Mechanisms of SARS-CoV-2 Inactivation Using UVC Laser Radiation.

Severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2) has had a tremendous impact on humanity. Prevention of transmission by disinfection of surfaces and aerosols through a chemical-free method is highly desirable. Ultraviolet C (UVC) light is uniquely positioned to achieve inactivation of pathogens. We report the inactivation of SARS-CoV-2 virus by UVC radiation and explore its mechanisms. A dose of 50 mJ/cm2 using a UVC laser at 266 nm achieved an inactivation efficiency of 99.89%, while infectious virions were undetectable at 75 mJ/cm2 indicating >99.99% inactivation. Infection by SARS-CoV-2 involves viral entry mediated by the spike glycoprotein (S), and viral reproduction, reliant on translation of its genome. We demonstrate that UVC radiation damages ribonucleic acid (RNA) and provide in-depth characterization of UVC-induced damage of the S protein. We find that UVC severely impacts SARS-CoV- 2 spike protein's ability to bind human angiotensin-converting enzyme 2 (hACE2) and this correlates with loss of native protein conformation and aromatic amino acid integrity. This report has important implications for the design and development of rapid and effective disinfection systems against the SARS-CoV-2 virus and other pathogens.

Development of a rapid plasma decontamination system for decontamination and reuse of filtering facepiece respirators.

The COVID-19 pandemic has caused a high demand for filtering facepiece respirators (FFRs), which has brought global challenges in sustaining the supply chain for FFRs. Because respirators are basic personal protective equipment to protect frontline healthcare workers against COVID-19, the chronic, global shortage of N95/N99 masks is one of the most urgent threats to our collective ability to save lives from the coronavirus. The reuse of masks may need to be considered as a crisis capacity strategy to ensure continued availability even though most of the masks are considered one-time use. Moreover, environmentalists warn that single-use masks add to the glut of plastic pollution, threatening the health of oceans and marine life. In this study, we develop a method to decontaminate respirators to reuse filtering facepiece respirators. Samples of SARS-CoV-2 are applied to the 4 × 4 cm2 samples of FFP2 and FFP3 respirator materials. The filtration efficiency of plasma treated samples is measured using a planar particle image velocimetry technique with a neutrally charged polydisperse aerosol particle of NaCl. The measured viral decontamination and filtration efficiencies show that the developed plasma decontamination system can achieve a 4-log reduction for the coronavirus without reducing the filtration efficiency of masks after 5-min plasma exposure. The developed plasma decontamination system demonstrates the feasibility to tackle the acute shortages of FFRs in many countries and their environmental and economic burdens against discarding reusable masks.

Persistent residual contamination in endoscope channels; a fluorescence epimicroscopy study.

BACKGROUND AND STUDY AIMS: The increasing demand for endoscopic procedures poses new contamination challenges, given developing antimicrobial resistance worldwide and potential viral or prion diseases in populations at risk. We examined working channels from reusable luminal endoscopes used in recent years. METHODS: Very sensitive fluorescence epimicroscopy was used to examine working channels from 6 decommissioned and 6 factory-new channels, as received, or following spiking and washing in the laboratory. RESULTS: After a single contamination and wash test cycle, new channels retained approximately 75 pg/mm(2) of proteins; through 7 subsequent cycles residual proteins fluctuated between 25 and 75 pg/mm(2). Decommissioned channels harbored 1 - 4 µg of proteins each, except in one gastroscope (33 µg), including up to 2 % amyloid proteins except in one gastroscope and one sigmoidoscope (with over 80 %); lumens showed wearing with established abraded biofilms in 3 cases. After spiking with scrapie-infected blood components and washing, residual protein levels in new channels varied following standard (17.23 pg/mm(2)), duplicated (2.39 pg/mm(2)) or extended (11.3 pg/mm(2)) washing; no changes were measured among the long-established contamination in old channels. CONCLUSIONS: Our observations suggest that wear effects in endoscope lumens may contribute to the adsorption of proteins, thus facilitating retention and survival of bacteria. As demonstrated by recent outbreaks worldwide despite recommended reprocessing, the development of antimicrobial-resistant bacterial strains, and the estimated prevalence of variant Creutzfeldt-Jakob disease (vCJD) in the UK particularly, combined with increasing demand for endoscopic procedures, call for sustained precautions and improved methods for the reprocessing of nonautoclavable, reusable surgical instruments.

Down-regulation of DNA mismatch repair enhances initiation and growth of neuroblastoma and brain tumour multicellular spheroids.

Multicellular tumour spheroid (MCTS) cultures are excellent model systems for simulating the development and microenvironmental conditions of in vivo tumour growth. Many documented cell lines can generate differentiated MCTS when cultured in suspension or in a non-adhesive environment. While physiological and biochemical properties of MCTS have been extensively characterized, insight into the events and conditions responsible for initiation of these structures is lacking. MCTS are formed by only a small subpopulation of cells during surface-associated growth but the processes responsible for this differentiation are poorly understood and have not been previously studied experimentally. Analysis of gene expression within spheroids has provided clues but to date it is not known if the observed differences are a cause or consequence of MCTS growth. One mechanism linked to tumourigenesis in a number of cancers is genetic instability arising from impaired DNA mismatch repair (MMR). This study aimed to determine the role of MMR in MCTS initiation and development. Using surface-associated N2a and CHLA-02-ATRT culture systems we have investigated the impact of impaired MMR on MCTS growth. Analysis of the DNA MMR genes MLH1 and PMS2 revealed both to be significantly down-regulated at the mRNA level compared with non-spheroid-forming cells. By using small interfering RNA (siRNA) against these genes we show that silencing of MLH1 and PMS2 enhances both MCTS initiation and subsequent expansion. This effect was prolonged over several passages following siRNA transfection. Down-regulation of DNA MMR can contribute to tumour initiation and progression in N2a and CHLA-02-ATRT MCTS models. Studies of surface-associated MCTS differentiation may have broader applications in studying events in the initiation of cancer foci.