Yersinia pestis – Confirmation

Consistent with O. Reg. 671/92 of the French Language Services Act, laboratory testing information on this page is only available in English because it is scientific or technical in nature and is for use only by qualified health care providers and not by members of the public.

Background
This page provides details on the identification of Yersinia pestis at Public Health Ontario (PHO). Y. pestis is the etiological agent of plague including bubonic, pneumonic, septicemic, pharyngeal, and meningeal plague.^{1, 2}Plague is a disease of public health significance (DoPHS) in Ontario and additional management considerations are found in the Ontario Public Health Standards Appendix 1 for Disease: Plague.

For information regarding other testing options, refer to the following PHO webpages:

Aerobic bacteria identification: Bacterial cultures – Aerobic – Reference ID/Confirmation
Aerobic bacteria susceptibility testing: Antimicrobial Susceptibility Testing – Aerobic Bacteria
Suspicious packages (for investigation of Agents of) Bioterrorism: Suspicious Packages (for Investigation of Agents of) Bioterrorism – RT PCR.

Note: Yersinia pestis serology testing is no longer available through PHO.

*Uncontrolled print copy. Valid only on day of print: 27 Oct 2026.

Testing Indications

Submitting laboratories must contact PHO’s Customer Service Centre by telephone at 416-235-6556 or 1-877-604-4567 for pre-approval.

Plague may be suspected clinically based on the following: ^1,2

Clinically compatible acute febrile illness with formation of painful buboes (lymphadenitis), severe respiratory symptoms, septicemia, and/or meningitis.
Travel to an endemic area or area experiencing an outbreak.
Relevant exposure history, e.g., infected fleas, infected or contaminated animals or animal products (animal husbandry, hunting, butchery, veterinary work), propagated material (laboratory workers, bioterrorism), or close contact of infected person.

Laboratory suspicion for Y. pestis should be raised based on the following: ^1,2,3

Relevant specimen source (e.g. lymph node aspirate, blood culture, respiratory specimen, CSF)
Gram-negative rod (usually plump, occurring singly or in pairs, may or may not show bipolar or “safety pin” appearance)
Non-motile
Slow growth on blood agar (absent or pinpoint colonies at 24 hours, 1-2 mm non-hemolytic whitish to slightly yellow colonies at 48 hours, and may have a “fried egg” appearance)
Non-lactose fermenter (colourless) on MacConkey agar
Positive for catalase and negative for oxidase, urea, and indole (performed in a biosafety cabinet)

Acceptance/Rejection Criteria

Acceptable specimens include:

Pure cultures are preferred
Properly labeled cultures: The patient and specimen identification on the requisition form must match the label on the culture container
Source of isolation indicated: The requisition must clearly state the source of the isolate (e.g. skin lesion swabs, blood cultures, respiratory secretions, stool)
Mixed cultures with prior approval from a PHO Microbiologist. Refer to Submission and Collection notes.
Primary specimen with prior approval from a PHO Microbiologist. Refer to Submission and Collection notes.

Rejected specimens include:

Mislabeled or unlabeled isolates
Missing specimen source information: Cultures without a clearly indicated isolate source on the requisition will be rejected
Non-viable cultures will not be tested
Mixed cultures (unless pre-approved by a PHO Microbiologist). Refer to Submission and Collection note below for details. If approval is not received and the sample is still sent, the submitting laboratory will be contacted stating the rejection
Primary clinical specimens (unless pre-approval by a PHO Microbiologist). Refer to Submission and Collection note below for details.

Specimen Collection and Handling

Specimen Requirements

Test Requested	Required Requisition(s)	Specimen Type	Minimum Volume	Collection Kit
Yersinia pestis – confirmation	Reference Bacteriology Requisition	Pure viable subculture of organism on appropriate media that supports the growth (e.g. Blood Agar)	N/A	Columbia sheep blood agar plate or other appropriate media

Submission and Collection Notes

Complete all fields of the requisition form, including:

Test(s) requested: select confirmation and identification
Culture information: indicate presumptive identification (if available) and Gram morphology; provide oxygen requirement for growth; relevant biochemical test results; indicate presence or absence of growth on MacConkey agar
Clinical diagnosis: indicate if patient is immunocompromised
Date of collection of primary specimen from which the isolate was recovered
Source of isolate

Label the culture container(s) with the patient’s first and last name, date of collection, and one other unique identifier such as the patient’s date of birth or Health Card Number. For additional information see: Criteria for Acceptance of Patient Specimens. Failure to provide this information may result in rejection or testing delay.

Submit only one specimen per patient. If there are multiple specimen sources and prioritization assistance is required, please consult with a PHO Microbiologist.

If pre-approved by a PHO Microbiologist to submit a culture in which suspected Yersinia pestis colonies are mixed with other colonies, ensure that the suspected colonies are clearly identified by marking their location(s) on the exterior of the culture plate (e.g. circle or using arrows with an indelible marker on the outside of the plastic petri dish).

Primary specimen submissions will be cancelled unless pre-approved by at PHO microbiologist. If primary specimen testing cannot be performed locally, please contact PHO’s Customer Service Centre (416-235-6556 or 1-877-604-4567) to discuss specimen selection, collection, appropriate container and transport to PHO’s laboratory with a PHO microbiologist.

Storage and Transport

Submit a fresh subculture to ensure viability on receipt. Place the sealed specimen in a biohazard bag and properly seal the bag. Store at 2-8°C prior to shipping. The requisition form must be placed in the outer pouch of the biohazard bag, separate from the specimen container. Samples should be shipped to PHO’s laboratory within 48 hours of isolation.

Important: All Yersinia pestis culture isolates are considered risk group 3 (RG3) agents and must be shipped in accordance with Category A shipping requirements of the Transportation of Dangerous Good Act.

Special Instructions

Yersinia pestis must be handled in accordance with the Canadian Biosafety Standards and shipped in accordance with the Transportation of Dangerous Goods Regulations.

Prior approval from PHO is required before submitting an isolate. Contact PHO’s Customer Service Centre at 416-235-6556 or 1-877-604-4567 before submitting the isolate.

Requisitions and Kit Ordering

Test Frequency and Turnaround Time (TAT)

Testing is performed daily (Monday to Friday) at PHO’s laboratory, Toronto location. Turnaround time for test results is up to 3 calendar days from receipt at PHO’s laboratory, Toronto location.

Isolates that are identified as Yersinia pestis at PHO are forwarded to the National Microbiology Laboratory (NML) for confirmatory testing. Turnaround time for test results from the NML is up to 4 calendar days for a preliminary result and 14 calendar days for a final result, from the date the sample is received by the NML.

STAT and Critical Specimens Testing

Urgent testing for the CLRN real-time PCR assay at PHO is available upon request by calling the PHO Laboratory Customer Service Centre at 416-235-6556 or 1-877-604-4567.

Test Methods

Yersinia pestis identification is performed at PHO using a laboratory-developed real-time polymerase chain reaction (PCR) test based on protocols developed by the Laboratory Response Network (LRN) in the United States and Canada.

Algorithm

Any isolate positive by the LRN PCR is considered presumptive identification for Yersinia pestis and is forwarded to the National Microbiology Laboratory (NML) for confirmatory testing.

Interpretation

The following table provides possible test results with associated interpretations:

Result	Interpretation
Detected	Yersinia pestis DNA DETECTED by real-time PCR. Presumptive positive. Specimen forwarded to NML for confirmatory testing.
Not Detected	Yersinia pestis DNA NOT DETECTED by real-time PCR. If further identification is required, please contact PHOL Customer Service Centre within 3 days at 1-877-604-4567 or 416-235-6556.
Inconclusive	Yersinia pestis real-time PCR result is inconclusive. Specimen forwarded to NML for further testing. If insufficient specimen remains, and Yersinia pestis is still suspected, submit additional specimen for testing.

Reporting

Results are reported to the ordering physician or authorized health care provider as indicated on the requisition.

Isolates identified as Yersinia pestis are reported to the Medical Officer of Health as per the Health Protection and Promotion Act.

If an isolate was identified as Yersinia pestis at PHO, submitters must follow the Human Pathogens and Toxins Act biosafety and biosecurity requirements pertaining to the declaration, handling, and disposal of Yersinia pestis samples.

References

Kingry LC, Gladney L, Petersen JM. Chapter 42, Yersiniaceae. In: Carroll KC, Pfaller MA, Karlowsky J, Landry ML, McAdam AJ, Patel R, eds. Manual of Clinical Microbiology, 13th ed. Washington, DC: Wiley & Sons Inc.; 2023. p. 1-18.
Public Health Agency of Canada. Pathogen Safety Data Sheets – Yersinia pestis. Available from: Pathogen Safety Data Sheets: Infectious Substances – Yersinia pestis - Canada.ca
Sentinel Level Clinical Laboratory Guidelines for Suspected Agents of Bioterrorism and Emerging Infectious Disease – Yersinia pestis. American Society for Microbiology (ASM). Revised March 2016. Available from: LRN Sentinel Level Clinical Laboratory Protocols | ASM.org