Wicking Assays for the Rapid Detection of West Nile and St. Louis Encephalitis Viral Antigens in Mosquitoes (Diptera: Culicidae)

Abstract

The recent outbreaks of West Nile (WN) encephalitis and St. Louis encephalitis (SLE) in the United States have highlighted the need for rapid and specific methods of detecting arboviral antigens in mosquitoes. We evaluated rapid, field-usable assays for detecting and differentiating WN and SLE viruses in mosquito pools, based on a patent-pending, immunochromatographic technology (VecTest) formatted on a dipstick. The device provides results in less than 20 min and can be used in laboratories with adequate containment facilities. In laboratory assessments, both the SLE and WN virus tests demonstrated sensitivity comparable with that of an antigen capture ELISA, but less than can be achieved with Vero cell plaque or reverse-transcriptase polymerase chain reaction assays. There was no evidence of cross-reaction when tested with high concentrations of heterologous flavivirus antigens or with Eastern equine encephalitis or Western equine encephalitis viruses. Both the WN and SLE dipstick tests delivered a clear positive result with a single positive specimen in a pool of 50 mosquitoes. This virus assay technology reduces the time required to obtain test results and will allow rapid medical threat assessment and effective targeting of vector control measures.

Recent progress in West Nile virus diagnosis and vaccination

“…It uses a detection dipstick coated with specific antibodies. Although it is less sensitive than the plaque assay in Vero cells or RT-PCR [111], it has the advantage that it gives a result in less than 20 minutes and it does not require sophisticated equipment.…”

Sources of Error in the Estimation of Mosquito Infection Rates Used to Assess Risk of Arbovirus Transmission

“…The total PFUs in the pool was then divided by 2.5 to generate a PFU/mL concentration for each pool to simulate typical homogenization and extraction methods (using 2.5 mL of diluent as for the VecTest). 13 Two viral assays with different virus detection ability were simulated. The first one was an assay that could only detect virus at concentrations ≥ 10 3.8 PFU/mL.…”

“…These assays differ in their power to discriminate among viruses and in their ability to detect the virus particles or plaque-forming units (PFU) at different concentrations. 13,14 The quantity of virus in a mosquito pool affects the outcome of the different assays because each assay has a threshold concentration below which virus usually cannot be detected. The quantity of virus in mosquitoes is affected by time and F igure 1.…”

West Nile Virus Infection in Farmed American Alligators (Alligator Mississippiensis) in Florida

“…The RT-PCR was performed using the ABI Prism 7000 Sequence Detection System (Applied Biosystems, Foster City, California) or the Bio-Rad Icycler IQ Real-Time Detection System (Bio-Rad, Hercules, California) according to manufacturers' specifications. Selected viral harvests from plaque assays were identified as WNV using a WNV-specific antigen detection assay (Medical Analysis Systems, Inc., Camarillo, California; Ryan et al, 2003).…”