Histoplasma DNA detection by PCR
General Information
Lab Name
Histoplasma DNA detection by PCR
Lab Code
HISDNA
Epic Name
Histoplasma DNA detection by PCR
Description
Detection of Histoplasma capsulatum DNA
Histoplasma capsulatum (or Ajellomyces capsulatum) causes Histoplasmosis which mainly affects lungs. Infected individuals generally exhibit no clinical manifestations or ill effects. Acute phase is manifested by non-specific respiratory symptoms that are cough or flu like. Chronic Histoplasmosis can manifest tuberculosis like symptoms and can also lead to disseminated disease that can be fatal if not treated.
First described in 1906 by Darling (1906), histoplasmosis is an infectious disease caused by inhaling the dimorphic fungi Histoplasma capsulatum microconidia. Although histoplasmosis is an endemic tropic systemic mycosis localized in the Americas and parts of Asia and Africa, several cases do occur in nonendemic areas. This disease mostly affects immunodeficient patients, particularly those with HIV. However, some cases of disseminated histoplasmosis in immunocompetent individuals have been described.
Diagnosis by detection of antigen (EIA) and detection of antibody are widely used. Direct microscopic examination and culture on Sabouraud media are the gold standard method for diagnosis. However, microscopic examination requires a good level of expertise. Moreover, the culture method is very slow (up to 4 months) and is hazardous (due to the presence of spores). Development in molecular biologic and polymerase chain reaction (PCR) techniques has permitted the detection of H. capsulatum DNA.
Histoplasma DNA detection by PCR [HISDNA] utilizes specifically designed primers that can amplify Histoplasma capsulatum specific ITS DNA. These specific primers are used in a nested PCR following amplification of the ITS2 region. More recently the laboratory has designed FRET probes specific for detecting Histoplasma capsulatum and have been able to detect closely related organisms such as Blastomyces species and Emergomyces species.
References
Kauffman et al. (2007) Histoplasmosis: Clinical and Laboratory Update in Clinical Microbiology Reviews p. 115-132
Synonyms
Fungal sequencing, Histoplasma detection, Histoplasma identification, Histoplasma PCR, Histoplasma sequencing, Histoplasmosis PCR, molecular Histoplasma, universal PCR
Components
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 |
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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