition of
this guide was published in 2010.
This Guide is divided into several sections. It includes a
contact directory for the PHOL, general information
including an overview of analyses available at the PHOL,
legal use of the guide, and details regarding the collection of
legal samples. Subsections devoted to food, water,
suspected sewage contamination, and environmental
Legionella testing offer details of test menu, test requisition
requirements, and collection and transportation
instructions. Detailed interpretation of PHOL environmental
test results is provided in table 17.
This edition was made possible through extensive
stakeholder participation, with distribution and engagement
of all thirty six local boards of health, the Association of
Supervisors of Public Health Inspectors of Ontario (ASPHIO),
Canadian Institute of Public Health Inspectors (CIPHI), the
Ministry of Health and Long Term Care – Public Health
Policy and Programs Branch (MOHLTC-PHPPB), and the
Ontario Multi-Agency Foodborne Outbreak/ Food Recall
Working Group laboratory subcommittee. The success of
future editions is dependant of this ongoing engagement
and generosity of all who are involved in the delivery of
environmental health programs and services in Ontario.
This guide will be formally revised on a biannual basis.
Smaller changes as tests are modified, deleted or added,
will be updated in the online version. Please refer to
www.oahpp.ca for the most recent edition. We welcome all
suggestions and input for future editions, and look forward
to ongoing collaborations with all of those involved in public
health in Ontario.
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iii
Note To Readers/Users
The Public Health Inspector’s Guide to the Principles and
Practices of Environmental Microbiology (the “Guide”) is
proprietary to Public Health Ontario (“PHO” or “Ontario
Agency of Health Protection and Promotion/OAHPP"). No
part of the Guide shall be used, reproduced, translated,
converted, adapted, stored in a retrieval system,
communicated or transmitted by any means, for any
commercial purpose, including without limitation, sale,
resale, license, rental or lease.
PHO does not make any representations, warranties or
guarantees express or implied, as to the accuracy or
completeness of the Guide. Users must be aware that
updates and amendments will be made from time to time
to the Guide. It is the responsibility of the users to
determine whether there have been any such updates or
amendments and PHO shall have no responsibility for
providing notice of any such updates or amendments.
Although the material in the Guide has been carefully
prepared, neither PHO nor any of its directors, officers,
employees or agents shall accept legal responsibility in any
manner whatsoever for the contents and/or the improper
use of this Guide.
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Table of Contents
Contact Directory
2
General information
Scope of the Guide
Analyses Not Available at the PHOL
Legal Use of the Guide
Legal Samples – Collection Instructions
3
3
3
4
fooD samPle analysis
Food – Microbiological Analysis Test Menu
Food – Test Requisition Requirements
Food – Collection and Transportation Instructions
Food – Laboratory Rejection Criteria
Environmental Swabs for Outbreak Investigations
– Collection and Transportation Instructions
Water samPle analysis
Water – Microbiological Analysis Test Menu
Water – Test Requisition Requirements
Water – Collection and Transportation Instructions
Drinking Water – Official Agency Submission
Drinking Water – Private Citizen Submission
Ice – Treated
Recreational Water Facilities and Hydrotherapy Pools
Public Beaches
Water – Laboratory Rejection Criteria
susPecteD seWaGe contamination samPle
analysis
Collection and Transportation Instructions
LegioneLLa investiGations
Legionella – Microbiological Analysis Test Menu
Legionella – Selection of Sampling Sites
Legionella – Environmental Sample Collection and
Transportation Instructions
5
6
7
10
11
12
14
16
16
17
18
19
20
21
22
24
24
Glossary – Water terms
27
interPretation of environmental
microbioloGical test results
32
ontario Public HealtH stanDarDs anD
suPPortinG leGislation
36
suPPortinG Documentation/references
37
contributors
38
aPPenDices
Appendix A: Supplies
Appendix B: Laboratory Submission Forms
A. Bacteriological Analysis of Water – Single
Sample Requisition for Official Agencies
B. Bacteriological Analysis of Water – Multiple
Sample Requisition for Official Agencies
C. Bacteriological Analysis of Drinking Water
for Private Citizen, SINGLE HOUSEHOLD ONLY
D. Food Bacteriology
E. Environmental Bacteriology Swab Tests
F. Potability of Water Release Form
G. Autorisation De Divulguer Des Renseignements
Sur La Potabilité De L'eau
H. Ontario Agency for Health Protection and
Promotion Legal Seal
I.Ontario Agency for Health Protection and
Promotion Regulated Water Seal
Appendix C: Major Foodborne Diseases – General
Features
Appendix D: List of Tables
39
40
41
42
43
44
45
46
47
48
48
49
53
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Contact Directory
customer service
For inquiries, call our customer service centre at 1-877-604-4567 or 416-235-6556 between 7:30 a.m. and 7:00 p.m. or the
After-hours Emergency Response Line: 416-605-3113. Visit the PHO website for more information:
www.oahpp.ca
PHo laboratory, toronto
telephone #
fax #
416-235-6556 or
1-877-604-4567
416-235-6552
81 Resources Road, Etobicoke, ON M9P 3T1
Hours of operation:
toll-free Helpline:
Mon. to Fri. 8:00 a.m – 5:00 p.m
Mon. to Fri. 7:30 a.m – 7:00 p.m
after-hours emergency response line:
416-605-3113
Key contacts:
Medical Microbiologist
Regional Microbiologist
Manager, Enteric, Environmental, Molecular Surveillance
and Sexually Transmitted Infections
Head Technologist, Environmental Microbiology
Environmental Microbiology - Laboratory
Program Coordinator Drinking Water Testing
Clinical and Environmental Microbiology Office
Manager, Clinical and Environmental Microbiology
PHo laboratories, reGional
Hours of operation:
Mon. to Fri., 8:00 a.m – 5:00 p.m
after-hours
emergency response line:
416-235-6063
613-547-1185
416-235-5707
416-235-5718
416-235-5716
519-455-9310
416-235-5712
416-235-5988
416-235-5951
416-235-5951
416-235-5951
519-455-3363
416-235-5951
416-235-5951
telephone #
fax #
416-605-3113
Hamilton
250 Fennell Avenue West, P.O. Box 2100, Hamilton ON L8N 3R5
905-385-5379
905-385-0083
Kingston
181 Barrie Street, P.O. Box 240, Kingston ON K7L 4V8
613-548-6630
613 547-1185
519-455-9310
519-455-3363
orillia
750 Memorial Avenue, P.O. Box 600, Orillia ON L3V 6K5
705-325-7449
705-329-6001
ottawa
2380 St. Laurent Boulevard, Ottawa ON K1G 6C4
613-736-6800
613-736-6820
Peterborough
99 Hospital Drive, P.O. Box 265, Peterborough ON K9J 6Y8
705-743-6811
705-745-1257
sault ste. marie
160 McDougald Street, Sault Ste. Marie ON P6A 3A8
705-254-7132
705-945-6873
sudbury
1300 Paris Street, Suite 2, Sudbury ON P3E 6H3
705-564-6917
705-564-6918
thunder bay
336 South Syndicate Avenue, Thunder Bay ON P7E 1E3
807-622-6449
807-622-5423
timmins
67 Wilson Avenue, Timmins ON P4N 2S5
705-267-6633
705-360-2006
london
St. Joseph’s Regional Mental Health Centre, 850 Highbury Avenue,
5th Floor, P.O. Box 5704, Terminal ‘A’, London ON N6A 4L6
2
416-235-5806
613-548-6630
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General Information
scoPe of tHe GuiDe:
The Guide has been developed to provide Ontario's public
health inspectors (PHIs) with a resource that outlines PHO
Laboratories environmental microbiological testing services.
This document includes information to outline the:
•
•
•
•
Tests performed
Specimen collection and transportation instructions
Laboratory acceptance and rejection criteria
Test result interpretation
Environmental testing available through the PHOL includes:
• Microbiological analysis of ready-to-eat foods
• Environmental swab analysis for pathogen specific
requests (outbreaks)
• Drinking water analysis for private citizens (total Coliform
and Escherichia coli only)
• Drinking water analysis requested by boards of health
(microbiological analysis only)
• Treated ice
• Recreational water facilities and hydrotherapy pools
• Public beach water testing
• Suspected sewage contamination samples
• Environmental sampling for Legionella species
• Identification of ticks removed from humans
(Parasitology)
Public Health Ontario Laboratories hold a drinking water
testing license (Ministry of the Environment) and drinking
water tests, which are limited to the scope of testing
(http://www.cala.ca/cala_directories.html), are accredited
by the Canadian Association for Laboratory Accreditation
Inc. (CALA) to ISO 17025.The PHOL uses references and
methods as guidance documents for testing from agencies
such as Health Canada (HC) references, American Public
Health Association (APHA) and the Association of Official
Analytical Chemists (AOAC).
analyses not available at tHe PHol
The PHOL do not test environmental samples for the
following parameters:
•
•
•
•
•
•
•
•
Algal toxin testing (Blue-green algae)
Chemical/ radiological analysis
Foreign objects
Fungal testing
Live infestations
Parasites and ticks from non-human sources
Virus detection in food and water
Giardia and Cryptosporidium in water
Board of health staff are encouraged to contact the
customer service centre at 1-877-604-4567 or 416-235-6556
for any further questions. Boards of health can also use
other resources in the province to assist with investigations
requiring these tests. Other resources available include:
• Botulism Reference Service, Health Canada:
(613) 957-0902 or After-hours: (613) 296-1139
• Canadian Association for Laboratory Accreditation (CALA):
www.cala.ca
– for a list of laboratories accredited with CALA in Ontario
• Canadian Food Inspection Agency (CFIA):
www.inspection.gc.ca
• Ministry of Health and Long Term Care – Public Health
Protection and Prevention Branch (MOHLTC-PHPPB):
environmental.health@ontario.ca
• Ministry of the Environment (MOE):
www.ene.gov.on.ca
• Ontario Ministry of Agriculture, Food, and Rural Affairs
(OMAFRA):
www.omafra.gov.on.ca
• Standards Council of Canada (SCC):
www.scc.ca
– for a list of laboratories accredited with SCC in Ontario
• University of Guelph Laboratory Services:
www.guelphlabservices.com
• Canadian Cooperative Wildlife Health Centre:
www.ccwhc.ca
leGal use of tHe GuiDe:
The microbial analysis and acceptable limits outlined for the
various parameters and sample type/source are based on
current standards where available and/or guidelines based
on current scientific evidence. References have been
included to assist the user in obtaining further information.
These guidelines are intended to serve as a general
statement regarding the environmental microbiological
testing services available at PHOL and are of no legal force
or effect. If a local board of health is considering submitting
environmental samples as part of an investigation with
overlapping jurisdictions (e.g. federally or provincially
inspected food plant combined with retail food outlet), and
there is a discrepancy between federal and/or provincial
microbial standards/acceptable limits, the local board of
health should consult with its legal counsel.
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leGal samPles – collection instructions:
According to the PHOL Quality Policy for Legal Specimens, a
Legal Sample is defined as:
Any laboratory specimen that has been submitted for testing
to the PHOL with an intact legal seal and whose chain of
custody has been maintained.
Chain of custody is defined as:
An unbroken documented chain of events from the time of
sample collection until it is processed and completed.
The PHOL must receive prior notification that a Legal
Sample has been collected and will be submitted.
Maintaining a proper chain of custody from collection of the
sample through transport, testing and reporting ensures the
integrity of the Legal Sample and that the laboratory results
will stand as valid evidence in a legal case.
If chain of custody is not maintained, the integrity of the
Legal Sample, or the test procedures performed may be
called into question, possibly rendering the laboratory
results inadmissible as evidence.
Therefore, it is essential to ensure that Legal Samples are
handled according to proper guidelines and adhere to the
following instructions:
• Notify the Medical Microbiologist, Laboratory Manager or
Designate in advance when Legal Samples are to be
submitted (refer to the Contact Directory).
• Affix and secure an official PHO and /or a law enforcement
agency legal seal to the Legal Sample such that the
sample cannot be opened without breaking the seal
(N.B. For details on how to obtain official legal seal, please
see appendix A.) The official legal seal must be initialed
and dated by the individual who has affixed the legal seal.
The sealed sample must be placed in an appropriate
tamper evident container.
• Once the Legal Sample is received in the laboratory, a
Chain of Custody form must be signed by the individual
delivering the samples and the legal seal verified as intact
by both the individuals delivering and receiving the
package. For Legal water samples, refer to Public Health
Ontario’s Instructions for Official Agencies Submitting
Water Samples to the Public Health Ontario Laboratories
(www.oahpp.ca).
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Food Sample Analysis
Food – Microbiological Analysis Test Menu
Tables 1 to 4 provide an overview of food testing performed
by the PHOL – Environmental Microbiology section, the
nature of each test, and the turn around time. Turn around
time is defined as the number of business days to complete
testing based on date of receipt in the laboratory. This turn
around time definition does not apply to tests that are onhold awaiting information to continue testing. Turn Around
Time (TAT) posted in this Guide are estimates for routine tests.
For some tests or at times of public health emergencies,
outbreaks or exigent circumstances, turn around times may
be longer than what is listed. PHO/OAHPP disclaims all
warranties whether express or implied in respect of the
information, including the TATs provided herein.
Table 5 provides a list of food related tests which the PHOL
refers to other laboratory facilities for testing.
Indicator bacteria testing is performed on processed, cooked,
ready-to-eat food items with the exception of heterotrophic
plate count which is not performed on raw foods (e.g., fruits
and vegetables) and cultured products (e.g., cheese).
Pathogen specific analyses are determined based on clinical
information obtained from submitters, and the food type.
Provision of clinical information on the test requisition is
critical in order for laboratories to provide the most
appropriate testing for outbreak investigations. A final report
is not issued until all tests for a submission are complete.
table 1: indicator bacteria
test
nature of test
turn arounD time
Heterotrophic Plate Count
Bacterial culture
2 days
Total Coliform Count
Bacterial culture
2 days
Escherichia coli Count
Bacterial culture
3 - 5 days
Total Gram Negative Count
Bacterial culture
1 day
table 2: foodborne Pathogens
nature of test
turn arounD time
Bacillus cereus
test
Bacterial culture
2 – 4 days
Campylobacter jejuni
Bacterial culture
4 – 6 days
Clostridium perfringens
Bacterial culture
1 – 5 days
Escherichia coli O157:H7
Bacterial culture
2 – 9 days
Listeria monocytogenes
Bacterial culture
4 – 8 days
Salmonella species
Bacterial culture
4 – 6 days
Shigella species (not routinely performed)
Bacterial culture
2 – 6 days
Staphylococcus aureus
Bacterial culture
Detection of bacterial toxins
2 – 4 days
2 days
Vibrio species
Bacterial culture
2 – 7 days
Yersinia enterocolitica
Bacterial culture
12 – 14 days
nature of test
turn arounD time
pH
Hydrogen ion measurement
with ion selective electrode
1 day
Phosphatase
Measurement of alkaline phosphatase
in dairy products by fluorometric assay
1 - 2 days
Water Activity – aw
Measurement of unbound water in food
by chilled-mirror dewpoint technique
1 day
table 3: Physiochemical tests
test
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table 4: molecular typing
test
Pulsed field gel electrophoresis (PFGE)
nature of test
turn arounD time
Molecular typing
3 – 5 days
table 5: referred tests
test
referral laboratory
Clostridium botulinum
Testing performed at:
Botulism Reference Service Health Canada
Room D457, Sir Frederick G. Banting Building, Building 22,
Tunney’s Pasture, PL2204E
251 Sir Frederick Banting Driveway
Ottawa, ON K1A 0K9
Telephone: (613) 957-0902
Cryptosporidium
Cyclospora
Giardia
Hepatitis A
Norovirus
Prion disease
Toxoplasma
Trichinella
The tests listed may be referred to other testing facilities. Please contact the
Toronto PHOL Environmental Laboratory at 416.235.5716.
Food – Test Requisition Requirements
The PHOL Food Bacteriology requisition (see Appendix B)
must be used to submit foods to the PHOL for analysis. It is
important to provide as much of the information requested.
submitter iDentification
Information required includes the local board of health
office address and the name, phone and fax number of the
PHI or other staff submitting the samples.
outbreaK number
An outbreak number should be assigned for all outbreak
related specimens. In some cases, the outbreak number is
the only way the laboratory can correlate results from
multiple submissions of foods and other related specimens
from a single outbreak.
PurPose of collection: (Mark the appropriate box)
• Routine surveillance: Regular, on-going sanitary
monitoring programs of food prepared/served in retail
food establishments. Samples collected in the absence of
foodborne disease.
• HaccP (Hazard analysis critical control Point): Food
collected in association with evaluating or monitoring
critical control points in food preparation, production and
processing.
6
• special survey: Special investigations of specific food items
conducted by local boards of health. Surveys are usually
negotiated with the laboratory prior to collection and
analysis may be altered to accommodate a specific need.
• complaint samples: Samples submitted for laboratory
evaluation associated with the investigation of foodborne
illness. Complaints usually come from the general public
to the local board of health office.
fooDborne illness: (Mark the appropriate box)
• Part of meal: Foods submitted in conjunction with a
suspect foodborne illness incident which are part of the
actual meal implicated in the outbreak.
• control samples: Foods submitted in conjunction with a
suspected foodborne illness incident which were not part
of the suspect meal, but which are of the same type
and/or prepared and stored under similar conditions as
those actually consumed.
• follow-up samples: Samples taken after remedial action to
determine whether or not the problem has been resolved.
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ePiDemioloGical information:
• Provide the predominant symptoms of illness in the space
provided.
• Provide the time and date of onset of symptoms.
• Provide the confirmed or suspected etiological agent
(if suspect indicate as such on requisition).
• Provide the time and date the suspect meal was
consumed.
• Provide the location(s) the suspect meal was prepared
and consumed.
samPle DescriPtion/iDentification:
• Samples must be identified by their common name on the
requisition in the column labeled Sample Description (e.g.
“Roast Beef”).
• In addition, a unique identifier must be present on the
sample bag and on the requisition under the column
labeled Sender’s Number. The number must uniquely
identify the food submission to assist tracking of samples
during outbreaks and is designated at the discretion of
the submitter. Duplication of numbers should be avoided
to prevent mismatching of samples.
• The marking must be sufficiently unique that the sample
cannot be confused with other similar samples in the
laboratory.
• Detailed information will facilitate food recalls should a
product be contaminated. The following information
should be collected when available (a photo or photocopy
of the product label would facilitate this data gathering
process):
o
o
o
o
o
o
o
o
o
o
o
o
o
o
o
Brand name
Common name
Unit size (e.g. 50g or 125 mL, etc.)
Lot number(s)
Dates: Best before/Expiry/Use by/Durable life/
Packed on/Manufacturing
Location of purchase
Universal product code (UPC)
Name of Manufacturer/Importer/Legal Agent (if
available on the packaging)
Label claims/preparation or serving instructions
Container type (e.g. vacuum pack, plastic rigid
container, etc.)
Shelf life of product (if possible)
Where the sample was taken (e.g. at the suspect place
of eating or the case's home)
Storage information (e.g. frozen, refrigerated, nonrefrigerated etc.)
Indicate if this is an opened or unopened sample
Indicate the number of samples/sub-samples taken
Food – Collection and Transportation Instructions
Table 6 provides information and sampling instructions for food.
table 6: food – collection and transportation instructions
Epidemiological and Risk Based Prioritization of Food Sample Collection in Foodborne
Outbreaks1
instructions
Precautions
Selection of food items for collection and submission in a foodborne
outbreak should be based on all available epidemiological and
laboratory information related to the outbreak.
• Epidemiological analyses, such as the calculation of attack rate,
will facilitate prioritization of those food items that are most
likely to be involved.
• If attack rates are unavailable, incubation period, clinical
syndrome and basic demographic information about related
cases should be used.
The integration of information about the clinical pathogen, if
known, should also facilitate prioritization of food items to be
collected and tested. Pathogen specific high risk foods are
presented in Appendix C.
• If risk stratification of food sources is not performed by the
submitting public health inspector, and there is an abundance
of specimens submitted for testing, prioritization will be
determined by the head technologist and medical
microbiologist of the PHOL Environmental Microbiology
section.
• The communication of all available clinical, epidemiological
microbiological information to the PHOL is of great importance
in guiding these decisions.
For full details please consult Foodborne disease outbreaks : guidelines for investigation and control. World Health Organization 2008.
http://whqlibdoc.who.int/publications/2008/9789241547222_eng.pdf Accessed February 14, 2010.
1
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table 6: food – collection and transportation instructions ...continued
Containers for Sampling and Submission of Foods
instructions
Precautions
Sterile plastic sample bags with round wire closure are the
preferred sample container for all food types.
Food samples found in the original container can be submitted in
the original packaging provided that the container is air-tight and
will not leak during transit.
• If sampling closed or intact samples, please consult with the
CFIA prior to submission.
If the original container has been opened or if the packaging is
fragile and may be damaged during transit, samples should be
transferred from the original container to a sterile plastic sample
bag with round wire closure.
• Styrofoam and box-board packaging used for fast-food
products are examples of containers that are not suitable for
bacteriological sampling. They are not air tight and may leak
causing cross contamination of samples and/ or external
contamination into the sample.
• These containers should not be put inside a sterile sample bag
with round wire closure since the outside surfaces will be
contaminated by handling and the sample may be
compromised in transit.
Preparation
As part of good aseptic technique, the time taken for the physical collection of the sample can be minimized by ensuring the following:
instructions
Precautions
Ensure all materials required are available.
Label sample bags before sample collection.
Investigate before sampling to determine plan of action that will
minimize sampling time.
Complete requisition(s) before or after, never during, sample
collection.
Collection of Sample(s)
Precautions
instructions
Use aseptic technique at all stages of sample collection.
• Failure to do so will compromise the laboratory results.
Collect samples that are representative of each item of food that is
being investigated.
• Submit a minimum of 200 grams of each sample if possible as
multiple tests may be required.
Meals that contain multiple discreet items may be collected
individually or combined for submission.
• Separation of meals or mixed foods must be performed at the
sampling stage. The laboratory will not separate components of
a sample received in the laboratory.
• If an individual component is of greater interest than other
components of a meal or mixed food, sample the items
individually and they will be tested as individual samples.
• If collected as a single sample, components should be collected
in approximately equal amounts and will be tested as one
sample.
Foods that contain multiple items (e.g. meat and gravy, sandwiches)
may be collected as a single sample.
The mouths of sterile plastic sample bags with round wire closures
should be rolled down several times and the wire ends twisted
together tightly to ensure and prevent leaking during transport.
8
• Leaking samples will be rejected by the laboratory.
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table 6: food – collection and transportation instructions ...continued
Large Lots of Food
Most commercially prepared foods are produced in very large lots.
instructions
Where possible, mix the lot of food before sampling.
Precautions
• If you encounter a lot of food that is larger than 2 kilograms,
take five sub-samples of 200 grams each and submit for
individual analysis.
• Ensure sub-samples are clearly and distinctly labeled with a
unique identifier for tracking.
Small Quantity Samples
Situations may be encountered where there is very little sample left.
instructions
In food poisoning cases, small samples will be processed by the
laboratory, but complete analysis may not be possible.
Precautions
• If you suspect, or have a confirmed etiological agent, indicate
the information on the requisition form and the specific
analysis will be given priority.
• In the absence of such information, the laboratory will exercise
judgment and will perform analyses that will most likely
provide useful information.
Sample Storage
instructions
Precautions
It is preferable to deliver samples to the laboratory as soon as
possible after collection.
• If there is a delay, samples must be stored in a secure location
where they cannot be tampered with or confused with other
food.
Shelf stable foods should be stored in a dry location protected from
light and extreme temperatures.
• Temperatures outside 15°C to 30°C may compromise the
sample(s) and impact the quality of laboratory results.
Frozen foods should be stored as close to the temperature at which
they were found as possible.
• Temperatures above freezing may result in thawing and impact
the quality of laboratory results.
Perishable foods should be stored at 2 to 4°C.
• Temperatures outside of this range may impact the quality of
laboratory results.
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table 6: food – collection and transportation instructions ...continued
Shipping Containers
instructions
Precautions
Ship all foods in containers with hard walls and lids secured in the
closed position. Shipping containers must be labeled with the
submitting organization, unique identifier and contents e.g. FOOD
SAMPLES on the outside of the container.
Shipping containers used for food samples should be dedicated to
food samples and not be used for other types of environmental
samples.
• Samples must be made secure inside the shipping container so
that there is no physical damage to the samples that may cause
leakage.
• Shipping containers should be decontaminated between uses
with an appropriate disinfectant and used according to the
manufacturer’s instructions.
Ship dry foods and other shelf stable foods at ambient temperature
in a closed container.
• Temperatures outside 15°C to 30°C may compromise the
sample(s) and impact the quality of laboratory results.
Ship frozen foods to the laboratory in an insulated container with
sufficient ice packs to maintain the frozen state.
• Temperatures above freezing may result in thawing and impact
the quality of laboratory results.
Ship perishable foods in insulated containers with sufficient cold
packs to maintain a temperature as close to 4°C as possible. If ice is
used, contain the ice in a manner that does not allow water contact
with the samples.
• Temperatures outside of this range may impact the quality of
laboratory results.
Transit Time
instructions
Precautions
Submit food samples to the laboratory as soon as possible or within
48 hours.
Samples collected during foodborne disease investigations that are
part of the implicated meal will not be rejected after 48 hours.
These samples are often the only opportunity to determine the
cause of an outbreak.
• Shipping time must be taken into consideration when
transporting samples.
• Routine Surveillance /HACCP samples older than 48 hours will
be rejected.
• Transit time needs to be considered in result interpretation as
prolonged transit time may compromise test results.
Food – Laboratory Rejection Criteria
Table 7 outlines the various reasons a food sample will be rejected by the PHOL. If rejected, a new sample must be submitted
with a completed requisition.
table 7: food – laboratory rejection criteria
DescriPtion
10
reason for rejection
Sample Type
• Specimen received from an unauthorized submitter e.g. private
citizen, or no board of health identifier
Sample Requisition
• Requisition received with incomplete information e.g. unique
identifiers missing from requisition, sample collection bag or
both
• Requisition received separated from the sample collection bag
with no unique identifier to link them
Sample Collection
• Inappropriate sampling container
• Raw foods requiring processing or cooking before consumption
unless linked to a specific outbreak and discussed with the
medical microbiologist prior to submission
• Insufficient amount of sample submitted
Sample Transport
•Sample leaked in transit
• Sample received with evidence of temperature abuse in transit
P ublic Healt H insP ec t o r ’s G uiD e t o t He P r inc iP les anD P r ac t ic es o f env ir o nm ent al m ic ro bio lo G y
Environmental Swabs for Foodborne Outbreak Investigations –
Collection and Transportation Instructions
Table 8 provides sampling instructions for environmental swabs. These instructions are limited to the use of swabs during
foodborne outbreak investigation.
Environmental swabs may be processed during foodborne outbreak investigations for the presence or absence of a specified
bacterial pathogen from a physical environment (e.g. cutting board, meat slicer etc.). These are available (for outbreaks and
foodborne illness(es) investigation only) from the Toronto Public Health Laboratory at 416-235-5716.
The environmental swabs available from the PHOL contain neutralizing solution as transport medium. This medium can
neutralize the following sanitizers: iodophores including chlorine compounds, quaternary ammonium compounds and
phenolic sanitizers.
table 8: sampling instruction for environmental swabs in foodborne outbreak investigations
samPlinG instructions
Precautions
1. Open the sterile swab container, grasp the handle end of the
swab being careful not to touch any portion that might be
inserted into the vial, and remove the swab aseptically.
2. Moisten the swab head and press out the excess solution against
the interior wall of the vial with a rotating motion.
3. Hold the swab handle to make a 30° angle contact with the
surface. Rub the swab head slowly and thoroughly over
approximately 50 cm2 of the surface three times, reversing
direction between strokes.
• If the sample area is large, multiple sections may be swabbed
with one swab.
• For an investigation where sampling areas are smaller and
physically different, a new swab should be used for each section.
4. After the area has been swabbed, using aseptic technique, return
the swab to the vial.
• Ensure the vial is closed tightly to prevent leakage and specimen
rejection.
5. Complete all sections of the Environmental Bacteriology Swab
Tests requisition (see Appendix B) with appropriate information
to ensure accurate tracking of the sampling.
6. Swabs should be stored and transported to the lab at 2 to 6°C
and optimally tested within 24 hours of collection.
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11
Water Sample Analysis
Water – Microbiological Analysis Test Menu
The Health Protection and Promotion Act, section 41 (4)
provides public health inspectors and medical officers of
health the ability to make, take and remove or require the
making, taking and removal of copies, samples or extracts
related to an examination, investigation, test or inquiry.
Sampling of drinking water may be required as part of an
outbreak investigation, a response to a report of a health
hazard or part of a routine inspection to ensure drinking
water is potable.
Boards of health staff are discouraged from collecting
routine samples from regulated drinking water systems as
this testing is performed by accredited and licensed
commercial laboratories on a regular basis.
All drinking water must be tested within 48 hours of
collection while non potable samples are tested within one
calendar day of collection. Samples should be kept cool and
delivered to the laboratory as soon as possible.
For samples requiring analysis for a bacteriological healthrelated parameter not included in Ontario Drinking Water
Quality Standards (Ontario Regulation 169/03) e.g.
Heterotrophic Plate Count (HPC) or Background, and
submitted under a Provincial Officer’s Order or Certificate of
Approval from a regulated drinking water system, please
contact the Program Coordinator Drinking Water Testing for
more information (refer to the Contact Directory).
12
If boards of health personnel wish to engage in a study
involving drinking water (e.g. to identify trends associated
with ground water contamination in various geographic
areas, etc.), the Program Coordinator, Drinking Water
Testing must be notified prior to sampling. Refer to section
5 of O. Reg 248/03 Drinking Water Testing Services
regulation for specific requirements associated with
drinking water research.
Table 9 provides an overview of water testing performed at
the PHOL for various types of water samples.
The table also identifies the type of sample container
required for submission and the laboratory turn around
time (TAT) for each test performed. Turn around time is
defined as the number of business days to complete testing
based on date of receipt in the laboratory. This turn around
time definition does not apply to tests that are on-hold
awaiting information to continue testing. Turn Around
Times posted in this Guide are estimates for routine tests.
For some tests or at times of public health emergencies,
outbreaks or exigent circumstances, turn around times may
be longer than what is listed.
PHO/OAHPP disclaims all warranties whether express or
implied in respect of the information, including the TATs
provided herein.
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table 9: Water – microbiological analysis test menu
samPle DescriPtion
samPle container
tests PerformeD
turn arounD time
Bottled Water *
PHOL water sample bottle – 200mL
Total coliform
Escherichia coli
Pseudomonas aeruginosa
Staphylococcus aureus
Heterotrophic plate count
2 – 4 days
2 – 4 days
2 – 5 days
2 – 6 days
2 – 5 days
Ice – Treated
Sterile plastic bag with round wire
closure
Total coliform
Escherichia coli
Heterotrophic plate count
(HPC performed on request for
treated water only)
2 – 4 days
2 – 4 days
2 – 5 days
Official Agency
- Drinking Water Analysis **
PHOL water sample bottle – 200mL
Total coliform
Escherichia coli
Heterotrophic plate count
(HPC performed on request for
treated water only)
2 – 4 days
2 – 4 days
2 – 5 days
Outbreak samples ***
Dependent on type of analysis
required
(Contact the PHOL, Toronto for more
information)
Dependent on type of analysis
required
(Contact the PHOL, Toronto for
more information)
Dependent on type
of analysis required
(Contact the PHOL,
Toronto for more
information)
Private Citizen
– Drinking Water Analysis
PHOL water sample bottle – 200mL
Total coliform
Escherichia coli
2 – 4 days
2 – 4 days
Public Beaches
PHOL water sample bottle – 200mL
Escherichia coli
2 – 4 days
Recreational Water Facilities
and Hydrotherapy Pools
PHOL water sample bottle – 200mL
Total coliform
Escherichia coli
Pseudomonas aeruginosa
Staphylococcus aureus
Heterotrophic plate count
2 – 4 days
2 – 4 days
2 – 5 days
2 – 6 days
2 – 5 days
Specific Etiological Agents
Refer to Table 10
Dependent on type of analysis
required
(Contact the PHOL, Toronto for more
information)
Dependent on type of analysis
required
(Contact the PHOL, Toronto for
more information)
Dependent on type
of analysis required
(Contact the PHOL,
Toronto for more
information)
Suspected Sewage
Contamination
PHOL water sample bottle – 200mL
Escherichia coli
2 – 4 days
NOTES:
* Commercial sealed products are not accepted. Contact the Canadian Food Inspection Agency if analysis of a sealed product is required.
** For samples submitted under an Ontario drinking water regulation, laboratories have an obligation to report test results as outlined in section 18 of the
Safe Drinking Water Act, 2002. The accompanying requisition must have all required information completed prior to receipt at the laboratory or the sample
will not be tested and a new sample and completed requisition submitted.
*** A Public Health Inspector or designate under the authority of the Medical Officer of Health may collect water samples from drinking water systems under
the Ontario drinking water regulations if required for investigational purposes in outbreak situations. These samples must be identified as “Non-Regulated”
with “Outbreak Investigation” identified on the requisition. These samples are not governed by the Safe Drinking Water Act requirements. The Ontario
Drinking Water Quality Standards do not apply and there is no requirement for testing to be performed by a licensed / accredited method. PHOL reporting
requirements apply.
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Table 10 provides a list of additional analyses which may be performed in outbreak situations or upon special request.
These MUST be submitted as NON-REGULATED requests only.
table 10: Water – microbiological analysis test menu – Pathogen specific analyses
samPle container
tests PerformeD
turn arounD time
Campylobacter species
PHOL water sample bottle – 200mL
Bacterial culture
4 – 6 days
Escherichia coli O157:H7
PHOL water sample bottle – 200mL
Bacterial culture
2 – 9 days
Legionella species
PHOL water sample bottle – 200mL
Potable – 1000mL (5X200mL)
Non-potable – 200mL
Bacterial culture
10 – 15 days
Pseudomonas aeruginosa
PHOL water sample bottle – 200mL
Bacterial culture
2 – 5 days
Salmonella species
PHOL water sample bottle – 200mL
Bacterial culture
4 – 6 days
Staphylococcus aureus
PHOL water sample bottle – 200mL
Bacterial culture
2 – 6 days
Vibrio cholerae
PHOL water sample bottle – 200mL
Bacterial culture
2 – 7 days
Yersinia species
PHOL water sample bottle – 200mL
Bacterial culture
12 – 14 days
analysis
Water – Test Requisition Requirements
Table 11 provides direction on the completion of the Official
Agency “Bacteriological Analysis of Water – Single Sample”
and “Bacteriological Analysis of Water – Multiple Sample”
requisitions. For detailed instructions related to the
completion of these forms and reporting requirements, refer
to Public Health Ontario’s Instructions For Official Agencies
Submitting Water Samples to the Public Health Ontario
Laboratories (http://www.oahpp.ca/services/watertesting.html).
When submitting samples for analysis to the PHO Laboratories
it is important that all samples submitted under O.Reg.319/08
and O.Reg.170/03 are properly identified so that the
appropriate reporting and licensing requirements can be met.
For general information refer to the Ministry of
Environment’s “Practices for the Collection & Handling Of
Drinking-Water Samples” (April 1, 2009 or as current)
http://www.ene.gov.on.ca/publications/4464e01.pdf
table 11: Water – test requisition requirements
Non Potable Water
samPle tyPe
samPle information –
non - Potable
(tick off)
rePortinG / testinG
requirements
comments
Public beach
Public beach
- PHO Reporting
Requirements
Public pools, spas, wading pools,
splash pads/spray pads, or water
slide receiving basins
Recreational water facility / hydrotherapy pool
- PHO Reporting
Requirements
Water suspected of being
contaminated with fecal material
Suspected sewage contamination
- PHO Reporting
Requirements
Samples must be
submitted in a
Sewage Sample
Collection Kit
Other non potable sources
Other
- PHO Reporting
Requirements
Indicate the
source
PHo = Public Health Ontario
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table 11: Water – test requisition requirements ...continued
Potable Water
samPle tyPe
tyPe of DrinKinG
Water systems
(tick off)
reason for
samPlinG
(tick off)
source of
DrinKinG
Water
(tick off)
aPPlicable
reGulations anD
rePortinG /
testinG
requirements
comments
Bottled water
Non Regulated
Applicable tick
box(es)
Bottled Water
- PHO Reporting
Requirements
Commercial sealed
products are not
accepted (contact CFIA)
Drinking water
system
O. Reg 170 / 03
under the SDWA
SDWA O. Reg 170/03
Applicable tick
box(es)
Applicable tick
box(es)
- O. Reg 170 / 03 and
O. Reg. 169/03 under
the SDWA
- O. Reg. 170/03
(upload exempt)
- ODWQS apply
- Licensed /
accredited methods
- All required fields of
the form must be
completed
- Unbroken chain of
custody required: Use
Regulated Water Seal or
complete “Relinquished
By” section on form
Drinking water
system
O. Reg 319 / 08
under the HPPA
HPPA
O. Reg 319/08
Applicable tick
box(es)
Applicable tick
box(es)
- O. Reg 319 / 08
under the HPPA
- O. Reg. 169/03
under the SDWA
- O. Reg. 319/08
(including upload to
LRMA)
- ODWQS apply
- Licensed /
accredited methods
- All required fields of
the form must be
completed
- Unbroken chain of
custody required: Use
Regulated Water Seal or
complete “Relinquished
By” section on form
Drinking water
systems that fall
outside Ontario
drinking water
regulations
HPPA Regulated
Premises
Or
Non Regulated
Applicable tick
box(es)
Applicable tick
box(es)
- O. Reg. 169/03
under the SDWA
- PHO Reporting
Requirements
- ODWQS apply
- Licensed /
accredited methods
Refer to Table A in
“Glossary Water Terms”
for definitions and
examples
Ice
Non Regulated
Applicable tick
box(es)
Other and
indicate “Ice” and
any applicable
tick box(es)
- PHO Reporting
Requirements
Treated ice only
Other potable
water sources
Non Regulated
Applicable tick
box(es)
Other
- PHO Reporting
Requirements
Indicate the source
Private well
Private Residence
Applicable tick
box(es)
Applicable tick
box(es)
- O. Reg. 169/03
under the SDWA
- PHO Reporting
Requirements
- ODWQS apply
- Licensed /
accredited methods
HPPa = Health Protection and Promotion Act
oDWqs = Ontario Drinking Water Quality Standards
PHo = Public Health Ontario
sDWa = Safe Drinking Water Act, 2002
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Water – Collection and Transportation Instructions
Table 12 provides information and sampling instructions for drinking water (both official agency and private citizens); ice
(treated); recreational water facilities and hydrotherapy pools and public beaches.
table 12: Water – collection and transportation instructions
Drinking Water – Official Agency Submissions
The PHOL tests drinking water samples for the microbiological indicators total coliforms and Escherichia coli. A heterotrophic plate count
(HPC) may be requested on a distribution system sample or other treated water if necessary. Refer to the Ministry of Health and LongTerm Care Drinking Water Protocol 2008 (or as current) for more information about drinking water protocols.
samPlinG instructions
Precautions
1. Complete the required information on the requisition.
The sample will not be tested if all of the required information is
not complete or accurate when received at the laboratory, and a
new sample and completed form will be required to be submitted.
2. Remove any aerator, screen or other attachment(s) from the
faucet. Alternatively, take a sample from an inside tap with no
aerator.
Do not take a sample from an outside faucet or the garden hose.
3. Disinfect the end of the faucet spout with an alcohol swab, or
diluted bleach solution (1 part household bleach to 10 parts
water), before running water to remove debris or bacteria.
Disinfecting the tap with a flame is not recommended because this
can damage faucets.
4. Turn on cold water and let it run for 2-3 minutes to remove
standing water from the plumbing system.
5. Wash hands with soap and water and dry them thoroughly prior
to taking the sample.
16
6. Examine the lid of the PHOL collection bottle.
If the tamper evident ring has separated from the cap use another
collection bottle.
7. Remove one of the barcodes on the bottle and apply it to the
top copy of the requisition in the “Barcode” field.
Samples will not be processed if the barcode is not affixed to the
top copy of the requisition. If the barcode is not able to be
removed, write the barcode number on the form.
8. Remove the sample bottle lid.
• Do not touch the inside of the lid
• Do not put the lid down
• Do not rinse out the bottle as this will remove the sodium
thiosulfate which neutralizes the chlorine residual before testing
in order to maintain bacterial viability during sample transport
to the laboratory.
• Do not ingest; for accidental exposure get medical attention.
9. Fill bottle to the 200mL fill line. Close the lid firmly.
If overfilled, pour excess water out so there is just 200mL of water
in the bottle.
10.Refrigerate the water sample or keep it in a cooler with frozen
ice packs if it can not be delivered to the laboratory within six (6)
hours.
• Do not freeze the sample.
• Samples received at the laboratory above 25°C will not be
tested.
11.Keep the water sample cool (but not frozen) until it is delivered
to the drop-off location.
All drinking water must be tested within 48 hours of collection for
the results to be suitable for the purposes of the Safe Drinking
Water Act, 2002.
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table 12: Water – collection and transportation instructions ...continued
Drinking Water – Private Citizen Submission
The PHOL tests Private Citizen drinking water from a “single household” for the microbiological indicators total coliforms and Escherichia coli.
The information below outlines the requirements for submitting private citizen drinking water samples for testing by non official agency
clients. Private Citizens are required to use the PHO collection kit which includes a Bacteriological Analysis of Drinking Water for Private
Citizen, SINGLE HOUSEHOLD ONLY requisition and barcoded collection bottle when submitting samples for testing. For more information
refer to the Water Testing Information page at http://www.oahpp.ca/services/water-testing.html.
samPlinG instructions
Precautions
1. Remove screen or other attachment(s) from tap.
2. Disinfect the end of the faucet with an alcohol swab or dilute
bleach solution (1 part household bleach to 10 parts water).
3. Run the cold tap for 2 - 3 minutes before collecting sample.
4. Fill the PHO Public Health Laboratory water collection bottle to
the 200 mL fill-line (Maximum amount) and recap bottle
immediately.
• Do not rinse the bottle before filling. It contains a preservative
(i.e. 0.8mM Sodium Thiosulphate. Do not ingest. For accidental
exposure get medical attention).
• Do not touch inner surface of cap or bottle with anything other
than the water collected.
5. Keep the sample cold (e.g. refrigerated and do not freeze). Bring
it to the drop off location as soon as possible in a cooler with
frozen ice packs.
• Ensure the form is completely and accurately filled in. Samples
will not be tested if the form is not complete when received at
the laboratory, and a new sample and completed form will be
required to be submitted.
• Results are unreliable if the sample is improperly collected,
stored or transported.
• Samples received at the laboratory above 25°C will not be
tested.
• All drinking water must be tested within 48 hours of collection
for the results to be suitable for the purposes of the Safe
Drinking Water Act, 2002.
6. In most cases, samples are transported from the drop off
location (e.g. the local board of health office) by courier to the
PHO Public Health Laboratory
For laboratory locations refer to www.oahpp.ca. For local board of
health locations refer to http://www.health.gov.on.ca/
important information regarding Private citizen submissions:
requisition
• The “Bacteriological Analysis of Drinking Water for Private Citizen, SINGLE HOUSEHOLD ONLY” form is not to be used for drinking water
from a drinking water source under one of Ontario’s drinking water regulations. Owner / operators of these systems must use licensed
and accredited commercial laboratories for their routine testing.
• The form must be complete and accurate upon receipt at the PHOL or the sample will not be tested and another sample with another
completed form submitted.
• The PHOL will not release results to Private Citizen clients by fax or over the telephone.
sample
• Private Citizens wishing to submit water they use for drinking from surface water sources such as lakes, rivers or streams should be
advised not to submit them unless the water has been properly treated first.
• Three consecutive samples, taken 1 to 3 weeks apart, are needed to determine the stability of the water supply. If the well shows
acceptable total coliform / E. coli counts, then sample three times a year.
• Samples are analyzed for the indicators of bacterial contamination total coliform and E.coli. Samples are not tested for other
contaminants, including chemical contaminants, and therefore may be unsafe to drink even when there is no significant evidence of
bacterial contamination. Clients are directed to consult their local board of health for information on testing for other contaminants.
• If technical difficulties should arise, it may be necessary to refer samples to another licensed laboratory within the PHOL system.
Submission of a drinking water sample with the test requisition grants consent to the transfer of the sample if necessary.
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17
results
• Submitters are to indicate on the form how they wish to receive their report:
a) By mail
b) Picked up at the laboratory if indicated on the requisition. Photo identification is required when picking up the report. Anyone
wishing to pick up the report on behalf of the submitter must have a Potability of Water Release form signed by the submitter, and
show photo ID when picking up the report. Refer to Appendix B or http://www.oahpp.ca/services/water-testing.html. Alternately, a
Freedom of Information request may be submitted to the PHO Privacy Officer (refer to www.oahpp.ca).
c) By Interactive Voice Response (IVR). The sample results and interpretation are available by calling 1-877-723-3426 and keying in the
barcode associated with the sample. Teletypewriter (TTY): 1-866-828-2238.
• Private Citizen clients requiring assistance with the interpretation of results or advice on the correct course of action are to contact their
local board of health.
table 12: Water – collection and transportation instructions ...continued
Ice – Treated
The PHOL tests treated ice samples for the microbiological indicators total coliforms and Escherichia coli and Heterotrophic Plate Count.
samPlinG instructions
Precautions
1. Complete the required information on the requisition and label
sample bag(s) with unique identifier that matches that on the
requisition.
Ensure all materials required for sample collection are available
prior to collection – sterile plastic sample bags with round wire
closure, test requisition, sterile collection device (if required).
2. Wash hands with soap and water and dry them thoroughly prior
to taking the sample.
3. Collect representative ice samples using either the collection
device located in the freezer unit or another sterile collection
device.
Use aseptic technique at all stages of sample collection. Failure to
do so may compromise the laboratory results.
4. Ensure the sample bag is properly secured to prevent sample
leakage.
5 Transport sample(s) and accompanying requisition to the
laboratory with a freezer pack in the appropriate transport
container.
18
Samples must be received in the laboratory within 48 hours of
collection.
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table 12: Water – collection and transportation instructions ...continued
Recreational Water Facilities and Hydrotherapy Pools
The PHOL tests recreational water facilities and hydrotherapy pool samples for the microbiological indicators total coliforms, Escherichia
coli, Pseudomonas aeruginosa, Staphylococcus aureus and Heterotrophic Plate Count. Refer to the Ministry of Health and Long-Term Care
Recreational Water Protocol 2008 (or as current) for more information about recreational water facilities and hydrotherapy pools
protocols.
Microbiological sampling may be conducted to evaluate and monitor cleaning and sanitizing procedures and to ensure that a health hazard
does not exist. Note: for hydrotherapy pools/baths, samples are to be obtained from a sanitized tub after fresh water has circulated for a
minimum of two minutes.
samPlinG instructions
Precautions
1. Complete the required information on the requisition.
2. Examine the lid of the PHOL collection bottle
If the tamper evident ring has separated from the cap use another
bottle.
3. Remove one of the barcodes on the bottle and apply it to the
top copy of the requisition in the “Barcode” field.
Samples will not be processed if the barcode is not affixed to the
requisition. If the barcode is not able to be removed, write the
barcode number on the form.
4. Hold the bottom of the PHOL sample bottle and aseptically
remove the cap.
• Do not touch the inside of the lid
• Do not put the lid down
• Do not rinse out the bottle as this will remove the sodium
thiosulfate which neutralizes the chlorine/bromine residual
before testing in order to maintain bacterial viability during
sample transport to the laboratory.
• Do not ingest; for accidental exposure get medical attention.
5. Plunge the bottle, neck downward, at a 45 degree angle to 30
cm below the water surface. Fill the bottle at that depth with a
sweeping forward motion ahead of the hand, bringing the bottle
up toward the water’s surface.
• Take samples in an appropriate location of the pool in order to
provide an accurate representation of the water in the pool.
• Keep the bottle away from your other hand and body parts.
• Avoid contamination from floating debris.
6. If necessary, discard some water so that the bottle is only filled
with water to the 200 mL fill line indicated on the bottle.
Carefully put the cap back on the bottle.
7. Transport sample(s) to the laboratory with a frozen freezer pack
in the appropriate transport container as soon as possible after
sample collection.
• DO NOT freeze the sample.
• Samples must be received in the laboratory within one calendar
day of collection.
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table 12: Water – collection and transportation instructions ...continued
Public Beaches
The PHOL tests public beach water samples for the microbiological indicator Escherichia coli. Refer to the Ministry of Health and Long-Term
Care Beach Management Protocol, 2008 (or as current) for more information about the protocol for public beaches.
Prior to the beginning of each beach season, notify the PHOL of the number of samples that will be submitted, day(s) of week for sampling,
and approximate time of day that samples will arrive at the laboratory. Note: Collect five (5) samples at designated sites at each identified
beach location. More than five (5) samples may be required for larger beaches.
samPlinG instructions
Precautions
1. Complete the required information on the requisition.
2. Remove one barcode from the bottle and apply it to the top
copy of the requisition in the “Barcode” field.
Samples will not be processed if the barcode is not affixed to the
requisition. If the barcode is not able to be removed, write the
barcode number on the form.
3. Examine the lid of the bottle.
If the tamper evident ring has separated from the cap use another
bottle.
4. Remove cap and ensure it is kept away from sources of
contamination.
• Do not touch the inside of the lid
• Do not put the lid down
5. Plunge bottle downward into water to 30 cm – turn bottle in
direction point of current.
• Where the depth of water is 1 to 1.5 meters, the samples are to
be obtained at 15 to 30 cm below the water surface.
• Where the depth of water is less than one (1) meter, obtain
samples as far off shore as possible, but within swimming area.
6. Remove bottle from water.
If the bottle is filled to the top, remove some of the water so it is
filled to 200mL line.
7. Replace cap.
8. Store and transport samples below between 1°C and 10°C
(preferably 4°C) within one calendar day of collection.
20
• DO NOT freeze the sample.
• Samples must be received at the laboratory within one calendar
day after collection.
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Water - Laboratory Rejection Criteria
Table 13 outlines the various reasons a water sample will be rejected by the PHOL, and a new sample must be submitted with a completed
requisition.
table 13: Water – laboratory rejection criteria
DescriPtion
reason for rejection
Sample Type
• Bottled water with intact seal
• Specimen received from an unauthorized submitter
• Private citizen collected non-treated surface water or drinking
water sample collected by a Private citizen from a regulated
source
• Specimen location from outside of Ontario
Sample Requisition
• Requisition received with incomplete and inaccurate information
• Unique identifier(s) missing from requisition, specimen
collection bottle or both
• Requisition received separated from the specimen bottle with
no unique identifier to link them
Sample Collection
•
•
•
•
•
Sample collected in non-PHOL collection bottle
Outdated collection kit received (Private citizen only)
Insufficient volume received for testing
Aesthetically unacceptable drinking water sample
Interfering substances identified during testing preventing
analysis
Sample Transport
•
•
•
•
•
•
•
Drinking water received greater than 48 hours after collection
Non potable water received greater than 1 calendar day
Specimen bottle or lid damaged in transit
Specimen leaked in transit
Specimen frozen upon receipt
Specimen indicated as collected from a hot water tap
Specimen received greater than 25°C (unless from a public
beach, recreational water facility or hydrotherapy pool)
Broken Chain of Custody
• Regulated drinking water –“Relinquished by” section of
requisition not completed or of Custody sample lacked an
initialed and dated seal affixed to the cap when received.
• Legal water samples –“Relinquished by” section of requisition
not completed and an initialed and dated seal not affixed to the
cap when received
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Suspected Sewage Contamination Sample
Analysis
Collection and Transportation Instructions
Table 14 provides instruction for the use of the Sewage
Sample Collection Kit which was introduced in 2007 and is
to be used by board of health staff when they are
investigating situations of possible sewage contamination.
For health and safety reasons, any samples identified as
“Suspected Sewage Contamination” that are not received in
this kit will not be processed and the board of health will be
requested to collect a new sample using the proper
requisition and kit.
The PHOL tests “Suspected Sewage Contamination
Samples” for the microbiological indicator Escherichia coli.
The Sewage Sample Collection Kit consists of:
• A sterile bar-coded PHOL water collection bottle
• A sealable biohazard bag for the sample with a pocket on
the side to hold the requisition
• A hard transport case labeled SEWAGE SAMPLES ONLY
containing absorbent material
• A soft sided blue transport bag (#TC 125-B) labeled with
the laminated tag with SEWAGE SAMPLES ONLY on one
side and the return address of the local PHOL on the
opposite side
• A “Bacteriological Analysis of Water Single Sample
Requisition for Official Agencies” (see Appendix B)
• An instruction sheet entitled “Collection and
Transportation Procedure for Sewage Sample Submitted
to the Ontario Public Health Laboratories”
table 14: suspected sewage contamination samples – collection and transportation instructions
Precautions
samPlinG instructions
22
1. Complete the required information on the requisition.
Samples will not be processed if the requisition is not completely
and accurately filled in when received at the laboratory, and a new
sample and completed form will be required to be submitted.
2. Remove one barcode from the bottle and apply it to the top copy
of the requisition in the “Barcode” field.
Samples will not be processed if the barcode is not affixed to the
requisition. If the barcode is not able to be removed, write the
barcode number on the form.
3. Examine the lid of the bottle.
If the tamper evident ring has separated from the cap use another
bottle.
4. Remove the cap.
• Do not touch the inside of the lid
• Do not put the lid down
• Do not rinse out the bottle as this will remove the sodium
thiosulfate which neutralizes the chlorine residual before testing
in order to maintain bacterial viability during sample transport
to the laboratory.
• Do not ingest; for accidental exposure get medical attention.
5. Wearing rubber/latex gloves, collect the suspected sewage
contaminated sample to the 200mL fill line.
• Do not overfill.
• If overfilled, remove some of the water so it is filled to 200mL
line.
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table 14: suspected sewage contamination samples – collection and transportation instructions ...continued
samPlinG instructions
Precautions
6. Tighten the cap on the collection bottle.
Leaking samples will not be processed; a new sample and
requisition will be required.
7. Wipe down exterior of collection bottle with disinfectant
A freshly prepared 1% solution of sodium hypochlorite, i.e.1 part
household bleach to 10 parts water is recommended
8. Remove and discard rubber/latex gloves. Wash / sanitize your
hands.
Do not put in biohazard bag along with sewage sample.
9. Seal the sewage specimen bottle in the biohazard bag supplied.
10.Put the completed requisition in the side pocket of the
biohazard bag containing the sewage sample.
Do not place the requisition inside the biohazard bag with the
sample.
11.Place the packaged sewage specimen in the hard transport case
labeled SEWAGE SAMPLES ONLY containing absorbent material
and a frozen ice pack. This container is placed inside a blue
laboratory sewage transport bag (#TC 125-B).
• The sewage transport bag will have a permanently attached
laminated tag on the top stating in red “SEWAGE SAMPLES
ONLY”.
• Do not send sewage samples in any other transport container or
with other environmental or medical specimens, as sample
integrity cannot be ensured, resulting in the rejection of all
samples submitted in this manner.
12.Store and transport samples between 1°C and 10°C (preferably 4°C) • DO NOT freeze the sample.
within one calendar day of collection.
• Samples must be received at the laboratory within one calendar
day after collection.
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Legionella Investigations
Legionella – Microbiological Analysis Test Menu
Table 15 provides an overview of the testing available and
turn around times for Legionella through the PHOL. Turn
around time is defined as the number of business days to
complete testing based on date of receipt in the laboratory.
This turn around time definition does not apply to tests that
are on-hold awaiting information to continue testing. Turn
Around Times posted in this Guide are estimates for routine
tests. For some tests or at times of public health
emergencies, outbreaks or exigent circumstances, turn
around times may be longer than what is listed.
PHO/OAHPP disclaims all warranties whether express or
implied in respect of the information, including the TATs
provided herein. Environmental testing for Legionella will
only be performed for an investigation of a confirmed
clinical case and this information must be provided to the
laboratory when environmental testing is requested.
table 15: Legionella – microbiological analysis test menu
samPle DescriPtion
samPle container
tests PerformeD
turn arounD time
Water – potable
PHOL water sample bottle – one liter
5 X 200mL
Legionella culture
10 – 15 days
Water – non-potable
PHOL water sample bottle – 200mL
Legionella culture
10 – 15 days
Environmental Swabs
Contact the PHOL – Toronto, Environmental
Microbiology Laboratory at 416-235-5716
Legionella culture
10 – 15 days
Solid material
Sterile plastic bag with round wire closure
Legionella culture
10 – 15 days
Legionella – Selection of Sampling Sites
The investigation, including environmental and
epidemiologic factors, should be considered in choosing the
sites to be sampled to identify the source of Legionella.
investigation and selection of sampling sites, but it is not a
list of mandatory sampling sites. Additional resources may
be found under Supporting Documentation/References.
Legionella outbreaks occur because of aerosolization and
inhalation of high numbers of Legionella. Investigators
should try to determine where in a water system there is
stagnation of water and subsequent aerosolization of
stagnant water. Any dead ends in a water system may allow
Legionella to grow to high numbers. Dead-ends may be
disconnected fixtures where pipes are capped instead of
completely removed. Any fixtures and water lines that are
not used on a regular basis may allow Legionella to grow to
high numbers.
• Potable water outside or on the boundary of the hospital
property
o Treatment plant (raw and refined water)
o Guard house or outlying facility if water is not fed there
from hospital
o Fire hydrant(s)
• General potable water system for hospital
o Incoming water pipe(s)
o Water softener (pre and post)
o Preheater (discharge side)
o Primary heater
o Circulating pump(s)
o Holding tanks (cold water, discharge side)
o Expansion tank for hot water (if possible)
o Back drain on sprinkler system(s) (trap to prevent back
flushing may be present and should be sampled)
o Fireline where it branches off main system (may be
multiple)
• Pharmacy
o Water used for respiratory therapy equipment
Samples most likely to be positive should be sampled and
submitted first. Random sampling may delay the
identification of the source of the outbreak. The most
obvious sampling sites would be water sources in close
proximity to infected individuals.
The following is a list of sampling sites recommended by the
Centers for Disease Control, Atlanta, for investigating
outbreaks (Barbaree JM, 1987). It can be a guide for
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• Air compressor system
o Vacuum water source
o Positive pressure equipment side
o Condensate from tanks(s)
o Water separator(s) (directly off compressors)
o Water source(s) near air intake(s)
o Air samples where patients were ill with legionellosis
• Potable water final distribution outlets
o Hemodialysis water source
o Before demineralizer
o After demineralizer
o Intensive care units
o Respiratory therapy (patient rooms)
o Cardiac
o Services with different geographical locations
o Ice maker (entry water)
• Air-conditioning system
o Air handling unit to service where disease occurred
(drain pan)
o Blowdown
o Water supply
• Whirlpool(s)
o Whirlpool (one nearest air intake system)
o Whirlpool drain
• Other
o Decorative fountain(s)
o Creeks, ponds, and sites of stagnant water.
Legionella – Environmental Sample Collection and Transportation
Instructions
Table 16 provides information and sampling instructions for
water, environmental swabs and solid material in the
investigation of Legionella.
notes:
• The PHOL should be notified prior to sample submission
to allow for preparation of materials required for testing.
Please contact the PHOL – Toronto, Environmental
Microbiology Laboratory at 416-235-5716.
• Samples should be stored at refrigeration temperature (2
to 6°C) and shipped to the laboratory as soon as possible.
• Each sample must be labeled with the unique identifier
that must also be affixed to the requisition. PHOL water
bottles have barcodes attached that can be used for this
purpose. Absence of a unique identifier linking sample(s)
to a sampling location on a requisition will result in
rejection of sample(s) by the laboratory.
• Drinking water analyses are performed in outbreak
situations or upon special request and MUST be
submitted as NON-REGULATED requests only.
table 16: Legionella – environmental sample collection and transportation instructions
Legionella Investigations –Water Sampling
samPlinG instructions
Precautions
1. Collect water in PHO water collection bottles:
For potable water, collect one liter of water. Five standard
bacteriology water bottles filled to the fill line will satisfy this
requirement.
For non potable waters, a single bottle of 200mL is sufficient.
• Taps or showers should not be allowed to run before sampling.
Turn water on and allow the water to run slowly into sample
bottles.
• Use of Personal Protective Equipment (PPE) should be guided by
local board of health internal health and safety guidelines.
2. Complete the requisition:
For liquid samples collected in water bottles, use either of the
following requisitions:
• Bacteriological Analysis of Water – Multiple Sample Requisition
for Official Agencies
• Bacteriological Analysis of Water – Single Sample Requisition for
Official Agencies
• Drinking water analyses are performed in outbreak situations or
upon special request and MUST be submitted as NONREGULATED requests only.
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table 16: Legionella – environmental sample collection and transportation instructions ...continued
Legionella Investigations – Environmental Swab Sampling
Environmental swabs can be used for Legionella outbreak investigations. These are available (for outbreaks only) from the PHOL, Toronto.
The environmental swabs available from the PHOL – Toronto contain neutralizing solution as transport medium. This medium can
neutralize the following sanitizers: iodophores including chlorine compounds, quaternary ammonium compounds and phenolic sanitizers.
Environmental swabs are processed during outbreak investigations for the presence or absence of Legionella.
samPlinG instructions
Precautions
1. If a water sample has not been collected, collect one first as
described in Legionella Investigations – Water Sampling.
• Swabs are usually collected from shower heads or other
implements that aerosolize water.
2. Remove shower head.
3. Open the sterile swab container; grasp the handle end of the
swab.
• Remove the swab aseptically being careful not to touch any
portion that might be inserted into the vial.
4. Moisten the swab head and press out the excess solution against
the interior wall of the vial with a rotating motion.
5. Use the head of the swab to scoop up any sediment that has
built up inside the shower head.
6. If there is no sediment in the shower head, swab the inside of
the shower head and the pipe to which the shower head was
attached to pick up biofilm.
For cooling towers, swabs can be used to sample any build-up of
slime or biofilm on the equipment:
Hold the swab handle to make a 30° angle contact with the
surface. Rub the swab head slowly and thoroughly over
approximately 50 cm2 of the surface three times, reversing
direction between strokes.
7. After the area has been swabbed, using aseptic technique,
return the swab to the vial.
• Ensure the vial is closed tightly to prevent leakage and specimen
rejection.
8. Complete all sections of the Environmental Bacteriology Swab
Tests requisition (see Appendix E) with appropriate information
to ensure accurate tracking of sampling.
Legionella Investigations – Solid Material Sampling
samPlinG instructions
Precautions
1. Sterile plastic bags with round wire closure can be used to
collect solid material as required. Such material may include
filter material from air filters or water filters.
• These bags are available at most board of health offices for the
collection of food samples.
2. For solid material collected in plastic bags use the requisition
entitled Environmental Bacteriology SwabTests (see Appendix E).
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Glossary – Water Terms
term
Definition
aesthetically unacceptable drinking water
Drinking water that is not clear, colourless or odourless
audit sample
A sample taken outside of the routine testing schedule as part of an audit
background bacteria in potable water
Bacteria which fail to ferment lactose on culture media designed for
coliform detection. They are also know as non-target organisms in
drinking water testing
bottled water
Potable water in commercially sealed containers
chain of custody
An unbroken documented chain of events from the time of sample
collection until it is processed and completed
cistern
A reservoir, tank or container for storing or holding water
compliance
A sample taken to fulfil the requirements of an Ontario drinking water
regulation
control sample for water testing
A sample taken prior to a change in a routine or prior to the application
of a corrective action
customer investigation
An investigation resulting from a verbal or written communication by a
customer
Designated facility
A facility as described in Ontario Regulation 170/03 Drinking Water
Systems
Distribution
The section of a drinking water system that transports drinking water to
the end user
Escherichia coli (e. coli)2
E. coli are coliform bacteria. They are Gram-negative, facultatively
anaerobic, non spore-forming, lactose-fermenting, rod-shaped bacteria
which are oxidase negative. Approximately 95% of E. coli strains can
produce the enzyme β-D-glucuronidase, which splits the substrate 5bromo-4-chloro-3-indolyl-β-D-glucuronide (i.e. B.C.I.G.) within 24-72 h at
36°C.
Historically, the presence of either or both E. coli and fecal coliforms has
been described as an indication of sewage or fecal contamination as
they are commonly isolated from the intestinal tract of warm-blood
animals. Currently, E. coli is considered to be the most specific indicator
of fecal contamination in the assessment of water quality for the
following reasons:
• E. coli isolates are normally present in the feces of warm-blooded
animals (including humans) at higher densities (107 to 108 cells per
gram) than other lactose-fermenting members of the family
Enterobacteriaceae.
• E. coli isolates may persist for periods of time, once outside the
human or warm-blooded animal intestine but they generally do not
multiply in water or wastewater in temperate climates.
• E. coli isolates have seldom been detected in environments which
have not been contaminated by fecal wastes. In contrast, other
thermotolerant organisms such as isolates of the genera Klebsiella,
Enterobacter and Citrobacter, have been associated from both fecal
material and other wastes including that from food and pulp and
paper.
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Glossary – Water Terms ...continued
28
term
Definition
Etiological Agent
The cause or origin of a disease
Frozen
Water at or below zero degrees C or containing obvious ice fragments
Heterotrophic Plate Count (HPC), Aerobic Plate Count (APC) or
Standard Plate Count (SPC)
The heterotrophic plate count is a process of estimating the number of
viable, heterotrophic bacteria (i.e. bacteria capable of growth aerobically
on Standard Methods Agar or a similarly formulated growth medium
when incubated at 35 +/- 0.5°C for 48 +/-3 hours) in water and
measuring the changes that occur to bacterial populations during water
treatment and distribution
HPPA O. Reg. 318/08 Drinking Water
Drinking water that is subject to the requirements as described in
Ontario Regulation 318/08 under the Health Protection and Promotion
Act until it is repealed
HPPA O. Reg 319/08 Drinking Water
Drinking water that is subject to the requirements as described in
Ontario Regulation 319/08 under the Health Protection and Promotion
Act
HPPA Regulated Premises
Premises/facilities regulated under HPPA, (Food premises, Recreational
water places, etc.) that fall outside the scope of O. Reg. 319/08
Hydrotherapy pool
Public spa as per Ontario Regulation 428/05 Public Spas
Ice
Potable water which has been cooled to a temperature less than 0°C
Interactive Voice Response (IVR)
A technology that allows a computer to interact with humans through
the use of voice and dual-tone multi-frequency signaling keypad inputs.
Private Citizen drinking water test results and the interpretation can be
accessed by calling a toll free telephone number and keying in the
sample barcode when prompted
Interfering substances
A substance in a water sample that either prevents the water from
passing through the membrane filter during the testing process or after
the incubation process, there is a substance on the membrane filter that
does not allow an accurate determination of the colony count
Legal sample
Any laboratory specimen that has been submitted for testing to the PHO
Laboratories with an intact legal seal and whose chain of custody has
been maintained
Municipal Drinking-Water System
Refer to the Safe Drinking Water Act, 2002
Non Municipal Drinking Water System
Refer to the Safe Drinking Water Act, 2002
Non Regulated Drinking Water
Drinking water systems that are not captured by specific regulation
where the public has access to the water and such systems are subject
of an investigation under the HPPA, public health inspectors may collect
drinking water samples in concurrence with the following sections of the
HPPA: Section 13, Section 41, Subsections (1), (2), (4), and (5). See Table
A Examples of Drinking Water that Fall Outside Ontario’s Drinking Water
Regulations
Non treated
Water that is not subjected to disinfection or other means of removing
contaminants
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Glossary – Water Terms ...continued
term
Definition
Official Agency
a) Board of health (public health unit): An official health agency
established by a group of urban and rural municipalities to provide a
more efficient community health program, carried out by full-time,
specially qualified staff, or
b) Ministry of Natural Resources Ontario Park: Ontario land dedicated to
the people of Ontario and visitors for their inspiration, education,
health, recreational enjoyment and other benefits with the intention
that these areas shall be managed to maintain their ecological
integrity and to leave them unimpaired for future generations. Refer
to Provincial Parks and Conservation Reserves Act, 2006
Outbreak investigation
A sample taken as part of a pubic health outbreak investigation
Outbreak number
The number assigned to an outbreak by public board of health staff for
tracking purposes
Overgrown
The crowding and /or confluent and/or non-identifiable microbial
growth on a plate, and there is no evidence E. coli, the culture is
considered overgrown with background bacteria
Overgrown with the presence of E. coli
Overgrown with the presence of E. coli The crowding and /or confluent
and/or non-identifiable microbial growth on a plate, but there is
evidence of E. coli as well.
Private Citizen
A person residing in Ontario who:
a) does not have access to a treated municipal drinking water supply and
b) draws their drinking water from a protected source within the
Province of Ontario for personal use or applies appropriate treatment
to water drawn from an unprotected source
Private Citizen Drinking Water
Drinking water from a “single household” and private residence as
defined in Ontario Regulation 171/03 Definitions of Words and
Expressions Used in the Act
Private Residence
Refer to Ontario Regulation 171/03 Definitions of Words and
Expressions Used in the Act (e.g. water source can include a well,
cisterns, treated surface water, etc.).
Pseudomonas aeruginosa
A Gram negative, non-fermentative, oxidase positive, rod shaped
bacteria, which grows well at 42°C, is actively motile, and under
appropriate incubation conditions produces a fluorescent pigment and
often a visible pigment.
Public Beach
Refer to Ontario Public Health Standards 2008 – ‘Beach Management
Protocol’, 2008 or current version
Recreational Water Facilities
Refer Ontario Public Health Standards, 2008. ‘Recreational Water
Protocol’, 2008 or as current
Regulated water seal
A fixture attached to the cap of a drinking water sample submitted
under a drinking water regulation that must be broken before access can
be obtained
Relinquish
To release a sample ensuring the chain of custody is not compromised
Resample
A sample taken from the same location. Usually in response to a
adverse test result or incident
Routine surveillance
Observation over a group, area or system on a regular basis
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Glossary – Water Terms ...continued
term
Definition
SDWA O. Reg 170/03 Drinking Water
Drinking water that is subject to the requirements as described in
Ontario Regulation 170/03 Drinking Water Systems (e.g. some
Municipal, Non-municipal and designated facilities)
Single Household
Potable water from any source which is protected from surface or
ground water contamination, regardless of any filtration procedure
and/or chemical disinfection applied, is aesthetically acceptable, and
serves fewer than 6 private residences but not to the public. For
analytical purposes, water passing through a point-of-use treatment
device will be regarded as a private water source.
Staphylococci sp.
Staphylococci are Gram-positive cocci of the family Micrococcaceae.
Staphylococcus aureus
S. aureus is a facultative anaerobe of the family Micrococcaceae that
produces a wide array of toxins and can coagulate rabbit plasma.
Surface water
Water collecting on the ground or in a stream, river, lake, etc.
Suspected sewage contamination
Water suspected of being contaminated by fecal material
Test for Potability
Total coliform and E. coli analysis
Total coliforms 2
Coliforms are described as Gram-negative, non spore-forming, rodshaped bacteria capable of fermenting lactose with the production of
acid and gas, within 48 hours, when grown in a lactose-containing liquid
culture medium incubated at 36+/-1.0°C. The total coliform count is the
quantification of colony forming units (CFU) that meet this definition
(including E. coli) per 100 mL of sample.
There are some lactose-fermenting strains of species within the genus
Aeromonas which produce a coliform-like reaction on media used for
the detection of bacterial indicators of contamination. Aeromonas
species are oxidase-positive and generally fail to produce gas during
lactose fermentation in liquid media. They do not, therefore, conform
to the classical definition of a coliform. Regardless, they are often
included in the total coliform count because the growth medium fails to
differentiate this organism from the true coliforms.
Some coliform bacteria are commonly isolated from the fecal material
found in the intestines of humans and other warm-blooded animals (eg.
E. coli) and coliforms, in general, may also be isolated from plants, soil
and sediments. The Total Coliform count, therefore, does not
necessarily provide specific evidence of fecal contamination.
Treated
Water that is subjected to disinfection or other means of removing
contaminants
Unauthorized Submitter – Official Agency
A submitter representing an agency other than a board of health or a
Ministry of Natural Resources Ontario Park
Unauthorized Submitter – Private Citizen
A person who submits a drinking water sample that does not meet the
definition of “Private Citizen” or “Single Household”
Unique identifier
A sample identifier that uniquely distinguishes it from any other sample
submitted. The sample barcode is used for this purpose.
Water well
A hole drilled or bored into the earth to obtain water
2
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Some information taken from Ministry of the Environment’s Laboratory Services Branch method MICROMFDC-3407
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table a: examples of Drinking Water that fall outside ontario Drinking Water regulations
The following list provides some examples which fall outside the scope of the Safe Drinking Water Act 2002, Drinking Water Systems
Ontario Regulation 170/03 and the Health Protection and Promotion Act, Small Drinking Water Systems Regulations (i.e. O. Reg 318/08 and
O. Reg 319/08).
Drinking water systems that are not captured by specific regulation where the public has access to the water and such systems are subject
of an investigation under the HPPA, public health inspectors may collect drinking water samples in concurrence with the following sections
of the HPPA: Section 13, Section 41, Subsections (1), (2), (4), and (5).
The following examples may be considered ‘drinking water systems’ in which drinking water samples must be analyzed at a licensed
laboratory using licensed drinking water testing methods, unless the example is entirely a plumbing system or they are facilities with no
‘works’ that have been established for providing potable water.
situation
examPle
Drinking water supplies that do not meet the specific
criteria of the Safe Drinking Water Act 2002 or Health
Protection and Promotion Act drinking water
regulations
• Seasonal campsite with fewer than 6 water connections, camps in
unorganized territory, certain recreational camps, rental cabins with fewer
than 6 units using the same water supply, etc.
• Raw water that it is not for human consumption and is not distributed to the
public (i.e. unfinished water in a treated drinking water system, new water
mains, etc.)
Facilities in the following circumstances
• Serviced by private drinking water systems that require health unit approval
for licensing or funding approval: Funeral homes, group and boarding homes,
homes for special care, migrant farm workers, etc.
Food premises in the following circumstances
• Small food take out and convenience stores where there is no direct access to
drinking water by the patrons, temporary food service facilities, farmers
markets, etc.
• Special event food premises or mobile food vending vehicles
Outbreak investigations
• Testing of a drinking water system as part of an outbreak investigation (e.g.
Legionella, E. coli O157:H7, Salmonella, etc.), regardless if the drinking water
system is regulated under drinking water regulations under the Safe Drinking
Water Act 2002 or the Health Protection and Promotion Act.
Plumbing in the following circumstances
• Samples are taken from a building’s plumbing (i.e. when the water enters
private property) although the water originated from a drinking water system
under an Ontario drinking water regulation. Example - a private residence in
a town or city, private apartment buildings, factory or food manufacturing
plant, etc. If a sample is collected to assess the water quality and not the
plumbing, then it is considered a drinking water sample and must be analyzed
at a licensed laboratory using licensed drinking water testing methods.
Private individual well water supplies
•
•
•
•
Water Containers
• Water haulage trucks (testing may be required as part of an inspection as per
Drinking Water Haulage Guidance Document)
• Private building water coolers
• Commercial bottled water / water bottling facilities
• Vending machines that dispense water, i.e. “Bottle your own water”
• Cisterns not supplying systems regulated under the Safe Drinking Water Act,
2002 or the Health Protection and Promotion Act
Enteric disease follow up
Assisting with addressing / identifying water safety concerns
Complaints
Landlord and tenant matters and samples are taken of the private water
supply for the issuance of a section 13 order for a violation of section 20
under the HPPA
• Treatment devices for samples taken to assess the ability of a newly installed
treatment process at a premise.
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Interpretation of Environmental
Microbiological Test Results
Table 17 summarizes acceptable limits and interpretation of
test results for all tests performed by the PHOL for foods,
water, suspected sewage contamination samples and
Legionella investigations.
Microbial standards for drinking water testing are based on
Ontario Drinking Water Quality Standards (O. Reg 169/03)
with input from the Ministry of Health and Long-Term Care
Environmental Health Policy and Programs Public Health
Division. Microbial standards for food testing, public beach
water testing, and Legionella environmental testing are
based on published evidence where available. In a
foodborne outbreak investigation, interpretation is
dependent on the pre-test probability of a food being
contaminated with the suspected or confirmed pathogen,
based on epidemiological information obtained during the
investigation.
For many bacterial organisms, such as E. coli O157:H7 and
Listeria monocytogenes, molecular typing by pulsed field gel
electrophoresis can be used to support a link between
clinical and environmental isolates.
table 17: interpretation of environmental microbiological test results
analysis
background bacteria
samPle tyPe/source
rePortinG limits
accePtable limits
Water – Potable
0 to “Overgrown” (only reported
on Regulated drinking water
samples submitted under an
“Order” or “Approval”)
< 200 per 100 mL
Recreational Water Facilities and
Hydrotherapy Pools
Only reported if “Overgrown”
N/A
laboratory test interpretation: Presence of background organisms is an unusual occurrence in
drinking water, which has been properly disinfected. High levels (>200 per 100 mL) reflect a
deterioration in water quality and an increase in health risk potential. May include bacterial
pathogens most of which are non-coliforms.
When samples are heavily contaminated (i.e. overgrown) with bacteria often found in the
environment, this condition interferes with the detection of coliforms or Escherichia coli that may
be present in the sample. The water may be unsafe to drink unless boiled or treated.
Bacillus cereus
Food
<100 to >2x105 per gram
<103 per gram
laboratory test interpretation: B. cereus is common in the environment and may be present in
food in small numbers. Isolation of >105 organisms per gram of food may be associated with
foodborne illness.
Campylobacter jejuni
Water
Presence/Absence
Absent
laboratory test interpretation: Transmission of Campylobacter via water is epidemiologically
implicated, but the organism has rarely been isolated from suspect sources.
Analysis of water samples for Campylobacter should only be undertaken when there has been
clinical confirmation of an outbreak and a water source has been incriminated epidemiologically.
Food
Presence/Absence
Absent
laboratory test interpretation: C. jejuni is a major cause of acute bacterial enteritis in humans. It is
often present in raw food of animal origin. This organism is highly susceptible to environmental
stresses and does not survive proper cooking or pasteurization. The majority of foodborne
outbreaks are the result of consumption of raw, under processed or cross-contaminated foods.
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table 17: interpretation of environmental microbiological test results ...continued
analysis
Clostridium botulinum
samPle tyPe/source
rePortinG limits
accePtable limits
Food/Feces/Serum
Presence/Absence
Absent
laboratory test interpretation: Human food poisoning is rare but case fatality rate (untreated) is
high. Demonstration of neurotoxin in serum or feces is necessary to confirm diagnosis. Detection of
the same neurotoxin-producing type of C. botulinum in suspect food is sufficient to implicate the
food as the source of illness.
Clostridium perfringens
Food
<100 to >2x106 per gram
<100 per gram
laboratory test interpretation: C. perfringens is part of the intestinal flora of all warm-blooded
animals and can be found in low numbers in the environment. Food poisoning is caused by the release
of enterotoxin in the intestine by sporulating organisms. A count of greater than or equal to 105
organisms per gram of food is associated with foodborne illness.
Coliforms
Water – Potable
0 to >80 per 100 mL
Private Citizen
samples: ≤5 per
100 mL
Official Agency
and bottled
water samples: 0
per 100 mL
Recreational Water Facilities and
Hydrotherapy Pools
<2 to > 160 per 100 mL
<2 per 100 mL
laboratory test interpretation: The presence of coliforms may be indicative of a contaminated
water supply. Coliforms occur naturally in soil and decaying vegetation; but may also be associated
with human or animal fecal contamination. Given the vulnerability of well water to external
influences, such as human, environmental and agricultural impacts, it is important to test water
frequently.
Food
<3 to >1.1 x 103 per gram
<103 per gram
laboratory test interpretation: Presence of coliforms in a processed, cooked, ready-to-eat food is
indicative of inadequate processing or post processing contamination resulting from poor handling
of the product. This acceptable limit cannot be applied to uncooked or raw foods such as salads and
salad ingredients, raw fruits and vegetables, etc.
Ice cream
<3 to >1.1 x 103 per mL
≤10 per mL
laboratory test interpretation: Presence of coliforms suggests recontamination following
pasteurization but could also indicate inadequate pasteurization.
escherichia coli
Water – Potable
0 to >80 per 100 mL
0 per 100 mL
Recreational Water Facilities and
Hydrotherapy Pools
<2 to > 160 per 100 mL
<2 per 100 mL
Public Beaches
<10 to >103 per 100 mL
<100 per 100 mL
laboratory test interpretation: The presence of Escherichia coli may indicate fecal contamination
from a human or animal source. There is a strong relationship between increased levels of
Escherichia coli in recreational waters and the incidence of swimming associated gastroenteritis.
Suspected Sewage Contamination
<103 to >1x106 per 100 mL
Not Applicable
laboratory test interpretation: Escherichia coli levels in water >10 per 100 mL are indicative of
fecal contamination
5
Food
<3 to >1.1 x 103 per gram
<3 per gram
laboratory test interpretation: The presence of Escherichia coli in food is indicative of direct or
indirect contamination of the food product with fecal material and thus, a potential for enteric
pathogens to also be present. Certain serotypes of Escherichia coli are capable of causing outbreaks
of diarrhea.
e.coli o157:H7
Food
Presence/Absence
Absent
laboratory test interpretation: The presence of E.coli O157:H7 in a processed, cooked or ready-toeat food is a definite health hazard.
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table 17: interpretation of environmental microbiological test results ...continued
analysis
Heterotrophic Plate count
(HPc)
samPle tyPe/source
rePortinG limits
accePtable limits
Water – Potable
<10 to >2x10 per mL
<500 per mL
3
laboratory test interpretation: An HPC >500 per mL in a distribution system or other treated water
is indicative of poor water quality and possible problems with the disinfection procedure.
Recreational Water Facilities and
Hydrotherapy pools
<10 to >2x103 per mL
See comments
under “Laboratory
Test Interpretation”
laboratory test interpretation: APHA recommends for public swimming pools that not more than
15% of samples collected during any 30-d period shall have an HPC count of 200/mL.
Food
<103 to >2.0 x 105 per gram
<105 per gram
laboratory test interpretation: It is reasonable to expect properly prepared and handled ready-toeat processed or cooked foods to have an HPC <104 per gram. An HPC >105 per gram is evidence of
temperature abuse – inadequate cooking or improper storage conditions. HPC cannot be applied to
raw, uncooked, unprocessed foods (e.g. fruits or vegetables) or cultured products (e.g. cheese).
Legionella species
Taps/sinks/spas/ showers
Presence/Absence
Significance
interpreted per
investigation in
consultation with
medical
microbiologist
laboratory test interpretation: Except for high risk populations, routine environmental monitoring
for Legionella is not recommended. Legionellae have predilection for sediments, slimy deposits and
rubber fittings in water distribution systems. Recovery of Legionellae from >30% of different
sampling sites is considered to indicate a hazard.
Listeria monocytogenes
Food (high risk foods, or in outbreak
investigation)
Presence/Absence
Absent
laboratory test interpretation: Reference Food Directorate, Health Canada Policy on Listeria
monocytogenes in Ready-to-eat Foods, FD-FSNP 0071 April 1, 2011
pH
Food
1.00 to 14.00
laboratory test interpretation: pH is a measure of food acidity, which varies with food type. Foods
with a pH ≤ 3.7 will not support the growth of bacterial foodborne pathogens. However, if the pH of
the food (jarred/canned products) is ≤ 4.6, all micro-organisms are inhibited with an aw ≤ 0.85.
Phosphatase
(alkaline phosphatase)
Dairy
Presence/Absence
Absent
laboratory test interpretation: The detection of residual alkaline phosphatase indicates a reduction
in pasteurization temperature of at least 1.5°C; a 5 minute reduction in holding time; or the
presence of ≥0.3% raw milk.
Pseudomonas aeruginosa
Recreational Water Facilities and
Hydrotherapy pools
0 to >100 per 100 mL
<10 per 100 mL
Water – Potable/Bottled
0 to >100 per 100 mL
0 per 100 mL
laboratory test interpretation: P. aeruginosa can be found in sewage effluents at levels of up to 105
per 100 mL.Well operated pools should not normally contain P. aeruginosa. If the count is over 10 P.
aeruginosa per 100 mL, repeat testing should be undertaken. Where repeated samples contain P.
aeruginosa the filtration and disinfection processes should be examined to determine whether
there are areas within the pool circulation where the organism is able to multiply. Where counts
exceed fifty, pool closure should be considered. P. aeruginosa is a primary cause of ear, eye and skin
infections among swimmers.
34
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table 17: interpretation of environmental microbiological test results ...continued
analysis
Salmonella species
samPle tyPe/source
rePortinG limits
accePtable limits
Water
Presence/Absence
Absent
laboratory test interpretation: When present, Salmonella are usually in low numbers in water.
Large sample volumes are necessary to allow for a concentration procedure. Presence of Salmonella
in potable water should not be tolerated, as such conditions allow the potential of the organism to
spread and multiply and become a significant health risk.
Food
Presence/Absence
Absent
laboratory test interpretation: The presence of Salmonella in a processed, cooked or ready-to-eat
food is a definite health hazard. The presence of Salmonella in a raw product, particularly poultry,
beef or pork, to be subjected to further processing prior to consumption, is not unusual.
Staphylococcus aureus
Recreational Water Facilities and
Hydrotherapy Pools
0 to >100 per 100 mL
<50 per 100 mL
Water – Potable/ Bottled
0 to >100 per 100 mL
0 per 100 mL
laboratory test interpretation: S. aureus is a major pathogen responsible for purulent infections of
the skin, eyes and ears. Its presence in swimming pools and hydrotherapy pools is a good index of
the health hazard associated with a high bather density.
Food
<100 to >2x105 per gram
<100 per gram
laboratory test interpretation: S. aureus is a major foodborne disease agent causing nausea,
vomiting, abdominal cramps and diarrhea. Preformed toxin in foods causes an onset of symptoms
within one to eight hours of ingestion of food. Recovery usually occurs within 24 hours. To be
implicated in foodborne disease, a suspect food should contain >105 S. aureus per gram. An
epidemiologically implicated food may be tested for staphylococcal enterotoxin. The presence of
enterotoxin is a definitive diagnosis. Detection of the same enterotoxin-producing isolates from a
suspect food and related clinical specimens is strong evidence that the food is directly implicated.
total Gram negative count
(tGnc)
Food
<103 to >2x105 per gram
<103 per gram
laboratory test interpretation: TGNC has same significance as presence of coliforms. Elevated
TGNC should not be detected in cooked or processed ready-to-eat foods. Absence of gram negative
organisms in food sample is another indicator of food safety.
Vibrio parahaemolyticus
Vibrio cholerae
non-01 Vibrio cholerae
Vibrio fluvialis
Vibrio vulnificus
Vibrio mimicus
Food
Presence/Absence
Absent
laboratory test interpretation: May be the result of insufficient cooking or recontamination from
mishandling. Implicated foods include crab, oysters, shrimp and lobster. Organisms can be recovered
from brackish coastal waters, sediment and sea-life in temperate estuarine environments. Specialized
culture techniques must be used to recover vibrios from food samples.
Water activity (aw)
Food
N/A
N/A
laboratory test interpretation: The availability of water for microbial, enzymatic or chemical
activity determines the shelf life of foods. This water availability is measured as water activity (aw).
Water activity is measured on a scale of 0 to 1, where 0 indicates no water and 1 indicates all water.
For a food to be considered safe to store at room temperature, it should have an aw ≤ 0.85. Bacterial
foodborne pathogens are inhibited at this aw; however, spoilage microorganisms would be inhibited
below aw 0.6.
Yersinia enterocolitica
Food/Water
Presence/Absence
Absent
laboratory test interpretation: Pathogenic biotypes have rarely been isolated from incriminated
foods or water, although Yersinia may be successfully isolated from clinical specimens. Isolation of a
pathogenic biotype from a ready-to-eat food can implicate that food as a vehicle of transmission in a
Yersinia outbreak.
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Ontario Public Health Standards and Supporting Legislation
• Ontario Public Health Standards, 2008. ‘Beach Management Protocol’, 2008 or as current.
http://www.health.gov.on.ca/en/pro/programs/publichealth/oph_standards/ophsprotocols.aspx
• Ontario Public Health Standards, 2008. ‘Drinking Water Protocol’, 2008 or as current
http://www.health.gov.on.ca/en/pro/programs/publichealth/oph_standards/ophsprotocols.aspx
• Ontario Public Health Standards, 2008. ‘Food Safety Protocol’, 2008 or as current.
http://www.health.gov.on.ca/en/pro/programs/publichealth/oph_standards/ophsprotocols.aspx
• Ontario Public Health Standards, 2008. ‘Recreational Water Protocol’, 2008 or as current
http://www.health.gov.on.ca/en/pro/programs/publichealth/oph_standards/ophsprotocols.aspx
• Health Protection and Promotion Act, 1990. Ontario Regulation 318/08 (Transitional - Small Drinking Water Systems.
http://www.e-laws.gov.on.ca/html/regs/english/elaws_regs_080318_e.htm
• Health Protection and Promotion Act, 1990. Ontario Regulation 319/08 (Small Drinking Water Systems.
http://www.e-laws.gov.on.ca/html/regs/english/elaws_regs_080319_e.htm
• Health Protection and Promotion Act, 1990. Ontario Regulation 428/05 (Public Spas).
http://www.e-laws.gov.on.ca/html/regs/english/elaws_regs_050428_e.htm
• Health Protection and Promotion Act, 1990. Regulation 562 (Food Premises).
http://www.e-laws.gov.on.ca/html/regs/english/elaws_regs_900562_e.htm
• Health Protection and Promotion Act, 1990. Regulation 565 (Public Pools).
http://www.e-laws.gov.on.ca/html/regs/english/elaws_regs_900565_e.htm
• Ontario Water Resources Act, 1990. Regulation 903 (Wells).
http://www.e-laws.gov.on.ca/html/regs/english/elaws_regs_900903_e.htm
• Safe Drinking Water Act, 2002.
http://www.e-laws.gov.on.ca/html/statutes/english/elaws_statutes_02s32_e.htm
• Safe Drinking Water Act, 2002. Ontario Regulation 169/03 (Ontario Drinking Water Quality Standards).
http://www.e-laws.gov.on.ca/html/regs/english/elaws_regs_030169_e.htm
• Safe Drinking Water Act, 2002. Ontario Regulation 170/03 (Drinking Water Systems).
http://www.e-laws.gov.on.ca/html/regs/english/elaws_regs_030170_e.htm
• Safe Drinking Water Act, 2002. Ontario Regulation 243/07 (Schools, Private Schools and Day Nurseries).
http://www.e-laws.gov.on.ca/html/regs/english/elaws_regs_070243_e.htm
• Safe Drinking Water Act, 2002. Ontario Regulation 248/03 (Drinking Water Testing Services).
http://www.e-laws.gov.on.ca/html/regs/english/elaws_regs_030248_e.htm
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P ublic Healt H insP ec t o r ’s G uiD e t o t He P r inc iP les anD P r ac t ic es o f env ir o nm ent al m ic ro bio lo G y
Supporting Documentation/References
• APHA. 2001. Compendium of Methods for the Microbiological Examination of Foods. 4th ed. American Public Health Association,
Washington, D.C.
• APHA. 2012. Standard Methods for the Examination of Water and Wastewater. 22nd ed. American Public Health Association,
Washington. D.C.
http://www.standardmethods.org
• Barbaree, J.M., G.W. Gorman, W.T. Martin, B.S. Fields, and W.E. Morrill. 1987. Protocol for sampling environmental sites for Legionella.
Appl. Environ. Microbiol. 53:1454-1458.
• Canadian Food Inspection Agency Policy on the Control of Listeria monocytogenes in Ready-to-Eat (RTE) Meat and Poultry Products
http://www.inspection.gc.ca/english/fssa/meavia/man/ch4/annexhe.shtml
• Food and Drug Administration, USA, 1998. Bacteriological Analytic Manual, 8th edition.
http://www.fda.gov/Food/ScienceResearch/LaboratoryMethods/BacteriologicalAnalyticalManualBA M/default.htm.
Accessed February 14, 2010.
• Health Canada. 2004. Fungal Contamination in Public Buildings: Health Effects and Investigation Methods.
http://hc-sc.gc.ca/ewh-semt/pubs/air/fungal-fongique/index-eng.php
• Health Canada. 2004. Policy on Listeria monocytogenes in Ready-to-Eat Foods. FD-FSNP 0071
http://www.hc-sc.gc.ca/fn-an/legislation/pol/policy_listeria_monocytogenes_2011-eng.php
• Health Canada. 1999. The Compendium of Analytical Methods.
http://www.hc-sc.gc.ca/fn-an/res-rech/analy-meth/microbio/index-eng.php
• Health Inspector’s Guide to the Principles and Practices of Environmental Bacteriology – 1989 – Editor: M.H.Brodsky, Chief
Environmental Bacteriology
• Health Inspector’s Guide to the Principles and Practices of Environmental Microbiology – 1997– Editor: M.H.Brodsky, Chief
Environmental Bacteriology
• Heymann DL, ed. (2008). Control of Communicable Diseases manual, 19th ed. Washington, DC, American Public Health Association
• Instructions For Official Agencies Submitting Water Samples to the Public Health Ontario Laboratories
http://www.oahpp.ca/services/water-testing.html
• Laboratory Guide for Gastroenteritis Outbreaks (2008) Public Health Laboratories, Ontario Agency for Health Protection and Promotion
– see website for most current version
http://www.oahpp.ca/services/documents/specimen-collection-guide/gastro_full_20080301.pdf
• Ministry of the Environment. Practices for the Collection and Handling of Drinking Water Samples.
http://www.portal.gov.on.ca/drinkingwater/stel01_046911.pdf
• PHO; Legionella workshop, Questions and Answers, 2012
http://www.oahpp.ca/resources/documents/presentations/2012june25/PHO%20Legionella%20Workshop%20June%202012%20-%20Q+A.pdf
• SMS Environmental.
http://www.sms-environmental.co.uk/swimming_pool_water.html#pseudomonas
• Specimen Collection Guide – Public Health Ontario Laboratories
http://www.oahpp.ca/services/specimen-collection-guide.html
• WHO; Legionella and the prevention of legionellosis, 2007
http://www.who.int/water_sanitation_health/emerging/legionella.pdf
• World Health Organization (2008). Foodborne Disease Outbreaks: Guidelines for Investigation and Control. Geneva, Switzerland, 2008
http://www.who.int/foodsafety/publications/foodborne_disease/fdbmanual/en/index.html
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Contributors
Dr. vanessa allen – Medical Microbiologist, Enteric,
Environmental, Molecular Surveillance, STI and Syphilis,
PHO
Peter boleszczuk – Head Technologist, Environmental
Microbiology, PHO
Helen Doyle – Manager of Environmental Health.
Community and Health Services Department, Regional
Municipality of York
sandy edelsward – Program Coordinator, Drinking Water
Testing, PHO
Dr. anna majury – Regional Microbiologist and Assistant
Professor, PHO
anne maki – Manager, Enteric, Environmental, Molecular
Surveillance and STI, PHO
allana murphy – Operational Lead, Environmental
Laboratory, PHO
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Appendices
Appendix A: Supplies
Board of health staff should ensure that environmental
sample collection and submission supplies are readily
available. If further supplies are required, they can be
obtained from the PHO – Regional Laboratory or from the
Toronto Laboratory Warehouse at 416-235-5820. Supplies
will be shipped within 1 – 5 days of request. Catalogue
numbers are as follows:
Water:
cat.no.
item
format
L-4020
Bacteriological Analysis of Water – Single Sample Requisition for Official Agencies
Pkg. of 125
L-3172
Bacteriological Analysis of Water – Multiple Sample Requisition for Official Agencies
Pkg. of 125
N-0040
Kit – Water – Bacteriology Test Private Citizen Drinking Water Collection Kits
Box of 125
TC 125-B
Sewage Collection Kits
Blue Transport Bag
M-5030
Bag – Stomacher
Sterile plastic sample bag with round wire closure
Request amount required
L-8001
Seal – Regulated water
Pkg. of 200
L-8002
Seal – Legal
Pkg. of 50
item
format
L-3180
Form – Food Bacteriology
Pkg. of 125
M-5030
Bag – Stomacher
Sterile plastic sample bag with round wire closure
Box of 250
food:
cat.no.
environmental swabs:
cat.no.
item
format
L-4004
Data Sheet – Environmental Swab Test
Environmental Bacteriology Swab Tests requisition
Pkg. of 250
Contact the PHOL
– Toronto,
Environmental
Laboratory at
416.235.5716
Environmental Swabs (test requisition included)
As requested
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Appendix B: Laboratory Submission Forms
Refer to the PHO website for the most current version of
the forms listed below.
http://www.oahpp.ca/resources/laboratory-materials.html
a. Bacteriological Analysis of Water – Single Sample
Requisition for Official Agencies
b. Bacteriological Analysis of Water – Multiple Sample
Requisition for Official Agencies
c. Bacteriological Analysis of Drinking Water for Private
Citizen, SINGLE HOUSEHOLD ONLY
D. Food Bacteriology
e. Environmental Bacteriology Swab Tests
f. Potability of Water Release Form
G. Autorisation De Divulguer Des Renseignements Sur La
Potabilité De L’eau
H. Ontario Agency for Health Protection and Promotion
Legal Seal
i. Ontario Agency for Health Protection and Promotion
Regulated Water Seal
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a. bacterioloGical analysis of Water – sinGle samPle requisition for official aGencies
Barcode
Date/Time Received
PHL No.
Bacteriological Analysis of Water – Single Sample Requisition for Official Agencies
FOR DRINKING WATER: THE REGULATION STATUS OF THE SAMPLE MUST BE INDICATED. IF REGULATED, ALL NON-SHADED AREAS MUST BE COMPLETED
OR THE SAMPLE WILL NOT BE ANALYZED BY THE LABORATORY AND ANOTHER SAMPLE WILL HAVE TO BE SUBMITTED.
A unique identifier (i.e. barcode) must be present on both the bottle and requisition when received at the laboratory or the sample will not be processed
Official Agency Address
Owner of the Water Supply
Agency Name:
Facility:
Bldg. No., Street, R.R., Box No.
Bldg. No., Street, R.R., Box No.
City, Town
City, Town
Postal Code
Province
Submitted By:
Submitted To:
Reason for Sampling
Sample Information-Drinking Water
Date Collected:
hh : mm
Time Collected:
Collected By:
AM
PM
Tel: (Working hrs.):
Fax:
Waterworks No.:
Type of Drinking Water Systems
(Check all applicable boxes)
Contact Name(s):
Public Health Lab
Comments/Additional Information:
Mandatory: tick one box
HPPA O. Reg 319/08
HPPA Regulated Premises
Non Regulated
Private Residence
SDWA O. Reg 170/03
Postal Code
Province
(
Control
Audit
Compliance
Resample
Complaint Investigation
Outbreak Investigation
Not assigned Assigned Outbreak Investigation
)
Etiological Agent if Known:
Test for Etiological Agent*
*Call laboratory before sampling
Ground Water (i.e. well) Cistern
Surface Water
Distribution
Bottled Water
Other:
Free or combined chlorine residual (mg/L):
Treatment
Non-treated
Treated
Tests Performed
∆
Date Collected:
Time Collected:
Collected By:
Escherichia coli (Cfu) per 100mL*
Date Read
100mL*
AM
PM
Public Beach
(
Circle
one
)
Recreational Water Facility /
Hydrotherapy Pool
Suspected Sewage Contamination
Other * (Please specify):
*Call laboratory before sampling
For Regulated Drinking Water or Legal Samples:
Relinquished By:
Relinquished By:
Date:
∆
Background (Cfu) per
Reported
By
hh : mm
Sampling Site:
HPC test Requested ∆
Total Coliforms (Cfu) per 100mL*
Count
Or, Test for Potability
Sample Information-Non-Potable
All potable samples must be <25°C when received at the lab. Samples must be received in the lab within 6
hours of collection if not refrigerated. Refrigerated non-potable samples must be received in the laboratory within
1 calendar day of collection and all drinking water must be received in the laboratory within 48 hours of collection
(time of collection must be indicated).
For Laboratories Only
if assigned, indicate number
Outbreak Number:
Source of Drinking Water
Circle
one
Sampling Site:
(After Hours)
For Lab Use:
(Print Name)
Time:
(Signature)
Date:
Received By:
Time:
Laboratory Comments / Date Reported
Pseudomonas aeruginosa (Cfu) per 100mL*
Staphylococcus aureus (Cfu) per 100mL*
Heterotrophic plate count (HPC) (Cfu) per mL**
Other:
per
mL
∆
4318-44 (12/11)
Presumptive Staphylococci (Cfu) per 100mL*
Authorized By:
(Technologist)
*Analysis by Membrane Filtration: **Analysis by Spread Plate. = Accredited test (drinking water)
These results relate only to the sample tested.
This information is being collected in compliance with the requirements of the Safe Drinking Act, 2002 and it’s regulations, and it will only be used for the purposes for which it is collected.
Public Health Ontario is subject to the Freedom of Information and Protection of Privacy Act and any information in its records may be subject to disclosure by the Ministry pursuant to the
Freedom of Information and Protection of Privacy Act.
∆
End of report
P ublic Hea lt H insP ec t o r ’s G uiD e t o t He P rinc iP les anD P r a c t ic es o f env iro nm ent al mic ro bio lo G y
41
42
4321-44 (05/12)
if assigned, indicate number
Waterworks No.:
Fax:
Tel: (After hrs.):
Tel: (Working hrs.):
Contact Name(s):
Province
City, Town
Bldg. No., Street, R.R., Box No.
Facility
Owner of the Water Supply
Postal Code
(
Circle
one
)
AM
PM
(
Circle
one
*Call laboratory before sampling
Other* (Please specify)
Suspected Sewage Contamination
Recreational Water Facility / Hydrotherapy Pool
Public Beach
Sampling site:
Collected by:
Time Collected: hh : mm
Date Collected:
)
Sample Information - Non-Potable
*Call laboratory before sampling
Test for Etiological Agent*
Or, Test for Potability
Etiological Agent if Known:
Control
Audit
Compliance
Resample
Complaint Investigation
Outbreak Investigation
(complete section below)
Outbreak Number:
Reason for Sampling
Mandatory: tick one box
HPPA O. Reg 319/08
HPPA Regulated Premises
Non Regulated
Private Residence
SDWA O. Reg 170/03
Type of Drinking Water Systems
Ground Water (i.e. well)
Cistern
Surface Water
Distribution
Bottled Water
(Please Specify)
Other:
Treatment
Non-treated
Treated
Source of Drinking Water
Sampling Site:
Collected By:
AM
Time Collected: hh : mm PM
Date Collected:
Ñ = Accredited test (drinking water)
Assigned
Not assigned
Postal Code
Public Health Lab
(Please Print)
Comments/Additional Information:
Submitted to:
Submitted by:
Province
City, Town
Bldg. No., Street, R.R., Box No.
Agency Name
Barcode
Time:
(Signature)
(Print Name)
Time:
Received By:
Authorized By:
(Technologist)
Date:
For Lab Use:
Date
Read:
Reported
By:
✓
Free or
HPC
combined
chlorine Requested
residual
(mg/L)
Ñ Total
Coliforms
E. coli
Ñ
Background
Ñ
Date Reported:
P. aeruginosa Presumptive S. aureus
Staphylococci
on 1mL. Vol.
by Spread Plate
Bacterial Counts (Cfu) Based on 100mL Vol. By Membrane Filtration Ñ HPC (Cfu) Based
End of report
Lab. No./
Date & Time Received
Sampling Precautions:
- All potable samples must be <25°C when received at the lab. Samples must be received in
the lab within 6 hours of collection if not refrigerated.
- Refrigerated non-potable samples must be received in the laboratory within 1 calendar day
of collection.
- All drinking water must be received in the laboratory within 48 hours of collection.
- Time of collection must be indicated.
These results relate only to the sample tested. This information is being collected in compliance with the requirements of the Safe Drinking Act, 2002 and its regulations,
and it will only be used for thepurposes for which it is collected. Public Health Ontario is subject to the Freedom of Information and Protection of Privacy Act and any information
in its records may be subjectto disclosure by the Ministry pursuant to the Freedom of Information and Protection of Privacy Act.
Date of Analysis:
Laboratory Comments:
Date:
Relinquished By:
Relinquished By:
For Regulated Drinking Water or Legal Samples:
Identification of
Collection Site
& Time Collected
FOR DRINKING WATER: THE REGULATION STATUS OF THE SAMPLE(S) MUST BE INDICATED. IF REGULATED, ALL NON-SHADED AREAS MUST BE COMPLETED OR THE
SAMPLE(S) WILL NOT BE ANALYZED BY THE LABORATORY AND ANOTHER SAMPLE(S) WILL HAVE TO BE SUBMITTED. A UNIQUE IDENTIFIER (I.E. BARCODE) MUST BE
PRESENT ON BOTH THE BOTTLE AND REQUISITION WHEN RECEIVED AT THE LABORATORY OR THE SAMPLE(S) WILL NOT BE PROCESSED
Official Agency Address
Sample Information - Drinking Water
Multiple Sample Requisition for Official Agencies
Bacteriological Analysis of Water
b. bacterioloGical analysis of Water – multiPle samPle requisition for official aGencies
P ublic Healt H insP ec t o r ’s G uiD e t o t He P r inc iP les anD P r ac t ic es o f env ir o nm ent al m ic ro bio lo G y
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Appendix C: Major Foodborne Diseases: General Features
Information extracted from World Health Organization (2008). Foodborne Disease Outbreaks: Guidelines for Investigation
and Control. Geneva, Switzerland, 2008
table 18: major foodborne Diseases – clinical features, mode of transmission and associated High risk foods
etioloGic aGent
Bacillus cereus
(Bacterial toxin)
incubation PerioD
symPtoms
a) Diarrheal syndrome:
8 – 16 hours
a) Diarrheal syndrome: acute diarrhea, nausea and abdominal pain.
b) Emetic syndrome:
1 – 5 hours
b) Emetic syndrome: acute nausea, vomiting and abdominal pain and
sometimes diarrhea.
mode of transmission and associated foods: Ingestion of food that has been stored at ambient
temperatures after cooking, permitting the growth of bacterial spores and toxin production. Many
outbreaks (particularly those of the emetic syndrome) are associated with cooked or fried rice that has
been kept at ambient temperature.
foods involved include: Starchy products such as boiled or fried rice, spices, dried foods, milk, dairy
products, vegetable dishes, and sauces.
Campylobacter jejuni
Typically 2 – 5 days
(range 1 – 11 days)
Fever, severe abdominal pain, nausea and diarrhea which can vary from
slight to profuse and watery sometimes containing blood or mucus.
mode of transmission and associated foods: Principally through ingestion of contaminated food. Main
food sources are raw milk and raw or undercooked poultry. Spread to other foods by cross-contamination
or contamination with untreated water; contact with animals or birds. Other sources of transmission are
contact with live animals (pets and farm animals). Person-to-person transmission occurs during the
infectious period that ranges from several days to several weeks.
foods involved include: Raw milk, poultry, beef, pork and drinking-water.
Clostridium botulinum
12 – 36 hours
(range several hours to 8
days)
Vomiting, abdominal pain, fatigue, muscle weakness, headache, dizziness,
ocular disturbance (blurred or double vision, dilated pupils, unreactive to
light), constipation, dry mouth and difficulty in swallowing and speaking,
and ultimately paralysis and respiratory or heart failure.
mode of transmission and associated foods: Ingestion of toxin pre-formed in food. This may occur when
raw or under-processed foods are stored in anaerobic conditions that allow growth of the organism. Most
outbreaks are due to faulty preservation of food (particularly in homes or cottage industries), e.g. canning,
fermentation, curing, smoking, or acid or oil preservation.
foods involved include: Vegetables, condiments (e.g. pepper), fish and fish products, meat and meat
products, honey, fruit and vegetable juices. Several outbreaks have occurred as a result of consumption of
uneviscerated fish, garlic in oil, and baked potatoes.
Clostridium perfringens
8 – 24 hours
Abdominal pain, diarrhea, rarely vomiting and fever.
mode of transmission and associated foods: Illness usually caused by cooked meat and poultry dishes
subject to time/temperature abuse. Dishes are often left for too long at ambient temperature to cool
down before storage, or cooled inadequately. This allows spores that survive the cooking process to
germinate and grow, producing large numbers of vegetative cells. If a dish is not reheated sufficiently
before consumption, the vegetative cells can cause illness.
foods involved include: Meat and poultry (boiled, stewed or casseroled).
Cryptosporidium parvum
2 – 4 days
Persistent diarrhea, nausea, vomiting and abdominal pain, sometimes
accompanied by an influenza-like illness with fever.
mode of transmission and associated foods: Spread through the fecal-oral route, person-to-person
contact or consumption of fecally contaminated food and water, bathing in contaminated pools.
foods involved include: Raw milk, drinking-water and apple cider.
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table 18: major foodborne Diseases – clinical features, mode of transmission and associated High risk foods ...continued
etioloGic aGent
incubation PerioD
symPtoms
a) Enteropathogenic
E. coli (EPEC)
a) EPEC: 1 – 6 days; as
short as 12 – 36 hours
a) EPEC adheres to the mucosa and changes its absorption capacity, causing
vomiting, diarrhea, abdominal pain and fever.
b) Enterotoxigenic
E. coli (ETEC),
producing a heat-labile
LT) and a heat-stable
(ST) enterotoxin
b) ETEC: 1 – 3 days; as
short as 10–12 hours
b) ETEC mediates its effects by enterotoxins. Symptoms include diarrhea
(ranging from mild to a severe, cholera-like syndrome), abdominal
cramps and vomiting, sometimes leading to dehydration and shock.
c) Enteroinvasive
E. coli (EIEC)
c) EIEC: 1 – 3 days; as
short as 10–18 hours
c) EIEC causes inflammatory disease of the mucosa and submucosa by
invading and multiplying in the epithelial cells of the colon. Symptoms
include abdominal pain, vomiting and watery diarrhoea (in <10% of cases
stools may become bloody and contain mucus).
escherichia coli
mode of transmission and associated foods: a – c) EPEC, ETEC, EIEC: consumption of food and water
contaminated with fecal matter. Time/temperature abuse of such foods increases risk of illness. Up to 25%
of infections in infants and young children in developing countries are due to E. coli, in particular ETEC and
EPEC (10 – 20% and 1 – 5% of cases at treatment centres, respectively). ETEC is a major cause of traveller’s
diarrhea in developing countries.
d) Enterohaemorrhagic
E. coli (EHEC) or
verocytotoxinproducing E. coli
(VTEC), also referred to
as Shiga-toxin
producing E. coli
(STEC), of which the
most commonly
recognized is E. coli
O157.
d) EHEC: 3 – 8 days,
median of 4 days
d) EHEC causes abdominal cramps and watery diarrhea that may also
develop into bloody diarrhea (hemorrhagic colitis). Fever and vomiting
may also occur.
mode of transmission and associated foods: d) EHEC is transmitted mainly through consumption of foods
such as raw or undercooked ground-meat products and raw milk from infected animals. Fecal contamination
of water and other foods, as well as cross-contamination during food preparation, will also lead to infection.
foods involved with the transmission of vtec: Include ground (minced) meat, raw milk, and vegetables.
Secondary transmission (person-to-person) may also occur during the period of excretion of the pathogen
which is less than a week for adults but up to 3 weeks in one-third of affected children.
giardia lamblia
7 – 10 days
(range 4 – 25 days)
Diarrhea (which may be chronic and relapsing), abdominal cramps, fatigue,
weight loss, anorexia and nausea. Symptoms may be caused by a protein
toxin.
mode of transmission and associated foods: Infected individuals excrete Giardia cysts in large numbers.
Illness is spread by fecal-oral route, person-to-person contact or fecally contaminated food and water.
Cysts have been isolated from lettuces and fruits such as strawberries. Infection also associated with
drinking-water from surface waters and shallow wells.
foods involved include: Water, home-canned salmon, fruit and vegetables and noodle salad.
Hepatitis a
25 – 28 days
(range 2 – 6 weeks)
Loss of appetite, fever, malaise, abdominal discomfort, nausea and
vomiting, followed by symptoms of liver damage (passage of dark urine,
pale stools, jaundice).
mode of transmission and associated foods: Spread by fecal-oral route, primarily person-to-person. Can
also be transmitted through food and water as a result of sewage contamination or infected foodhandlers. Risk of transmission is greatest during the second half of the incubation period until a few days
after the appearance of jaundice.
foods involved include: Shellfish, raw fruit and vegetables, bakery products.
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table 18: major foodborne Diseases – clinical features, mode of transmission and associated High risk foods ...continued
etioloGic aGent
Listeria monocytogenes
incubation PerioD
symPtoms
3 – 70 days
Influenza-like symptoms such as fever, headache and occasionally
gastrointestinal symptoms.
mode of transmission and associated foods: A substantial proportion of cases of listeriosis are foodborne.
foods involved include: Raw milk, soft cheese, meat-based paste, jellied pork tongue, raw vegetables and
coleslaw.
Salmonella species
6 – 48 hours, occasionally
up to 4 days
The principal symptoms are fever, headache, nausea, vomiting, abdominal
pain and diarrhea.
mode of transmission and associated foods: Main route of transmission is by ingestion of the organisms
in food (milk, meat, poultry, eggs) derived from infected food animals. Food can also be contaminated by
infected food-handlers, pets and pests, or by cross-contamination as a result of poor hygiene.
Contamination of food and water from the feces of an infected animal or person may also occur. Problems
caused by initial contamination may be exacerbated by prolonged storage at temperatures at which the
organism may grow. Direct person to-person transmission may also occur during the course of the
infection.
foods involved include: Unpasteurized milk, raw eggs, poultry, meat, spices, salads and chocolate.
Salmonella typhi and
Salmonella paratyphi
types a–c.
10 – 20 days (range 3 days
to 8 weeks)
Systemic infections characterized by high fever, abdominal pains, headache,
vomiting, diarrhea followed by constipation, rashes and other symptoms of
generalized infection.
mode of transmission and associated foods: Ingestion of food and water contaminated with fecal
matter. Food-handlers may carry the pathogen and be a source of food contamination. Secondary
transmission may also occur.
food associated with salmonella typhi include: Prepared foods, dairy products (e.g. raw milk), meat
products, shellfish, vegetables, and salads.
Shigella dysenteriae
S. flexneri, S. boydii,
S. sonnei
1 – 3 days, up to 1 week
for S. dysenteriae
Abdominal pain, vomiting, fever, diarrhea ranging from watery (S. sonnei) to
dysenteric with bloody stools, mucus and pus (S. dysenteriae and, to a lesser
extent S. flexneri and S. boydii).
mode of transmission and associated foods: Food and water contaminated with fecal matter. Person-toperson transmission through the fecal–oral route is an important mode of transmission. Food can be
contaminated by food-handlers with poor personal hygiene or by use of sewage/wastewater for
fertilization.
foods involved include: Uncooked foods that have received extensive handling, such as mixed salads and
vegetables, water and raw milk.
Staphylococcus aureus
2 – 6 hours
Intoxication, sometimes of abrupt and violent onset. Severe nausea,
cramps, vomiting and prostration, sometimes accompanied by diarrhea.
mode of transmission and associated foods: Consumption of foods containing the toxin. Foods are
contaminated by food-handlers. If storage conditions are inadequate, the bacteria may multiply to
produce toxin. Intoxication is often associated with cooked food e.g. meat, in which competitive bacteria
have been destroyed.
foods involved include: Prepared foods subject to handling in their preparation (ham, chicken and egg
salads, cream-filled products, ice cream, cheese).
Vibrio cholerae
o1 and o139
1 – 3 days
Profuse watery diarrhea, which can lead to severe dehydration, collapse and
death within a few hours unless lost fluid and salt are replaced; abdominal
pain and vomiting.
mode of transmission and associated foods: Food and water contaminated through contact with fecal
matter or infected food handlers. Contamination of vegetables may occur through sewage or wastewater
used for irrigation. Person-to-person transmission through the fecal-oral route is also an important mode
of transmission.
foods involved include: Seafood, vegetables, cooked rice and ice.
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table 18: major foodborne Diseases – clinical features, mode of transmission and associated High risk foods ...continued
etioloGic aGent
Vibrio parahaemolyticus
incubation PerioD
symPtoms
9–25 hours, up to 3 days
Profuse watery diarrhea, abdominal pain, vomiting, and fever. A dysenteric
syndrome has been reported from some countries, particularly Japan
mode of transmission and associated foods: Associated with consumption of raw or undercooked fish
and fishery products or cooked foods subject to cross contamination from raw fish.
Vibrio vulnificus
12 hours – 3 days
Profuse diarrhea with blood in stools. Organism is associated with wound
infections and septicaemia may originate from the gastrointestinal tract or
traumatized epithelial surfaces.
mode of transmission and associated foods: All known cases are associated with seafood, particularly
raw oysters.
Yersinia enterocolitica
24 – 36 hours
(range 1 – 11 days)
Abdominal pain, diarrhea, mild fever, sometimes vomiting
mode of transmission and associated foods: Illness is transmitted through consumption of pork products
(tongue, tonsils, gut), cured or uncured, as well as milk and milk products.
viral gastroenteritis
Many different viruses
can cause viral
gastroenteritis, including
adenoviruses,
coronaviruses,
rotaviruses, parvoviruses,
caliciviruses and
astroviruses. Those
viruses most commonly
associated with
foodborne outbreaks are
norovirus and hepatitis A.
15 – 50 hours
Diarrhea and vomiting, which is often severe and projectile with sudden
onset.
mode of transmission and associated foods: Gastroenteritis viruses usually spread by fecal–oral route.
Food and drinking-water may be contaminated either at source when exposed to sewage/wastewater in
the environment or used for irrigation, or by an infected food-handler. Filter-feeding shellfish most
common food contaminated at source, but a wide range of different cooked and uncooked foods have
been implicated in secondary contamination by food-handlers.
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Appendix D: List of Tables
table 1: Food – Microbiological Analysis Test Menu
– Indicator Bacteria
table 2: Food – Microbiological Analysis Test Menu
– Foodborne Pathogens
table 3: Food – Microbiological Analysis Test Menu
– Physiochemical Tests
table 4: Food – Microbiological Analysis Test Menu
– Molecular Typing
table 5: Food – Microbiological Analysis Test Menu
– Referred Tests
table 6: Food – Collection and Transportation Instructions
table 7: Food – Laboratory Rejection Criteria
table 8: Sampling Instruction for Environmental Swabs in
Foodborne Outbreak Investigations
table 9: Water – Microbiological Analysis Test Menu
table 10: Water – Microbiological Analysis Test Menu
– Pathogen Specific Analyses
table 11: Water – Test Requisition Requirements
table 12: Water – Collection and Transportation
Instructions
table 13: Water – Laboratory Rejection Criteria
table 14: Suspected Sewage Contamination Samples –
Collection and Transportation Instructions
table 15: Legionella – Microbiological Analysis Test Menu
table 16: Legionella – Environmental Sample Collection and
Transportation Instructions
table 17: Interpretation of Environmental Microbiological
Test Results
table 18: Major Foodborne Diseases - Clinical Features,
Mode of Transmission and Associated High Risk
Foods
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Notes
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february 2013