Specimen Submission for Diagnostic Testing

Public health department laboratory staff may be able to forward specimens to CDC for specialized testing, including a range of assays for Mycoplasma pneumoniae. CDC uses molecular testing with multiplex quantitative Polymerase Chain Reaction (qPCR) as the primary laboratory procedure for M. pneumoniae identification. Both culture and serological testing are not used as routine diagnostics in the CDC laboratory. Refer to the M. pneumoniae diagnostic methods for more information.

All necessary information for investigating suspected M. pneumoniae cases and providing specimens to the CDC laboratory for diagnostic testing are highlighted below. Note that certain tests, PCR and culture in particular, will have better yields if specimen collection occurs before starting treatment with antimicrobials.

Approved Specimen Types

CDC’s laboratory accepts any of the following types of specimens for M. pneumoniae testing from public health department laboratories (only if the specimen type does not fall within the category of “specimen rejection criteria” as outlined below):

Combined nasopharyngeal (NP) and oropharyngeal (OP) swabs in viral transfer media (VTM) or universal transfer media (UTM)
NP swabs in VTM/UTM
OP swabs in VTM/UTM
Sputum
Tissue
Bronchial lavage fluid
Bronchial washings
Cerebrospinal fluid
Purified nucleic acids
Bacterial isolates

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Specimen Rejection Criteria

The following are reasons why specimens are most commonly rejected by CDC’s laboratory for M. pneumoniae testing:

Specimens collected using calcium alginate swabs, cotton swabs, or swabs with wooden shafts (use only sterile Dacron, rayon, or nylon swabs with plastic shafts; if available, use flocked swabs)
Specimens not transported at proper temperature (refer to Specimen Submission for shipment requirements)
Specimens with insufficient volume for testing (200µL specimen volume is required for completion of diagnostic testing but additional volume is preferred for further testing such as isolate typing)
Inappropriate specimen types
Specimens arriving with no documentation, insufficient documentation, or erroneous documentation (refer to Specimen Submission for necessary documentation)

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Specimen Collection Protocol

Developed by the Unexplained Respiratory Disease Outbreak (URDO) working group, the Specimen Collection Guidelines [8 pages] detail how to collect tissue specimens, as well as upper (NP, OP) and lower (sputum, bronchial lavage fluid) respiratory tract specimens.

Before collecting a specimen, ask the patient (and patient’s parent, if applicable) for permission to swab him or her for M. pneumoniae and other respiratory pathogen testing if you have a patient who meets at least one of the following criteria:

Cough for ≥ 3 days and fever ≥ 100.0°F (reported or documented)
Chest x-ray confirmed pneumonia
Erythema multiforme, Stevens-Johnson syndrome, or toxic epidermal necrolysis

Collect an OP and NP swab specimen

Use only sterile Dacron, rayon, or nylon swabs with plastic shafts. If available, use flocked swabs. DO NOT use calcium alginate swabs or swabs with wooden sticks, as they may contain substances that inactivate some agents and inhibit some molecular assays.

For OP swab specimens:

Insert swab into the posterior pharynx and tonsillar areas. Rub swab over both tonsillar pillars and posterior oropharynx and avoid touching the tongue, teeth, and gums.

For NP swab specimens:

Insert flexible shaft swab through the nares parallel to the palate (not upwards) until resistance is encountered or the distance is equivalent to that from the ear to the nostril of the patient indicating contact with the nasopharnyx. Gently rub and roll the swab.

Collecting a Nasopharyngeal Swab Clinical Specimen is a pertussis video that demonstrates proper techniques for collecting and transporting a clinical specimen obtained by swabbing the posterior nasopharynx. While it is not specific to M. pneumoniae, the technique is the same.

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