Zika Virus Diagnostics in Endemic Areas: Conventional Methods, Biofluid Kinetics, and Emerging Point-of-Care Technologies

Zika virus (ZIKV) is an Aedes-borne flavivirus that remains an important public health concern because infection can cause congenital Zika syndrome and neurological complications, including Guillain–Barré syndrome. Laboratory diagnosis is challenging because ZIKV infection may resemble dengue and chikungunya clinically, while serological assays can show substantial cross-reactivity with other flaviviruses. This narrative review summarizes conventional and emerging approaches to ZIKV diagnosis, with emphasis on their application in endemic and resource-limited settings. Molecular nucleic acid amplification tests (NAATs), particularly real-time reverse-transcription polymerase chain reaction (RT-qPCR), are preferred for detecting acute infection when viral RNA is present. However, the period of RNA detectability in serum or plasma is relatively short, and a negative molecular result does not necessarily exclude recent infection. Viral RNA may persist longer in other specimens, including urine and whole blood, while ZIKV RNA may remain detectable in semen for several months in some infected men. Serological methods, particularly IgM assays, can support diagnosis after the acute phase but are limited by flavivirus cross-reactivity and prolonged antibody persistence. Plaque reduction neutralization testing (PRNT) may assist interpretation in selected circumstances, although it may not reliably distinguish ZIKV from other flaviviruses in individuals with previous flavivirus infection or vaccination. Emerging approaches, including reverse-transcription loop-mediated isothermal amplification (RT-LAMP), recombinase polymerase amplification (RPA), CRISPR-Cas-based detection systems, microfluidic platforms, biosensors, and paper-based or smartphone-assisted assays, offer potential advantages for decentralized testing. Nevertheless, many of these technologies remain under development or require further clinical validation before widespread implementation. Future diagnostic strategies should integrate analytical performance with specimen selection, timing of testing, epidemiological context, quality assurance, affordability, and feasibility of deployment in endemic regions.