Longer amplicons provide better sensitivity for electrochemical sensing of viral nucleic acid in water samples using PCB electrodes

Abstract

The importance of monitoring environmental samples has gained a lot of prominence since the onset of COVID-19 pandemic, and several surveillance efforts are underway using gold standard, albeit expensive qPCR-based techniques. Electrochemical DNA biosensors could offer a potential cost-effective solution suitable for monitoring of environmental water samples in lower middle income countries. In this work, we demonstrate electrochemical detection of amplicons as long as $${503}\,\hbox {bp}$$ 503 bp obtained from Phi6 bacteriophage (a popular surrogate for SARS-CoV-2) isolated from spiked lake water samples, using ENIG finish PCB electrodes with no surface modification. The electrochemical sensor response is thoroughly characterised for two DNA fragments of different lengths ($${117}\,\hbox {bp}$$ 117 bp and $${503}\,\hbox {bp}$$ 503 bp ), and the impact of salt in PCR master mix on methylene blue (MB)-DNA interactions is studied. Our findings establish that length of the DNA fragment significantly determines electrochemical sensitivity, and the ability to detect long amplicons without gel purification of PCR products demonstrated in this work bodes well for realisation of fully-automated solutions for in situ measurement of viral load in water samples.

Viricidal Efficacy of a 405‐nm Environmental Decontamination System for Inactivation of Bacteriophage Phi6: Surrogate for SARS‐CoV‐2

“…Phi6 is a dsRNA lytic bacteriophage of the Cystoviridae virus family which infects Pseudomonas bacteria ( 40 , 41 ). It possesses similarities to that of coronaviruses, namely, it is of similar size, has spike proteins and is enveloped by a lipid membrane ( 40 , 41 , 42 ), and thus has been suggested and utilized as a surrogate for the study of SARS‐CoV‐2 in multiple publications ( 35 , 42 , 43 , 44 , 45 , 46 , 47 , 48 , 49 , 50 , 51 ).…”

Electrochemical detection of myeloperoxidase (MPO) in blood plasma with surface-modified electroless nickel immersion gold (ENIG) printed circuit board (PCB) electrodes

“…To accomplish this, we electrodeposited methylene blue dye diluted in DI water on the PCB electrode, without applying large voltages or using corrosive chemicals. Methylene blue dye is a phenothiazine dye and is used as a redox probe in many applications due to its electrochemical activity [25,27,[36][37][38][39]. Typically, a high potential (>0.8 V) and low pH (5) are needed for the formation of cation radicals that initiate the polymerization process, and ensure stability of the polymer film [40].…”

Biosensors for waterborne virus detection: Challenges and strategies

“…Traditional methods such as nucleic acid amplification techniques and immunology-based methods such as real-time quantitative polymerase chain reaction (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA) can be time-consuming and require well-trained personnel and sophisticated equipment, preventing their broad implementation [ 8 ]. In regard to low- and middle-income countries (LMICs) with limited resources, the need for accessible and affordable virus detection methods is becoming even more urgent [ 9 ]. Biosensors are powerful analytical tools with relatively low cost, fast, and even near real-time response compared to the aforementioned conventional analytical methods [ 10 ].…”