What is the PCR (Polymerase Chain Reaction) HTLV (Human T-lymphotropic virus) 1-2 test?

PCR HTLV 1-2 Test: A Diagnostic Tool for HTLV Infection

The PCR HTLV 1-2 test is a molecular diagnostic method that detects and quantifies proviral DNA of Human T-lymphotropic virus types 1 and 2, allowing for differentiation between these two virus types when serological tests are inconclusive or indeterminate. 1

Test Description and Purpose

The PCR (Polymerase Chain Reaction) HTLV 1-2 test serves several important clinical purposes:

• Virus Detection and Differentiation: It can detect and distinguish between HTLV-1 and HTLV-2 infections, which is crucial as these viruses have different clinical implications 2
• Confirmation of Infection: It serves as a confirmatory test when serological tests yield indeterminate or inconclusive results 1
• Proviral Load Quantification: It measures the proviral load (PVL), which is the proportion of infected lymphocytes, an important marker for disease progression risk 3

Testing Methodology

The PCR HTLV 1-2 test employs several techniques:

• Standard PCR: Uses type-specific primers targeting the tax gene region to detect and differentiate HTLV-1 from HTLV-2 2
• Real-time PCR: Provides quantitative measurement of proviral load with high sensitivity 4
• Droplet Digital PCR (ddPCR): A newer technique offering absolute quantification with very low variability (coefficient of variation 0.97-8.2%) 3
• Multiplex Real-time PCR: Allows simultaneous detection, genotyping, and quantification of HTLV-1, HTLV-2, and even HTLV-3 in a single reaction 5

Clinical Applications

The test has several important clinical applications:

• Resolving Indeterminate Western Blot Results: Up to 50% of samples with indeterminate Western blot results in high-risk populations may be positive for HTLV by PCR 6
• Disease Risk Assessment: Higher proviral loads are associated with increased risk of developing HTLV-associated diseases, particularly HAM/TSP (HTLV-1 Associated Myelopathy/Tropical Spastic Paraparesis) 4
• Monitoring Disease Progression: Regular monitoring of proviral load helps in assessing disease activity and progression 1
• Differentiating HTLV Types: PCR can reliably distinguish between HTLV-1 and HTLV-2 infections, which is important as HTLV-1 is associated with more severe diseases 2

Diagnostic Algorithm

The CDC and recent guidelines recommend a two-step testing approach for HTLV diagnosis 1:

1. Initial Screening: Enzyme immunoassay (EIA) using HTLV-1 whole-virus lysate antigens
2. Confirmatory Testing:
   • Western Blot (WB) or Radioimmunoprecipitation Assay (RIPA) for repeatably reactive specimens
   • PCR testing when Western Blot results are indeterminate or to differentiate between HTLV-1 and HTLV-2

For cost-effectiveness and improved accuracy, some experts propose an alternative algorithm 7:

1. Two EIAs for screening
2. Real-time PCR for confirmation
3. Western Blot only for PCR-negative samples that were EIA-positive

Clinical Significance

The ability to detect and quantify HTLV proviral load has important implications:

• Patients with HAM/TSP typically have significantly higher proviral loads (average 13,450 copies/10^6 cells) compared to asymptomatic carriers (average 6,665 copies/10^6 cells) 4
• The test helps identify individuals who may need closer monitoring for disease development
• It assists in counseling infected individuals about transmission prevention measures 1

Limitations and Considerations

• PCR testing requires specialized laboratory equipment and expertise
• False negatives can occur with very low proviral loads
• The test should be interpreted in conjunction with clinical findings and other laboratory results
• Different PCR methodologies may have varying sensitivities and specificities

The PCR HTLV 1-2 test represents a significant advancement in HTLV diagnostics, providing more definitive results than serological testing alone, particularly in cases with indeterminate Western blot results.