Releasing Intracellular NS1 from Mosquito Cells for the Detection of Dengue Virus-Infected Mosquitoes.

Lie Cheng,Hsin-Wei Chen,Hsing-Han Li,Jih-Jin Tsai,Matthew P Su,Wei-Liang Liu,Chao-Ying Pan,Chun-Hong Chen,Shih-Cheng Wu

doi:10.3390/v12101105

Abstract

Dengue virus (DENV), the pathogen that causes dengue fever, is mainly transmitted by Aedes aegypti. Surveillance of infected mosquitoes is a major component of integrated mosquito control methods for reducing the risk of vector-born disease outbreaks. However, a specialized rapid test for DENV detection in mosquitoes is not currently available. Utilizing immunoblotting, we found that the secretion of NS1 from both a DENV-infected mosquito cell line and mosquito bodies was below the detection threshold. However, when Triton X-100 was used to lyse infected mosquitoes, intracellular NS1 was released, and could then be effectively detected by the NS1 rapid test. The distribution of DENV NS1 in intrathoracically infected mosquitoes was different from that of orally infected mosquitoes. Next, we performed sensitivity tests by bisecting mosquitoes longitudinally; one half of each mosquito was subjected to the NS1 rapid test while the other half was used for qPCR confirmation. This modified test had a sensitivity of nearly 90% from five days post-infection onwards, while DENV had escaped from the midgut barrier. This adapted test offers a valuable, easy-to-use tool for mosquito surveillance, which is a crucial component of DENV disease control.

Highlights

Dengue virus (DENV) is a flavivirus with four distinct serotypes (DENV 1–4), and is predominantly transmitted by Aedes aegypti (A. aegypti) and Aedes albopictus
If the secreted form of non-structural protein 1 (NS1) infection status using the NS1 rapid test may be possible in a manner similar to that achieved via the or saliva
We found that 0.5% and 1% Triton X-100-phosphate-buffered saline (PBS) allowed intracellular NS1 to be released in sufficient quantities for rapid test detection (Figure 2D)

Summary

Introduction

Dengue virus (DENV) is a flavivirus with four distinct serotypes (DENV 1–4), and is predominantly transmitted by Aedes aegypti (A. aegypti) and Aedes albopictus. While several methods exist for detecting dengue infection in humans, such as virus isolation, ELISA, qPCR and the dengue non-structural protein 1 (NS1) rapid test [9,10,11,12,13], there are currently no readily available and specialized rapid tests for detecting DENV in mosquitoes. Adapting one of these tests for use in mosquitoes could be of great value for mosquito control programs

Methods

Results

Conclusion