Abstract
The rapid spread of Zika virus (ZIKV) represents a global public health problem, especially in areas that harbor several mosquito species responsible for virus transmission, such as Brazil. In these areas, improvement in mosquito control needs to be a top priority, but mosquito viral surveillance occurs inefficiently in ZIKV-endemic countries. Quantitative reverse transcription PCR (qRT-PCR) is the gold standard for molecular diagnostic of ZIKV in both human and mosquito samples. However, the technique presents high cost and limitations for Point-of-care (POC) diagnostics, which hampers its application for a large number of samples in entomological surveillance programs. Here, we developed and validated a one-step reverse transcription LAMP (RT-LAMP) platform for detection of ZIKV in mosquito samples. The RT-LAMP assay was highly specific for ZIKV and up to 10,000 times more sensitive than qRT-PCR. Assay validation was performed using 60 samples from Aedes aegypti and Culex quinquefasciatus mosquitoes collected in Pernambuco State, Brazil, which is at the epicenter of the Zika epidemic. The RT-LAMP had a sensitivity of 100%, specificity of 91.18%, and overall accuracy of 95.24%. Thus, our POC diagnostics is a powerful and inexpensive tool to monitor ZIKV in mosquito populations and will allow developing countries to establish better control strategies for this devastating pathogen.
Highlights
ZIKV is an arbovirus member of the genus Flavivirus in the family Flaviviridae
We determined the ability of reverse transcription LAMP (RT-LAMP) to detect ZIKV in A. aegypti under controlled conditions
Sequencing results and BLAST analysis demonstrated that ZIKV reverse transcriptase (RT)-LAMP amplicons match 100% with virus circulating in Brazil (Fig. 6), confirming the specificity of the RT-LAMP for ZIKV. These results indicated that our ZIKV RT-LAMP assay represents a robust and affordable diagnostic platform that can be used as a surveillance tool for mosquitoes infected with ZIKV
Summary
ZIKV is an arbovirus member of the genus Flavivirus in the family Flaviviridae. The ZIKV genome consists of a single positive-sense single-stranded RNA (+ssRNA), with approximately 11 Kb in length. ZIKV surveillance in insect vectors is an important tool for identifying viral circulation and potential entry points, contributing to prevent outbreaks of disease[17]. This virus has spread rapidly in developing countries that lacks good sanitation infrastructure and harbors several mosquito species competent for ZIKV transmission. QRT-PCR is expensive, requires highly specialized manpower, and involves costly and sophisticated equipment for amplification and detection of the viral genome These drawbacks make the technique unsuitable for large-scale applications in low-resource settings areas, which negatively impact the establishment of effective disease control programs[23,24]. Many of the developed ZIKV LAMP assays still require special equipments for virus detection, which limits its applicability in low-resource scenarios
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