Development and evaluation of a visible reverse transcription-loop-mediated isothermal amplification (RT-LAMP) for the detection of Asian lineage ZIKV in field-caught mosquitoes

Padet Siriyasatien; Supaporn Wacharapluesadee; Kanyarat Kraivichian; Charuai Suwanbamrung; Nataya Sutthanont; Ana Cantos-Barreda; Atchara Phumee

doi:10.1016/j.actatropica.2022.106691

Development and evaluation of a visible reverse transcription-loop-mediated isothermal amplification (RT-LAMP) for the detection of Asian lineage ZIKV in field-caught mosquitoes

Acta Trop. 2022 Dec:236:106691. doi: 10.1016/j.actatropica.2022.106691. Epub 2022 Sep 11.

Authors

Padet Siriyasatien¹, Supaporn Wacharapluesadee², Kanyarat Kraivichian³, Charuai Suwanbamrung⁴, Nataya Sutthanont⁵, Ana Cantos-Barreda⁶, Atchara Phumee⁷

Affiliations

¹ Center of Excellence in Vector Biology and Vector Borne Diseases, Department of Parasitology, Faculty of Medicine, Chulalongkorn University, Bangkok 10330, Thailand; Department of Parasitology, Faculty of Medicine, Chulalongkorn University, Bangkok 10330, Thailand.
² Thai Red Cross Emerging Infectious Diseases Clinical Centre, King Chulalongkorn Memorial Hospital, Faculty of Medicine, Chulalongkorn University, Bangkok 10330, Thailand.
³ Department of Parasitology, Faculty of Medicine, Chulalongkorn University, Bangkok 10330, Thailand.
⁴ School of Public Health, Walailak University, Nakhon Si Thammarat, Thailand; Excellent Center for Dengue and Community Public Health (EC for DACH), Walailak University, Nakhon Si Thammarat 80160, Thailand.
⁵ Department of Medical Entomology, Faculty of Tropical Medicine, Mahidol University Bangkok 10400, Thailand.
⁶ Department of Biochemistry and Molecular Biology-A, Faculty of Veterinary Medicine, Regional Campus of International Excellence "Campus Mare Nostrum", University of Murcia, Espinardo, Murcia 30100, Spain.
⁷ Excellent Center for Dengue and Community Public Health (EC for DACH), Walailak University, Nakhon Si Thammarat 80160, Thailand; Department of Medical Technology, School of Allied Health Sciences, Walailak University, Nakhon Si Thammarat 80160, Thailand. Electronic address: atchara.ph@wu.ac.th.

PMID: 36103950
DOI: 10.1016/j.actatropica.2022.106691

Abstract

The Zika virus (ZIKV) infection is an emerging and re-emerging arbovirus infection that is transmitted to humans through the bite of infected mosquitoes. Early detection of ZIKV in mosquitoes is one of the prerequisite approaches for tracking the spread of the virus. Therefore, this study aims to develop and validate a visual reverse transcription-loop-mediated isothermal amplification (RT-LAMP) method called ZIKV-RT-LAMP, for detecting ZIKV in field collected mosquito samples from Thailand. A single-tube ZIKV-RT-LAMP assay was developed to detect Asian lineage ZIKV RNA. The detection limit and cross-reactivity of ZIKV were investigated. The hemi-nested RT-PCR (hn-RT-PCR) and the colorimetric LAMP kit (cLAMP kit) were performed as reference assays. The detection limit of the ZIKV-RT-LAMP assay was 10^-6 ffu/ml or pfu/ml, making it highly specific and 100 times more sensitive than the hn-RT-PCR and cLAMP kits. The ZIKV-RT-LAMP assay detected the Asian lineage of ZIKV RNA without cross-reactivity with other arthropod-borne viruses. The sensitivity and specificity of the ZIKV-RT-LAMP assay were 92.31% and 100%, respectively. The ZIKV-RT-LAMP is a simple, rapid, and inexpensive method for detecting ZIKV in field-caught mosquitos. In the future, extensive surveys of field-caught mosquito populations should be conducted. Early detection of ZIKV in field-caught mosquitoes provides for prompt and effective implementation of mosquito control strategies in endemic areas.

Keywords: Mosquitoes; RT-LAMP; Thailand; Zika virus.

MeSH terms

Animals
Culicidae* / genetics
Humans
Molecular Diagnostic Techniques
Nucleic Acid Amplification Techniques / methods
RNA
RNA, Viral / genetics
Sensitivity and Specificity
Zika Virus Infection* / diagnosis
Zika Virus* / genetics

Substances

RNA, Viral
RNA

Supplementary concepts

LAMP assay