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Review Article

Advances in Diagnostic Methods for Zika Virus Infection

[+] Author and Article Information
Carlos A. Herrada, Md. Alamgir Kabir, Rommel Altamirano

Department of Computer Engineering and
Electrical Engineering and Computer Science,
Florida Atlantic University,
Boca Raton, FL 33431;
Asghar-Lab, Micro and
Nanotechnology in Medicine,
College of Engineering and Computer Science,
Boca Raton, FL 33431

Waseem Asghar

Department of Computer Engineering and
Electrical Engineering and Computer Science,
Florida Atlantic University,
Boca Raton, FL 33431;
Asghar-Lab, Micro and
Nanotechnology in Medicine,
College of Engineering and Computer Science,
Boca Raton, FL 33431;
Department of Biological Sciences,
Florida Atlantic University,
Boca Raton, FL 33431
e-mail: wasghar@fau.edu

1Corresponding author.

Manuscript received March 13, 2018; final manuscript received July 31, 2018; published online November 5, 2018. Assoc. Editor: Yaling Liu.

J. Med. Devices 12(4), 040802 (Nov 05, 2018) (11 pages) Paper No: MED-18-1054; doi: 10.1115/1.4041086 History: Received March 13, 2018; Revised July 31, 2018

The Zika virus (ZIKV) is one of the most infamous mosquito-borne flavivirus on recent memory due to its potential association with high mortality rates in fetuses, microcephaly and neurological impairments in neonates, and autoimmune disorders. The severity of the disease, as well as its fast spread over several continents, has urged the World Health Organization (WHO) to declare ZIKV a global health concern. In consequence, over the past couple of years, there has been a significant effort for the development of ZIKV diagnostic methods, vaccine development, and prevention strategies. This review focuses on the most recent aspects of ZIKV research which includes the outbreaks, genome structure, multiplication and propagation of the virus, and more importantly, the development of serological and molecular detection tools such as Zika IgM antibody capture enzyme-linked immunosorbent assay (Zika MAC-ELISA), plaque reduction neutralization test (PRNT), reverse transcription quantitative real-time polymerase chain reaction (qRT-PCR), reverse transcription-loop mediated isothermal amplification (RT-LAMP), localized surface plasmon resonance (LSPR) biosensors, nucleic acid sequence-based amplification (NASBA), and recombinase polymerase amplification (RPA). Additionally, we discuss the limitations of currently available diagnostic methods, the potential of newly developed sensing technologies, and also provide insight into future areas of research.

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Figures

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Fig. 1

Geographical distribution of ZIKV outbreak from 2007 to 2017. This image illustrates the spread of ZIKV from its inception in the small island of Yap to its conclusion in the Americas [18] (Reprinted from The Lancet with permission from Elsevier © 2017).

Grahic Jump Location
Fig. 2

Zika virus external morphology and viral proteins. This illustration depicts the encoded structural components contain the capsid and membrane proteins [29] (Reprinted with permission from The American Society for Microbiology © 2017). The nonstructural portion of the ORF includes an envelope protein and 7 other proteins; NS1, NS2a, NS2b, NS3, NS4a, NS4b, and NS5 [30] (Reprinted with permission of Creative Commons Attribution BY 4.0).

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Fig. 3

CDC recommended tests for ZIKV detection. (a) In RT-qPCR, viral RNA extracted from an infected patient's sample is placed into a reaction mix containing all of the necessary factors for amplification such as ZIKV specific primers (to bind to the RNA template), reverse transcriptase (to make cDNA from RNA), DNA polymerase and dNTPs (to amplify the DNA), a buffer solution (to maintain an optimal environment for the polymerase), and an intercalating fluorescent dye for quantification. Amplification takes place through thermal cycling, and its product is later identified and quantified based on fluorescence. (b) In MAC ELISA, the activity of IgM antibodies is measured in response to ZIKV infection. The process starts when a patient's blood sample is incubated in a well plate containing antibodies against IgM. If IgM is present, then it will strongly bind to the antibodies in the plate. If not, then the patient's sample will be washed away and there would be no reaction when the secondary HRP antibody is added to the plate. (c) PRNT tests are carried out to confirm serological results. In PRNT, a patient's serum undergoes a series of dilutions that are then added to a ZIKV viral suspension, mixed, and incubated alongside cell cultures. If antibodies against ZIKV are present, then there will be a reduction in the number of observable ZIKV plaques [64] (Reprinted with permission of Creative Commons Attribution BY 4.0).

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Fig. 4

Localized surface plasmon resonance biosensor schematics and ZIKV sensitivity. (a) Qdot bimetallic nanohybrids are made by coupling plasmonic nanoparticles such as gold to quantum dots. These hybrids are then infused alongside ZIKV specific DNA loops to the molecular beacon of the biosensor. (b) LSPR relies on the resonance that occurs through the hybridization of viral ZIKV RNA with the complementary DNA loop sequence found in the molecular beacon. This in turn induces a signal that can be picked up, amplified and quantified by nearby quantum dot nanocrystals [77]. (Reprinted with permission of Elsevier © 2017).

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Fig. 5

CRISPR/CAS9 biomolecular sensors. (a) Primers and toehold sensors are selectively designed to match specific ZIKV strains. These sensors, alongside their primers are then infused and freeze-dried into paper effectively completing the biosensor. (b) Adding a patient's sample (fluid) to the paper will cause it to become moisturized and active. If a specific ZIKV strain is present, then a color change from yellow to purple will take place on the surface of the paper, which would be indicative of a positive reaction. This sensor is specific enough to differentiate between closely related flaviviruses and even different ZIKV lineages within a single base resolution, when coupled to a CRISPR/Cas9-based module [78] (Reprinted with permission of Elsevier © 2016).

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Fig. 6

Isothermal amplification technologies; RT-LAMP and RPA amplification. (a) Patients' samples are collected and lysed to extract viral nucleotides. (b) The lysed samples are then filtered through a porous silica membrane inside the microfluidic cassette to isolate ZIKV nucleic acids. Once the nucleotides have been isolated, a colorless leuco crystal violet indicator is pumped into the reaction chamber (where the filter membrane is located) to serve as a marker for amplification. (c) The inside of the device consists of a thermos that has been modified to hold a microfluidic cassette, a heat sink, an isothermal heat source (Mg-Fe alloy), and a heat sink to dissipate heat, while the outside (d) consists of a 3D printed lid for sample collection, and a water port to trigger the isothermal reaction. (e) Photographs depicting the isothermal amplification reactor prior and after use. If ZIKV is found, then the colorless crystal violet indicator reacts and becomes violet in color [81] (Reprinted with permission of ACS Publications: https://pubs.acs.org/doi/10.1021/acs.analchem.6b01632). (f) This modified 3D printer is able to perform high throughput nucleotide extraction, and isothermal amplification all within the same enclosure. Due to its configuration, you can run up to twelve patient samples per run, as this printer holds 96 well plates. The heated bed underneath the plate maintains the temperature at a constant rate, as to not affect the amplification process. (g) RPA amplification can also be performed within a thermos like structure as seen in RT-LAMP. However, this process is not as efficient, as you cannot run as many samples in it [85] (Reprinted with permission of Elsevier © 2018).

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