Specimen Collection and Handling

Specimen Requirements

Test Requested Required Requisition(s) Specimen Type Minimum Volume Collection Kit

E. coli

Suspected Sewage Contamination Water is water suspected of being contaminated by fecal material. Samples are submitted by a public health unit only.

200 mL (to the fill line on the bottle)

Water is collected in a PHO laboratory water collection bottle. Other collection bottles are not accepted. Samples must be submitted in the Sewage Sample Collection Kit: a blue laboratory sewage transportation bag (TC-125-1B) labelled with a “SEWAGE SAMPLES ONLY” tag.

Submission and Collection Notes


Complete the required information on the requisition (i.e. submitting agency name including the sub office if this office submitted the sample, date and time collected and sample type). 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.


Remove one barcode from the bottle and apply it to the top copy of the requisition in the “Barcode” field. Remove a second barcode and apply it in the corresponding location on the second copy of the form. Samples will not be processed if the barcodes are not affixed to the requisition. If the barcode is not able to be removed, write the barcode number in the “Barcode” box on the form.


Examine the lid of the bottle. If the tamper evident ring has separated from the cap use another bottle.


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.
  • 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 seek medical attention.

Wearing rubber/latex gloves collect the suspected sewage contaminated sample to the 200 mL fill line.

  • Do not overfill.
  • If overfilled, remove some of the water so it the bottle is filled to 200 mL line.

Tighten the cap on the collection bottle. Leaking samples will not be processed; a new sample and requisition will be required.


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.


Remove and discard rubber/latex gloves. Do not put the used gloves in the plastic bag along with sewage sample. Wash/sanitize your hands.


Seal the sewage sample bottle in the plastic bag supplied.


Put the completed requisition in the side pocket of the plastic bag containing the sewage sample. Do not place the requisition inside the plastic bag with the sample.


Place the packaged sewage sample 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-125-1B).  Do not send sewage samples in any other transport container or with other environmental or medical samples, as sample integrity cannot be ensured.

Timing of Specimen Collection

Ensure adequate time to collect and transport the sample to the laboratory. It must be tested within one calendar day of collection.


Samples are submitted by a public health unit only. Samples will not be tested if the acceptance criteria are not met. Refer to the Public Health Inspector’s Guide to Environmental Microbiology Laboratory Testing.

Storage and Transport

Retain a copy of the completed requisition that includes the barcodes. Specimens should be stored at 2-8 °C following collection and shipped to PHO laboratory in a blue laboratory sewage transport bag (TC-125-1B). Pack with frozen ice packs. Samples received at the laboratory above 25 °C will not be tested. Do not freeze samples.

Samples must be shipped within 24 hours of collection and tested within one calendar day.

For health and safety reasons, any sample identified as “Suspected Sewage Contamination” will not be accepted unless received in the Sewage Sample Collection Kit. A new sample should be collected and submitted using the proper requisition and kit.

Special Instructions

This test is only offered to public health unit clients.

Instructions For Official Agencies Submitting Water Samples to the Public Health Ontario Laboratory provides step by step instructions for completing the documentation and submitting/relinquishing the sample. For legal samples the chain of custody must be complete when the sample is received at the laboratory (Refer to Instructions For Official Agencies Submitting Water Samples to the Public Health Ontario Laboratory).


Requisitions and Kit Ordering

Test Frequency and Turnaround Time (TAT)

Samples are routinely accepted at the laboratory Monday – Friday during regular operating hours. Please contact the PHO laboratory Customer Service Centre prior to the submission of samples that will be received outside of these hours.

The turnaround time is up to 4 days for E. coli.

STAT and Critical Samples Testing

Please contact the PHO laboratory Customer Service Centre prior to the submission of samples that will be received outside of regular operating hours or the PHO Laboratory Duty Officer at (416) 605-3113 if outside of Customer Service’s hours. Samples of an urgent nature (e.g. STAT) will be processed and read with no undue delay.

Test Methods

Specimens are tested by the Membrane Filtration method (modified from the American Society for Microbiology’s Bacteriological Examination of Waters: Membrane Filtration Protocol.

By submitting a sample with a PHO Laboratory requisition, the submitter accepts Public Health Ontario’s methodology.


Results are reported to the submitter as indicated on the requisition.

Additional Information

For general inquiries, please contact the Public Health Ontario Laboratory Customer Service Centre.

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Updated 13 June 2022