Haemophilus influenzae – Confirmation of Identification and Serotyping

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This page provides identification, confirmation of identification and serotyping information for Haemophilus influenzae at Public Health Ontario (PHO). Haemophilus influenzae is the causative agent of various infections, ranging from serious conditions such as meningitis, bloodstream infections and pneumonia to milder infections such as ear infection and bronchitis.

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

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Testing Indications

Submit the bacterial isolate for confirmation if Haemophilus influenzae is strongly suspected but is displaying unusual phenotypic characteristics that warrant further verification.

Note: Serotyping is performed on isolates recovered from sterile sites only.

Acceptance/Rejection Criteria

Primary specimens are not acceptable; only pure viable 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.
Mis-labelled or un-labelled culture will not be tested.

Specimen Collection and Handling

Specimen Requirements

Test Requested	Required Requisition(s)	Specimen Type	Minimum Volume	Collection Kit
Haemophilus influenzae – Confirmation of ID and Serotyping	Requisition for Specimen Containers and Supplies	Pure viable subculture of organism on appropriate media that supports the growth	N/A	Agar plate or slant tube
Haemophilus influenzae – Confirmation of ID and Serotyping	Reference Bacteriology Requisition	Swab of the pure subculture	N/A	Amies charcoal transport medium

Submission and Collection Notes

Complete all fields of the requisition form, including the following mandatory fields:

Test(s) Requested: Select confirmation and identification
Culture Information: Indicate the presumptive identification and Gram morphology; provide oxygen requirement for growth; oxidase test result; catalase test result; indicate of presence or absence of growth on MacConkey agar
Clinical Information: Indicate if patient is immunocompromised
Date of Collection of primary specimens from which isolate was recovered
Source of isolate or the primary specimen from which isolate was recovered

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 bacterial culture submitted for identification confirmation is provided on a swab in Amies charcoal transport media or broth media (instead of on a plate) the turnaround time will be delayed by at least 24 hours.

If susceptibility testing is required, the submitter must call within 5 days of receiving the final identification confirmation result and provide necessary information and justification for susceptibility testing request.

Do not submit multiple isolates from the same specimen/site of infection.

Primary specimens are unacceptable as they should be processed in the originating lab.

If full bacterial identification and susceptibility testing is clinically indicated- such as in cases involving immunocompromised patients, it must be clearly indicated on the requisition.

Limitations

Culture must be viable. Use a fresh subculture and ship in conditions to ensure viability on receipt.

Storage and Transport

Place the sealed culture in a biohazard bag and properly seal. Store at 2-8°C while awaiting shipment. The requisition form must be placed in the outer pouch of the biohazard bag, separate from the culture container. Transport a fresh subculture to ensure viability on receipt. It should be shipped to PHO within 48 hours of isolation. All cultures must be shipped in accordance to the Transportation of Dangerous Good Act.

Special Instructions

Requisitions and Kit Ordering

Test Frequency and Turnaround Time (TAT)

For Haemophilus influenzae cultures, daily testing (Monday to Saturday) is conducted at PHO, Toronto location.

Serotyping result is available within 24 hours of receipt in the laboratory, exclude Sunday.

Turnaround time is up to four business days for identification confirmation and biotyping.

Test Methods

Confirmation of identification is based on correlated results from the following methods:

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

Serotyping is performed on isolates recovered from sterile sites only; while biotyping is performed on islates recovered from all sites.

Serotyping is performed using the slide agglutination method. Biotyping is performed using biochemical tests.

Interpretation

The following table provides possible test results:

Test	Reported as:
Identification	[Genus species]
Serotyping for Haemophilus influenzae	a or b or c or d or e or f or non-typeable
Biotyping for Haemophilus influenzae	I or II or III or IV or V or VI or VII or VIII

Reporting

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.

Specimens that are positive for Haemophilus influenzae from sterile sites of infection are to be reported to the Medical Officer of Health as per Health Protection and Promotion Act.