Strategies in Ebola virus disease (EVD) diagnostics at the point of care

Abstract

Ebola virus disease (EVD) is a devastating, highly infectious illness with a high mortality rate. The disease is endemic to regions of Central and West Africa, where there is limited laboratory infrastructure and trained staff. The recent 2014 West African EVD outbreak has been unprecedented in case numbers and fatalities, and has proven that such regional outbreaks can become a potential threat to global public health, as it became the source for the subsequent transmission events in Spain and the USA. The urgent need for rapid and affordable means of detecting Ebola is crucial to control the spread of EVD and prevent devastating fatalities. Current diagnostic techniques include molecular diagnostics and other serological and antigen detection assays; which can be time-consuming, laboratory-based, often require trained personnel and specialized equipment. In this review, we discuss the various Ebola detection techniques currently in use, and highlight the potential future directions pertinent to the development and adoption of novel point-of-care diagnostic tools. Finally, a case is made for the need to develop novel microfluidic technologies and versatile rapid detection platforms for early detection of EVD.

Keywords: Ebola virus; diagnostics; immunoassay; microfluidics; polymerase chain reaction.

Figure 1

The current unprecedented and past EVD epidemiological outbreak maps and models; (a) image of the regions affected by the 2014 West African Ebola Outbreak (Aylward et al. 2014). Regional maps of confirmed and probable cases, clockwise from top right: (regions with declared epidemics) Guinea, Sierra Leone and Liberia; (regions with subsequent transmission events) Mali, Texas and New York in the USA and Spain. Reprinted with copyright permission from Aylward et al. (2014). (b) 2014 outbreak growth of confirmed, probable and suspected cases of EVD in each affected country under epidemic crisis (Sierra Leone, Liberia and Guinea). Reprinted with copyright permission from Gire et al. (2014). (c) Temporal sequence of Ebola infection within a 48-h duration in an infected cell, demonstrating the events prior to detectable threshold. Green and orange particles represent common serological targets, VP40 and GP, respectively. Viral RNA is represented by blue and red transcripts. Reprinted with copyright permission from Roca et al. (2015).

Figure 2

The Genomic and Protein maps of Ebolavirus (Alazard-Dany et al. 2006). (top) The overall ~19kbp genomic structure of Ebola with: a nucleoprotein (NP), a polymerase cofactor (VP35), a matrix protein (VP40), a transmembrane glycoprotein (GP), the GP gene also codes for an edited mRNA for a soluble glycoprotein (sGP), a transcriptional activator (VP30), a viral envelope-associated protein (VP24) and a RNA–dependent RNA polymerase (L). (b) A rendered Ebola virus protein schematic, displaying the viral proteins in their appropriate roles of the overall viral structure. Reprinted with copyright permission from Lee and Saphire (2009).

Figure 3

RT-LAMP final cyclic reaction and elongation steps; (a) following the FIP primer binding, there is another round of polymerization. A reverse transcriptase binds the FIP primer making a cDNA copy. This step is continued with the BIP primer, which makes a template DNA strand from cDNA polymerization. The outside reaction is repeated in multiple rounds, allowing for mass amplification of the Ebola RNA target. Reprinted with copyright permission from Notomi et al. (2000). (b) Results of RT-LAMP for Ebola (Kurosaki et al. 2007) showing sensitivity of the amplified in vitro Ebola RNA at only 20 copies. (c) The duration of the Ebola genome amplification by RT-LAMP have shown to detect Ebola within 20 min. Reprinted with copyright permission from Kurosaki et al. (2007).

Figure 4

Magnetic bead actuation on a chip and whole-blood sampling isotachophoresis; (a) Functionalized magnetic beads with anti-CD4-antibodies in an HIV ELISA detection assay. Magnetic beads bind and capture CD-4 cells, and are actuated to each reaction chamber when a magnetic field is applied a colour change indicates a seropositive reaction in the final well. Reprinted with copyright permission from Wang et al. (2014). (b) Complex isotachophoresis on-a-chip designed to capture and isolate parasite DNA from infected leukocytes, using electric fields to generate electrophoretic separation of ionic samples. Reprinted with copyright permission from Marshall et al. (2011).

Figure 5

Immuno-PCR (iPCR) assay: iPCR technique with both variations of Enzyme-Linked Immunosorbent Assay (ELISA), the standard for Ebola serological diagnosis; iPCR differs from ELISA by using DNA amplification of an indicator a DNA for ultrasensitive. Reprinted with copyright permission from Akter et al. (2014). (b) The sensitivity of iPCR technique is enhanced using functionalized gold nanoparticle conjugated with multiple single stranded DNA oligonucleotides. Reprinted with copyright permission from Chen et al. (2009).

Figure 6

Alternative biosensing technologies for rapid and early diagnosis of Ebola. (a) Quartz crystal microbalance immunosensors are composed of a gold electrode layer with surface antibody capture agents, placed on top of a piezoelectric quartz crystal. Resonance frequency changes are measured as target binds to the antibody-coated surface, allowing for label-free detection. Reprinted with copyright permission from Kierny et al. (2012). (b) Surface plasmon resonance (SPR) biosensor schematic, composed of a channel with specific antibodies fixed on a gold layer. A measurement in changes of light reflected from an optic source allows for potentially highly sensitive diagnostic capability. Reprinted with copyright permission from Cooper (2002). (c) A nanostructured photonic crystal-based sensor functionalized with gp120 to selectively capture and detect HIV copies from whole blood. Reprinted with copyright permission from Shafiee et al. (2014). (d) Break junction functionalized with aptamer to capture cancer biomarkers through electrical impedance. Reprinted with copyright permission from Ilyas et al. (2012).