Bacterial DNA Detection by PCR

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General Information

Lab Name
Bacterial DNA Detection by PCR
Lab Code
BCTDNA
Epic Ordering
Bacterial DNA detection by PCR
Description

Detection and identification of Bacterial DNA (16S rRNA target).

Accurate identification of bacterial isolates is an essential task of the clinical microbiology laboratory that enables initiation of proper antimicrobial therapy. For many organisms, traditional phenotypic identification may be difficult, laborious and time-consuming. This issue is further confounded by phenotypic variation within species, many newly described pathogenic species and the limited battery of phenotypic tests available to distinguish among established and potential bacterial pathogens. It is sometimes essential to identify bacterial isolates to species level in order to rule out species that are drug resistant or more pathogenic, to detect unsuspected pathogens, ascribe pathogenicity to species so far considered to be nonpathogenic and to identify new bacterial species.

With more than 20,000 sequences available in public databases (e.g. NCBI), 16S ribosomal RNA gene (rRNA) sequencing is considered by current taxonomists to be the gold standard in bacterial identification and classification. 16S rRNA gene contains conserved regions useful for the design of broad-range PCR primers that can amplify various fragments of the 16S rRNA gene from pathogenic and nonpathogenic bacteria. These fragments include hypervariable regions containing species-specific signature sequences useful for bacterial identification to species level.

In many specimens bacteria can be seen by microscopy of tissue sections but are very difficult to grow due to their fastidious nature, or are not viable as a result of antimicrobial therapy. Some specimens may never reveal the presence of a bacterial pathogen because of low abundance and/or lack of viability. The use of PCR to detect this DNA extracted directly from clinical specimens facilitates the identification of these pathogens.

REFLEXIVE TESTING: When suspected pathogen microorganisms are detected, identification procedures are performed, as appropriate for the organism and specimen.

Bartonella DNA detection by PCR [BRTDNA]

Enterobacterales DNA Detection [ENBDNA]

Legionella DNA detection by PCR [LEGDNA]

Mycobacterium avium complex DNA Detection [MAVDNA]

Mycoplasma and Ureaplasma DNA Detection by PCR [MPLDNA]

M. tuberculosis complex DNA Detection [TBCDNA]

Treponema pallidum DNA detection by NAAT [TPLDNA]

Tropheryma whipplei DNA detection by PCR [TWHDNA]

MULTIPLE TEMPLATE REFLEXES, if requested by ordering physician:

For more information on this test, to download a requisition form, and for a list of acceptable and unacceptable specimens, please refer to: Molecular Diagnosis Microbiology Section Website

Synonyms
16S PCR, 16S sequencing, bacteria 16S, bacteria identification, bacteria PCR, bacteria sequencing, bacterial 16S, bacterial identification, bacterial PCR, bacterial sequencing, broad range bacteria PCR, broad range bacterial PCR, molecular bacteria, molecular bacterial
Components

Interpretation

Method

DNA extraction, nucleic acid purification, polymerase chain reaction (PCR), sequencing

Reference Range
See individual components
Guidelines

Ordering & Collection

Specimen Type
Tissue (Fresh frozen or paraffin-embedded), Fluid (see Acceptable Specimens for details)
Collection

Acceptable specimens are listed below. Please see this page for complete specimen collection and handling instructions.

Specimens should be collected into a DNA free container labeled with at least two identifiers.

Acceptable Specimens

  • Fresh frozen tissue
  • Fresh frozen fluid: any body fluid is acceptable if it is not listed under Unacceptable Specimens. See common examples below.
  • Formalin Fixed Paraffin-embedded tissues (FFPE, PET): blocks, scrolls, and unstained slides are acceptable
  • eSwabs and UTM (universal transport media)
  • Sodium polyanethol sulfonate (SPS, Wampole Isolator Tubes) acceptable with disclaimer

Common acceptable body fluid examples: cerebrospinal fluid, pleural fluid, pericardial fluid, urine, bronchial lavage, joint fluid, bone marrow, vitreous fluid, etc.

Unacceptable Specimens

  • Sputum, blood, serum, plasma, stool
  • No citrated or heparinized solutions
  • Tissues floating in formalin
  • Swab/fluid collected in tube containing agar
Handling Instructions

Fresh frozen tissue/fluid specimens should be submitted and maintained on dry ice.

Formalin Fixed Paraffin-embedded tissues (FFPE, PET) can be sent ambient or with ice packs during warmer summer months to prevent melting.


Optimal Quantity:

  • Fresh Tissue: 0.3-1.0 cm^3
  • Fluid: 0.2-1 mL
  • Formalin Fixed Paraffin-embedded Tissue (FFPE/PET): blocks are preferred and will be sent back to client upon completion of testing
  • Scrolls/unstained slides: cross-sectional area >1cm^2 send 10 sections of 10µm thickness, if <1cm^2 send 20 sections if available

Please note: We do not need a separate specimen aliquot for each test ordered. Only a single specimen aliquot or block of optimal quantity is necessary for performing multiple tests. If multiple aliquots or blocks of optimal quantity are sent, up to 2 will be pooled.

Quantity
requested: See Optimal Quantity above
minimum: Specimens below optimal quantity are acceptable for testing, however, diagnostic yield is generally proportional to specimen size.

Processing

Processing

UWMC/HMC – store and send specimen refrigerated and freeze specimens at -20°C upon arrival in UW-ML Microbiology. If specimen storage and transport will exceed 8 hours, freeze at -20°C.

Freeze specimens at -20°C upon arrival

Outside Laboratories: Freeze specimens in sterile containers at -20°C. Transport all specimens on ice.

Performance

LIS Dept Code
Micro Molecular Diag (MMD)
Performing Location(s)
UW-MT Microbiology, Molecular Diagnostics
206-520-4600

Clinical Lab, Room # NW177,
University of Washington Medical Center,
1959 NE Pacific street, Seattle, WA 98195

Frequency
Fresh frozen tissues/fluids result in 2-3 business days after receipt of specimen. Formalin Fixed Paraffin-embedded tissues result in 3-4 business days after receipt of specimen.
Available STAT?
No

Billing & Coding

CPT codes
87801
LOINC
76575-0
Interfaced Order Code
UOW4294