Bacterial cultures – Anaerobic – Reference Identification/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.

This page provides Anaerobic Identification/Confirmation testing information for bacterial cultures at Public Health Ontario (PHO). For information regarding Aerobic Identification/Confirmation testing information for Bacterial cultures, refer to the following PHO webpages:

See – Algorithm: Clostridium species isolates from sites OTHER than STOOL, GASTRIC ASPIRATE, or WOUND that are requested for “botulinum toxin testing” or “botulism testing” are NO LONGER tested by the Botulism Reference Service Centre of Canada; these isolates will continue to be identified at PHO’s laboratory. 

Testing Indications

Acceptance/Rejection Criteria

  • Primary specimens are not acceptable. Only pure cultures are accepted.
  • If the source of isolation of the culture is not indicated in the requisition, it will be rejected.
  • Mixed or non-viable cultures will not be tested. A written report will be issued to indicate that the test has been rejected.
  • Mislabeled or unlabeled culture will not be tested. A report will be sent to the submitting laboratory stating “Culture received unlabeled” or “Requisition identification does not match the identification information on the culture received. Please resubmit”.
  • The submitting laboratory will be contacted regarding isolates or specimens from critical sites (e.g. brain abscess) that are submitted inappropriately (e.g. improper transport conditions or mislabeled/unlabeled).
  • Isolates submitted that are not in appropriate anaerobic transport system at room temperature will not be tested.
  • Do not submit multiple isolates from the same specimen/site of infection - for polymicrobic infections, only submit the two most predominant isolates or clinically significant pathogens. Laboratories should perform preliminary identification tests including catalase (15% hydrogen peroxide) and Gram stain.  Note growth characteristics on all isolates on the Reference Bacteriology Requisition .

Specimen Collection and Handling

Specimen Requirements

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

Bacterial Cultures-Anaerobic-Reference ID/Confirmation

Pure viable subculture of 24 – 48 hours growth of organism on appropriate pre-reduced medium submitted in an anaerobic transport kit or anaerobic swab2,3



Submission and Collection Notes


Please be sure to complete all fields on the Reference Bacteriology Requisition including:

  1. Test requested
  2. Date of primary specimen collection
  3. Source of isolation (very important):
    1. Type of urine specimen(s) (e.g. midstream, indwelling catheter, cystoscopy etc.)
    2. The site and type of wound swab; be as specific as possible (e.g. animal bite, surgical wound, decubitus ulcer, etc.)
    3. The type of specimen if other than blood culture, urine or wound swab, please provide as much detail as possible
    4. The number of consecutive blood cultures positive for the submitted isolate when isolates are from blood cultures 
  4. Any previous antibiotic treatment 
  5. Gram stain description, aero tolerance test results and catalase reaction
  6. Presumptive identification
  7. Indicate if isolate is part of a polymicrobic infection 

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.


If the healthcare provider requires susceptibility testing, the submitter must call within 5 days of receiving final results and provide necessary information and justification for susceptibility testing request.


Do not submit multiple isolates from the same specimen/site of infection - for polymicrobic infections, only submit most clinically significant pathogen(s) for testing. 


Primary specimens are unacceptable as they should be processed in the originating lab. A report will be issued to indicate that the test has been cancelled.


Where full bacterial identification and susceptibility testing is clinically indicated, such as for an immunocompromised patient, please clearly indicate this on the requisition.


Laboratories should perform preliminary identification tests including catalase (15% hydrogen peroxide) and Gram stain. Note growth characteristics of the isolate being submitted on the requisition form.

Storage and Transport

Place the  culture in a biohazard bag and seal. Culture should be transported at room temperature to the laboratory within 48 hours after isolation or propagation through anaerobic incubation. All cultures must be shipped in accordance to the Transportation of Dangerous Good Act.

Requisitions and Kit Ordering

Test Frequency and Turnaround Time (TAT)

Anaerobic cultures are tested Monday – Friday at PHO’s laboratory-Toronto.

Turnaround time is 6-8 days from date of receipt at PHO’s laboratory.

Test Methods

All bacterial culture isolates are identified/confirmed by conventional identification methods:

  • Gram stain morphologic examination
  • Biochemical testing
  • Matrix-Assisted Laser Desorption/Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS)

If the conventional identification methods are not able to provide definitive identification of the isolate being tested, 16S rRNA gene PCR and sequence analysis will be performed on the isolate and identified to the most specific level possible.


  • Species level identification will be attempted for Lactobacillus spp. isolated from systemic infections only (e.g. blood, pleural fluid etc.).  Lactobacillus spp.from urine cultures will not be processed6.
  • Identification will be performed on organisms commonly considered to be contaminants/commensal flora only on special request by the healthcare provider based on clinical relevance. Contact PHO’s laboratory  Customer Service for these requests. e.g. Propionibacterium spp.and Cutibacterium spp. from single Positive Blood Culture,1, 2-5 Wound swab and Drainage2.

Special Note on Potential Botulinum Toxin producing Clostridium species:

  • Potential Botulinum Toxin producing Clostridium species (includes C. botulinum, C. sporogenes, C. barrati, C. butyricum and C. argentinense) from sites OTHER than STOOL, GASTRIC ASPIRATE, or WOUND, if isolate received with requests “Clostridium botulinum toxin” or “Botulism testing” or similar on the requisition, the toxin testing request will be cancelled and followed up with report to the client with the following note:

Clostridium botulinum Toxin testing is not done on this isolate.

If botulism is suspected, submit clinical specimens (such as blood, stool, vomit, or wound swabs) or suspect food samples from symptomatic patients directly to the Botulism Reference Service (BRS) in Ottawa, Canada, for toxin testing. Evidence of the toxin may help confirm a diagnosis of infant, foodborne or wound botulism.

Please refer to the following guides:


The Laboratory Test Report will show the identification of the isolate : “ Isolate identified as [Genus species]”

Results are reported to the ordering physician, authorized health care provider (General O. Reg 45/22, s.18) or submitter as indicated on the requisition.


  1. Institute for Quality Management in Healthcare (IQMH): Consensus Practice Recommendations – BACT-Blood Cultures. Revision date: 2017-09-28.
  2. Institute for Quality Management in Healthcare (IQMH): Consensus Practice Recommendations – BACT Superficial wound swabs. 2013-12-09.
  3. Miller JM, Binnicker MJ, Campbell S, Carroll KC, et al. A Guide to Utilization of the Microbiology Laboratory for Diagnosis of Infectious Diseases: 2018 Update by the Infectious Diseases Society of America and the American Society for Microbiology. Clinical Infectious Diseases, Volume 67, Issue 6, 31 August 2018, Pages e1–e94,
  4. Clinical and Laboratory Standards Institute (CLSI). Principles and procedures for blood culture; approved guideline. CLSI document M47-A, Vol. 27, No. 17. Wayne, Pennsylvania, USA. 2007.
  5. Baron EJ, Weinstein MP, Dunne Jr WM, Yagupsky P, Welch DF, Wilson DM. 2005. Cumitech 1C, Blood Cultures IV. Coordinating ed., EJ Baron. ASM Press, Washington DC.
  6. McCarter YS, Burd EM, Hall, GS, Zervos M. 2009. Cumitech 2C, Laboratory Diagnosis of Urinary Tract Infections. Coordinating ed. SE Sharp. ASM Press, Washington, DC.
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Updated 18 April 2024