A reverse transcription loop-mediated isothermal amplification for broad coverage detection of Asian and African Zika virus lineages

Boon-Teong Teoh,Kim-Ling Chin,Chee-Sieng Khor,Keivan Zandi,Juraina Abd-Jamil,Annelies Wilder-Smith,Shih-Keng Loong,Nurhafiza Zainal,Sing-Sin Sam,Kim-Kee Tan,Nur-Izyan Samsudin,Sazaly Abubakar

doi:10.1186/s12879-020-05585-4

Abstract

BackgroundEarly detection of Zika virus (ZIKV) infection during the viremia and viruria facilitates proper patient management and mosquito control measurement to prevent disease spread. Therefore, a cost-effective nucleic acid detection method for the diagnosis of ZIKV infection, especially in resource-deficient settings, is highly required.MethodsIn the present study, a single-tube reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the detection of both the Asian and African-lineage ZIKV. The detection limit, strain coverage and cross-reactivity of the ZIKV RT-LAMP assay was evaluated. The sensitivity and specificity of the RT-LAMP were also evaluated using a total of 24 simulated clinical samples. The ZIKV quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assay was used as the reference assay.ResultsThe detection limit of the RT-LAMP assay was 3.73 ZIKV RNA copies (probit analysis, P ≤ 0.05). The RT-LAMP assay detected the ZIKV genomes of both the Asian and African lineages without cross-reacting with other arthropod-borne viruses. The sensitivity and specificity of the RT-LAMP assay were 90% (95% CI = 59.6–98.2) and 100% (95% CI = 78.5–100.0), respectively. The RT-LAMP assay detected ZIKV genome in 9 of 24 (37.5%) of the simulated clinical samples compared to 10 of 24 (41.7%) by qRT-PCR assay with a high level of concordance (κ = 0.913, P < 0.001).ConclusionThe RT-LAMP assay is applicable for the broad coverage detection of both the Asian and African ZIKV strains in resource-deficient settings.

Highlights

Detection of Zika virus (ZIKV) infection during the viremia and viruria facilitates proper patient management and mosquito control measurement to prevent disease spread
We described an improved Reverse transcription loop-mediated isothermal amplification (RT-loop-mediated isothermal amplification (LAMP)) assay which took into account the detection of ZIKV strains from both the African and Asian lineages
The sequences of the RT-LAMP primers were further compared with an alignment of 463 ZIKV genomes retrieved from GenBank (Additional file 4: Figure S2)

Summary

Introduction

Detection of Zika virus (ZIKV) infection during the viremia and viruria facilitates proper patient management and mosquito control measurement to prevent disease spread. The Aedes mosquito-borne Zika virus (ZIKV) was discovered in Uganda in 1947 [1] and until recently, caused only sporadic human infections in Africa and Asia characterized by mild and self-limiting disease [2,3,4]. In 2013–2014, ZIKV rapidly spread to other countries in the Pacific, including French Polynesia [6], New Caledonia [7], the Cook Islands, Solomon Islands and Easter Island [8, 9]. It was during the Brazil outbreak that health authorities observed a surge in adverse pregnancy outcomes including congenital microcephaly, cerebral calcification and fetal growth restriction, among infants born to ZIKV-infected mothers [13]. The possible associations of ZIKV infection to these health disorders compelled the World Health Organization to declare ZIKV a Public Health Emergency of International Concern on February 2016 [15]

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Conclusion