Tropheryma whipplei DNA detection by PCR
General Information
Lab Name
Tropheryma whipplei DNA detection by PCR
Lab Code
TWHDNA
Epic Name
Tropheryma whipplei DNA detection by PCR
Description
Detection of Tropheryma whipplei DNA
Whipple's disease is an infectious illness associated with a wide variety of intestinal and extraintestinal manifestations. Since untreated disease leads to death and because adequate antimicrobial therapy for diseases with similar clinical presentations differs significantly in the duration and choice of drugs, a reliable laboratory diagnosis is of considerable interest. The causative agent has been propagated in human macrophages deactivated with interleukin-4, but stable subcultures have been established only very recently. Based on its 16S ribosomal DNA (rDNA) sequence, it was classified as a member of the gram-positive bacteria with high G+C content and named Tropheryma whipplei (formerly Tropheryma whippelii).
Recently, T. whipplei DNA has been found in duodenal biopsy tissue, gastric juice, and saliva specimens, but not in dental stone, of patients without Whipple's disease. Despite recent success in culturing T. whipplei, the clinical suspicion of Whipple's disease is usually confirmed by periodic acid-Schiff staining of inclusions in macrophages or by electron microscopy showing the characteristic trilamellar membrane of T. whipplei.
Molecular tests are significantly more sensitive than histopathology for intestinal and extraintestinal specimens. T. whipplei-specific seminested PCR utilizes a highly conserved segment of the hsp65 gene for the specific detection of T. whipplei.
References
La Scola et al. 51 (4):(2001) Description of Tropheryma whipplei gen. nov., sp. nov., the Whipple’s disease bacillus in International Journal of Systematic and Evolutionary Microbiology, pp1471–1479
Synonyms
16S PCR, molecular Tropheryma whipplei, Tropheryma whipplei identification, Tropheryma whipplei PCR, Tropheryma whipplei sequencing, Twhip PCR, universal PCR, Whipple’s disease, Whipple’s PCR
Components
| Code | Name |
|---|---|
| TWHSUM | T whipplei PCR: Summary |
| TW16RS | T whipplei PCR: Detection, 16S rRNA |
| TW16ID | T whipplei PCR: Identification, 16S rRNA |
| TWHSRS | T whipplei PCR: Detection, hsp65 |
| TWHSID | T whipplei PCR: Identification, hsp65 |
| TWHSD | T whipplei PCR: Specimen Description |
| TWHSPI | T whipplei PCR: External Identifier |
| TWHSR | T whipplei PCR: Special Requests |
| TWHSC | T whipplei PCR: Specimen Comments |
| TWHNAE | T whipplei PCR: Specimen DNA Extraction |
| TWHREV | T whipplei PCR: Pathologist Review |
| TWHME | T whipplei 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
Collection
Acceptable specimens are listed below. Please see Molecular Microbiology Specimen Submission for complete specimen collection and handling instructions.
Shipping/Handling
- Fresh frozen tissue/fluid specimens should be collected into a DNA free container labeled with at least two identifiers and 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.
Acceptable Specimens
- Fresh frozen tissue
- Fresh frozen fluid: any body fluid is acceptable if it is not listed under Unacceptable Specimens.
- Common examples include: cerebrospinal fluid, pleural fluid, pericardial fluid, urine, bronchial lavage, joint fluid, bone marrow, vitreous fluid, etc.
- Formalin Fixed Paraffin-embedded tissues (FFPE, PET): blocks, scrolls, and unstained slides. Note that fresh tissue is considered the optimal specimen of choice, as it reduces the chance of introducing exogenous DNA templates or microorganisms during embedding/fixation, and because formalin fixation dramatically reduces the sensitivity of molecular assays due to reduced template yield and quality.
- Sputum: acceptable - except for Bacterial PCR reflex NGS [BCTDNA]/Bacterial DNA Detection by PCR (without reflex to NGS) [NRBDNA], Fungal PCR reflex NGS [FUNDNA]/Fungal DNA Detection by PCR (without reflex to NGS) [NRFDNA], and Nontuberculous Mycobacteria DNA Detection [NTMDNA]* assays
- eSwabs**, UTM (universal transport media), body fluid/bone marrow in EDTA (not including blood)
- Sodium polyanethol sulfonate (SPS, Wampole Isolator Tubes) acceptable with disclaimer
*Mycobacterium avium complex DNA Detection [MAVDNA] can be ordered on sputum
**Fungal PCR reflex NGS [FUNDNA] and Fungal DNA Detection by PCR (without reflex to NGS) [NRFDNA] may have interference due to some lots of eSwabs which have been found to contain Saccharomyces cerevisiae DNA, resulting in false positive detection. Clinical correlation and/or retesting with a different collection method is advised. The detection of S. cerevisiae from eSwab specimens can interfere with our ability to rule out other fungal DNA.
Unacceptable Specimens
- Blood, serum, plasma, stool/rectal swabs
- No citrated or heparinized solutions
- Tissues floating in formalin
- Swab/fluid collected in tube containing agar
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 over scrolls/unstained slides 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.
Fresh tissue is the optimal specimen of choice, as it reduces the chance of introducing exogenous DNA templates or microorganisms during embedding/fixation. Formalin fixation dramatically reduces the sensitivity of the assays due to reduced template yield and quality.
Forms & Requisitions
Molecular Microbiology Order Form
Handling Instructions
Please see Molecular Microbiology Specimen Submission for complete specimen collection and handling instructions.
Quantity
requested: See "Collection" for Optimal Quantity
minimum: Specimens below optimal quantity are acceptable for testing, however, diagnostic yield is generally proportional to specimen size.
Processing
UWMC/HMC: Store and send fresh tissue/fluid specimens refrigerated, if specimen storage and transport will exceed 8 hours, freeze at -20°C. Freeze all fresh tissue/fluid specimens at -20°C upon arrival in UW Molecular Microbiology.
Performance
LIS Dept Code
Micro Molecular Diag (MMD)
Performing Location(s)
| UW-MT |
Microbiology, Molecular Diagnostics
206-520-4600 ---------------------------------------- Shipping Address Attn: Molecular Microbiology Performing Lab Address Clinical Microbiology Lab, NW177 |
Contact Information Please e-mail us with any questions or comments you may have. Your inquiry will be answered as soon as possible. email: molmicdx@uw.edu The Molecular Microbiology lab is open from Monday-Friday, 7am-4pm PDT. Billing inquiries and requests for faxed reports can be made to our Client Services Department at (206) 520-4600 or (800) 713-5198. For results or other inquiries, we can be reached by phone at the following numbers:
For assistance during weekends, holidays and after hours, please contact Lab Medicine Resident at (206) 598-6190 |
|---|
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