Gold-Oligonucleotide Nanoconstructs Engineered to Detect Conserved Enteroviral Nucleic Acid Sequences.

Veeren M Chauhan,Mohamed M Elsutohy,Jonathan W Aylott,C Patrick Mcclure,William L Irving,Neil Roddis

doi:10.3390/bios11070238

Abstract

Enteroviruses are ubiquitous mammalian pathogens that can produce mild to life-threatening disease. We developed a multimodal, rapid, accurate and economical point-of-care biosensor that can detect nucleic acid sequences conserved amongst 96% of all known enteroviruses. The biosensor harnesses the physicochemical properties of gold nanoparticles and oligonucleotides to provide colourimetric, spectroscopic and lateral flow-based identification of an exclusive enteroviral nucleic acid sequence (23 bases), which was identified through in silico screening. Oligonucleotides were designed to demonstrate specific complementarity towards the target enteroviral nucleic acid to produce aggregated gold–oligonucleotide nanoconstructs. The conserved target enteroviral nucleic acid sequence (≥1 × 10−7 M, ≥1.4 × 10−14 g/mL) initiates gold–oligonucleotide nanoconstruct disaggregation and a signal transduction mechanism, producing a colourimetric and spectroscopic blueshift (544 nm (purple) > 524 nm (red)). Furthermore, lateral-flow assays that utilise gold–oligonucleotide nanoconstructs were unaffected by contaminating human genomic DNA, demonstrated rapid detection of conserved target enteroviral nucleic acid sequence (<60 s), and could be interpreted with a bespoke software and hardware electronic interface. We anticipate that our methodology will translate in silico screening of nucleic acid databases to a tangible enteroviral desktop detector, which could be readily translated to related organisms. This will pave the way forward in the clinical evaluation of disease and complement existing strategies to overcome antimicrobial resistance.

Highlights

We describe the development of a multimodal gold–oligonucleotide nanoconstruct-based biosensor composed of gold nanoparticles and an in silico designed oligonucleotide sequence that can determine the presence of a target nucleic acid sequence, which is conserved amongst all known enteroviruses, by demonstrating a colourimetric change that can be detected spectroscopically or through the use of lateral flow
When oligonucleotides hybridise with the target nucleic acid sequences, gold nanoparticles disaggregate, initiating a signal transduction pathway (Figure 1), which can be characterised via colourimetric, spectroscopic, and lateral flow assays
Gold–oligonucleotide nanoconstructs sensitive to enteroviral nucleic acids were fabricated via a systematic methodology, which included identification of target nucleic acid and in silico oligonucleotide design, synthesis of gold nanoparticles and functionalisation of their surfaces, and characterisation of the signal transduction pathway

Summary

Introduction

The enterovirus genus currently consists of 15 species (Enterovirus A-L and Rhinovirus A-C) [1], with potentially new species still being identified [2,3]. These species are associated with mild and serious disease [4], which include the common cold [5] and poliomyelitis [6], respectively. Unlike enteroviral infections, can be effectively treated with antibiotics [7]; if they are left untreated, they can become life-threatening [8]. An improved understanding of infectious organisms [11] and their detection [12] could enhance therapeutics that effectively cure and alleviate the symptoms of disease

Methods

Results

Conclusion