Direct detection of human adenovirus or SARS-CoV-2 with ability to inform infectivity using DNA aptamer-nanopore sensors

Abstract

Viral infections are a major global health issue, but no current method allows rapid, direct, and ultrasensitive quantification of intact viruses with the ability to inform infectivity, causing misdiagnoses and spread of the viruses. Here, we report a method for direct detection and differentiation of infectious from noninfectious human adenovirus and SARS-CoV-2, as well as from other virus types, without any sample pretreatment. DNA aptamers are selected from a DNA library to bind intact infectious, but not noninfectious, virus and then incorporated into a solid-state nanopore, which allows strong confinement of the virus to enhance sensitivity down to 1 pfu/ml for human adenovirus and 1 × 10 4 copies/ml for SARS-CoV-2. Applications of the aptamer-nanopore sensors in different types of water samples, saliva, and serum are demonstrated for both enveloped and nonenveloped viruses, making the sensor generally applicable for detecting these and other emerging viruses of environmental and public health concern.

Aptamers for Viral Detection and Inhibition

“… 259 , 261 , 267 The resulting higher binding affinity can facilitate the development of rapid, inexpensive, and sensitive strategies for virus sensing early after infection, which is critical for curbing the spread of highly contagious infectious diseases like COVID-19. 281 − 283 Additionally, aptamers can be strategically evolved to distinguish infectious virus particles from noninfectious forms, 231 which provides a unique and novel solution to address the problem for people being able to know when they are no longer infectious and can come out of quarantine, as nucleic acid tests are known to generate false positive results from the presence of nucleic acid molecules from degraded viruses. 284 , 285 These emerging platforms and technologies are well worth further investigations for the development of better viral sensors to mitigate future epidemics and pandemics.…”

“…(b) Scheme of infectious virus detection using aptamer-functionalized nanopore sensors. Reproduced with permission from ref ( 231 ). Copyright 2021 American Association for the Advancement of Science.…”

“…In addition to aforementioned aptamers that were selected by targeting specific SARS-CoV-2 surface antigens (N or S protein), Lu and colleagues have evolved an aptamer, called SARS2-AR10, that was selected against intact virions by performing a counter selection using UV-inactivated virus particles. 231 As a result, the aptamer can distinguish active/infectious SARS-CoV-2 virus from the noninfectious form. The SARS2-AR10 aptamer was integrated with a solid-state nanopore system, which renders strong confinement to the virus, to selectively detect intact SARS-CoV-2 containing samples with an LoD of 1 × 10 4 copies/mL ( Figure 7 b).…”

Recent advances of functional nucleic acid-based sensors for point-of-care detection of SARS-CoV-2

“…The assay has a 100-fold lower LOD than the lateral flow assay, 100% accuracy for clinical samples, and, more importantly, a cost of $0.40 per test, making it an attractive strategy for large-scale testing in resource-limited settings. Furthermore, for the direct detection of intact virions, Peinetti et al [ 61 ] selected DNA aptamers that bind intact infectious viruses. The aptamer is integrated into a solid nanopore, creating a strong constraint on the virus and strongly enhancing the sensitivity to SARS-CoV-2 intact virions.…”

One Solution for All: Searching for Universal Aptamers for Constantly Mutating Spike Proteins of SARS‐CoV‐2

“…With such benefits, it comes as no surprise that multiple labs around the world have produced aptamers relevant to diagnostic and therapeutic solution to SARS‐CoV‐2. Thus far, more than a dozen SELEX experiments have been conducted against the S protein, and over fifty aptamers have been derived [18–28] . Most of the aptamers display excellent affinity for the S protein, with K d (dissociation constant) values predominantly ranging between 1–20 nM.…”