Bacterial DNA Detection by PCR

General Information

Lab Name

Bacterial DNA Detection by PCR

Lab Code

BCTDNA

Epic Name

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.

BRTDNA (Bartonella PCR)
ENBDNA(Enterobacterales DNA Detection)
LEGDNA (Legionella PCR)
MAVDNA (Mycobacterium avium complex PCR)
MPLDNA (Mycoplasma PCR)
TBCDNA (Mycobacterium tuberculosis complex DNA)
TPLDNA (Treponema pallidum DNA)
TWHDNA (Tropheryma whipplei PCR)

MULTIPLE TEMPLATE REFLEXES, if requested by ordering physician:
NGS16S (Bacterial detection by 16S NGS)

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

Code Name
BT16RS Bacterial PCR: Detection, 16S rDNA
BT16ID Bacterial PCR: Identification, 16S rDNA
BTSI Bacterial PCR: Specimen Description
BTSPI Bacterial PCR: External Identifier
BTSR Bacterial PCR: Special Requests
BTSC Bacterial PCR: Specimen Comments
BTNAE Bacterial PCR: Specimen DNA Extraction
BTREV Bacterial PCR: Pathologist Review
BTME Bacterial PCR: Method Note

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)

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
  • No SPS (Sodium Polyanethole Sulfonate) e.g. Wampole Isolator Tubes
  • 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

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