REAL-TIME REVERSE TRANSCRIPTASE–POLYMERASE CHAIN REACTION QUANTIFICATION OF WEST NILE VIRUS TRANSMITTED BY CULEX PIPIENS QUINQUEFASCIATUS

DANA L. VANLANDINGHAM Department of Pathology, University of Texas Medical Branch, Galveston, Texas; Department of Microbiology and Immunology, Tulane University, New Orleans, Louisiana

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BRADLEY S. SCHNEIDER Department of Pathology, University of Texas Medical Branch, Galveston, Texas; Department of Microbiology and Immunology, Tulane University, New Orleans, Louisiana

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KIMBERLY KLINGLER Department of Pathology, University of Texas Medical Branch, Galveston, Texas; Department of Microbiology and Immunology, Tulane University, New Orleans, Louisiana

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JOSEPH FAIR Department of Pathology, University of Texas Medical Branch, Galveston, Texas; Department of Microbiology and Immunology, Tulane University, New Orleans, Louisiana

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DAVID BEASLEY Department of Pathology, University of Texas Medical Branch, Galveston, Texas; Department of Microbiology and Immunology, Tulane University, New Orleans, Louisiana

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JING HUANG Department of Pathology, University of Texas Medical Branch, Galveston, Texas; Department of Microbiology and Immunology, Tulane University, New Orleans, Louisiana

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PATRICIA HAMILTON Department of Pathology, University of Texas Medical Branch, Galveston, Texas; Department of Microbiology and Immunology, Tulane University, New Orleans, Louisiana

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STEPHEN HIGGS Department of Pathology, University of Texas Medical Branch, Galveston, Texas; Department of Microbiology and Immunology, Tulane University, New Orleans, Louisiana

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Transmission experiments are a critical component of vector competence studies. In this study, a real-time reverse transcriptase-polymerase chain reaction (RT-PCR) was used to enumerate the amount of West Nile virus (WNV) secreted in mosquito saliva following oral infection. Culex pipiens quinquefasciatus were allowed to feed on WNV-infected blood, and saliva was collected on days 14 and 21 post-infection (pi). The amount of virus at these two time points varied significantly, with mean equivalent plaque-forming units (pfu) of approximately 30,500 on day 14 pi and 5,800 on day 21 pi. Individual mosquitoes secreted up to 2 × 105 pfu of virus. Titer of whole mosquitoes and immunofluorescence assay of salivary glands from mosquitoes collected at these two time points were also used for supplemental comparison. This report describes the first use of a real-time RT-PCR to quantify the amount of WNV in mosquito saliva.

Author Notes

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