Plate-Based High-Throughput Fluorescence Assay for Assessing Enveloped Virus Integrity.

Viruses are a considerable threat to global health and place major burdens on economies worldwide. Manufactured viruses are also being widely used as delivery agents to treat (gene therapies) or prevent diseases (vaccines). Therefore, it is vital to study and fully understand the infectious state of viruses. Current techniques used to study viruses are often slow or nonexistent, making the development of new techniques of paramount importance. Here we present a high-throughput and robust, cell-free plate-based assay (FAIRY: Fluorescence Assay for vIRal IntegritY), capable of differentiating intact from nonintact enveloped viruses, i.e, infectious from noninfectious. Using a thiazole orange-terminated polymer, a 99% increase in fluorescence was observed between treated (heat or virucide) and nontreated. The FAIRY assay allowed for the rapid determination of the infectivity of a range of enveloped viruses, highlighting its potential as a valuable tool for the study of viruses and interventions against them.

Bridging Flocculation of a Sterically Stabilized Cationic Latex as a Biosensor for the Detection of Microbial DNA after Amplification via PCR.

There is a high demand for rapid, sensitive, and accurate detection methods for pathogens. This paper demonstrates a method of detecting the presence of amplified DNA from a range of pathogens associated with serious infections including Gram-negative bacteria, Gram-positive bacteria, and viruses. DNA is amplified using a polymerase chain reaction (PCR) and consequently detected using a sterically stabilized, cationic polymer latex. The DNA induces flocculation of this cationic latex, which consequently leads to rapid sedimentation and a visible change from a milky-white dispersion to one with a transparent supernatant, presenting a clear visible change, indicating the presence of amplified DNA. Specifically, a number of different pathogens were amplified using conventional or qPCR, including Staphylococcus aureus, Escherichia coli, and Herpes Simplex Virus (HSV-2). This method was demonstrated to detect the presence of bacteria in suspension concentrations greater than 380 CFU mL-1 and diagnose the presence of specific genomes through primer selection, as exemplified using methicillin resistant and methicillin susceptible Staphylococcus aureus. The versatility of this methodology was further demonstrated by showing that false positive results do not occur when a PCR of fungal DNA from C. albicans is conducted using bacterial universal primers.

Broad-Spectrum Extracellular Antiviral Properties of Cucurbit[n]urils.

Viruses are microscopic pathogens capable of causing disease and are responsible for a range of human mortalities and morbidities worldwide. They can be rendered harmless or destroyed with a range of antiviral chemical compounds. Cucurbit[n]urils (CB[n]s) are a family of macrocycle chemical compounds existing as a range of homologues; due to their structure, they can bind to biological materials, acting as supramolecular "hosts" to "guests", such as amino acids. Due to the increasing need for a nontoxic antiviral compound, we investigated whether cucurbit[n]urils could act in an antiviral manner. We have found that certain cucurbit[n]uril homologues do indeed have an antiviral effect against a range of viruses, including herpes simplex virus 2 (HSV-2), respiratory syncytial virus (RSV) and SARS-CoV-2. In particular, we demonstrate that CB[7] is the active homologue of CB[n], having an antiviral effect against enveloped and nonenveloped species. High levels of efficacy were observed with 5 min contact times across different viruses. We also demonstrate that CB[7] acts with an extracellular virucidal mode of action via host-guest supramolecular interactions between viral surface proteins and the CB[n] cavity, rather than via cell internalization or a virustatic mechanism. This finding demonstrates that CB[7] acts as a supramolecular virucidal antiviral (a mechanism distinct from other current extracellular antivirals), demonstrating the potential of supramolecular interactions for future antiviral disinfectants.