Low ozone concentration and negative ions for rapid SARS-CoV-2 inactivation

Ozone is a powerful anti-bacterial, anti-fungal and anti-viral agent, yet exposure to high levels of ozone can pose risks to human/animal health and, in the long term, corrode certain objects. In order to overcome these risks, we evaluated the potential of using a relatively short exposure of a low concentration of ozone to disinfect an indoor environment in the absence of individuals and animals. ICON3 by O3ZONO/M2L, a new disinfection device generating both ozone and negative ions, was selected to assess the potential of this strategy to inactivate different viral isolates of SARS-CoV-2.Tests under controlled laboratory conditions were performed in a system consisting of an ozone-proof airtight plastic box inside a biological safety cabinet, where suspensions of two strains of SARS-CoV-2 were exposed to ozone and negative ions and virucidal activity was measured by means of two complementary methodologies: viral replication capacity and viral titer determination.These studies revealed that low concentration ozone (average 3.18 ppm after the peak) inactivated up to >99% of SARS-CoV-2 within 20 minutes of exposure. Under controlled conditions, similar ozone exposure was recreated with ICON3 in different volume rooms (15, 30, 60 m3) where a linear relationship was observed between the room volume and the time of continuous ozone/ions flow required to reach and maintain the desired ozone levels used in the laboratory studies.These studies suggest that ICON3 may have the potential for use in the disinfection of SARS-CoV-2 in indoor environments in the absence of individuals and animals, under properly controlled and monitored safety conditions.

ER stress and autophagy induced by SARS-CoV-2: The targets for melatonin treatment

Coronavirus disease 19 (COVID-19) is a viral disease caused by the new coronavirus SARS-CoV-2. Like other coronaviral infections, SARS-CoV-2 causes oxidative and ER stress triggering cellular response pathways, mainly PERK and IRE1 branches of the UPR. This excessive oxidative stress and the increasing of unfolded and misfolded proteins induce autophagy. Once this process is triggered, the blockage of the fusion of autophagosomes and lysosomes induced by virus leads to an incomplete autophagy. Double-membraned vesicles, which create a membranous support for viral RNA replication complexes, are formed. Melatonin is a pleiotropic molecule, which reduces oxidative and ER stress, regulates immune system, and modulates autophagy pathway. Thus, melatonin reinforces UPR and unlocks autophagy blockage, allowing autophagosomes to bind to lysosomes, completing the process of autophagy and decreasing viral replication capacity. Based on these activities of melatonin the recommendation of melatonin for patients with COVID-19 should be seriously considered, especially in elderlies and patients with different comorbidities, which are the highest risk population for serious cases.

Double trouble? Gag in conjunction with double insert in HIV protease contributes to reduced DRV susceptibility

HIV protease is essential for processing the Gag polyprotein to produce infectious virions and is a major target in antiretroviral therapy. We have identified an unusual HIV-1 subtype C variant that contains insertions of leucine and asparagine (L38↑N↑L) in the hinge region of protease at position 38. This was isolated from a protease inhibitor naïve infant. Isothermal titration calorimetry showed that 10% less of L38↑N↑L protease was in the active conformation as compared with a reference strain. L38↑N↑L protease displayed a ±50% reduction in KM and kcat. The catalytic efficiency (kcat/KM) of L38↑N↑L protease was not significantly different from that of wild type although there was a 42% reduction in specific activity for the variant. An in vitro phenotypic assay showed the L38↑N↑L protease to be susceptible to lopinavir (LPV), atazanavir (ATV) and darunavir in the context of an unrelated Gag. However, in the presence of the related Gag, L38↑N↑L showed reduced susceptibility to darunavir while remaining susceptible to LPV and ATV. Furthermore, a reduction in viral replication capacity (RC) was observed in combination with the related Gag. The reduced susceptibility to darunavir and decrease in RC may be due to PTAPP duplication in the related Gag. The present study shows the importance of considering the Gag region when looking at drug susceptibility of HIV-1 protease variants.

Mother-to-Child HIV Transmission Bottleneck Selects for Consensus Virus with Lower Gag-Protease-Driven Replication Capacity

ABSTRACT In the large majority of cases, HIV infection is established by a single variant, and understanding the characteristics of successfully transmitted variants is relevant to prevention strategies. Few studies have investigated the viral determinants of mother-to-child transmission. To determine the impact of Gag-protease-driven viral replication capacity on mother-to-child transmission, the replication capacities of 148 recombinant viruses encoding plasma-derived Gag-protease from 53 nontransmitter mothers, 48 transmitter mothers, and 47 infected infants were assayed in an HIV-1-inducible green fluorescent protein reporter cell line. All study participants were infected with HIV-1 subtype C. There was no significant difference in replication capacities between the nontransmitter (n = 53) and transmitter (n = 44) mothers (P = 0.48). Infant-derived Gag-protease NL4-3 recombinant viruses (n = 41) were found to have a significantly lower Gag-protease-driven replication capacity than that of viruses derived from the mothers (P < 0.0001 by a paired t test). High percent similarities to consensus subtype C Gag, p17, p24, and protease sequences were also found in the infants (n = 28) in comparison to their mothers (P = 0.07, P = 0.002, P = 0.03, and P = 0.02, respectively, as determined by a paired t test). These data suggest that of the viral quasispecies found in mothers, the HIV mother-to-child transmission bottleneck favors the transmission of consensus-like viruses with lower viral replication capacities. IMPORTANCE Understanding the characteristics of successfully transmitted HIV variants has important implications for preventative interventions. Little is known about the viral determinants of HIV mother-to-child transmission (MTCT). We addressed the role of viral replication capacity driven by Gag, a major structural protein that is a significant determinant of overall viral replicative ability and an important target of the host immune response, in the MTCT bottleneck. This study advances our understanding of the genetic bottleneck in MTCT by revealing that viruses transmitted to infants have a lower replicative ability as well as a higher similarity to the population consensus (in this case HIV subtype C) than those of their mothers. Furthermore, the observation that “consensus-like” virus sequences correspond to lower in vitro replication abilities yet appear to be preferentially transmitted suggests that viral characteristics favoring transmission are decoupled from those that enhance replicative capacity.