Biology subjects

Leibowitz, J. L.

Publications and source records attributed to Leibowitz, J. L..

4 recordsLinked to original sources

bioRxiv2023

Uncovering the Vital Role of Lipid Droplets in Coronavirus Replication: A Novel Insight Arising from the Biological Activity of 2-Bromopalmitate

The emergence of viral infections with global impact highlights the urgent need for broad-spectrum antivirals. In this study, we evaluated the effect of palmitoylation inhibitors [2-bromopalmitate (2-BP), cerulenin, and 2-fluoro palmitic acid (2-FPA)] and the enhancer palmostatin B on the replication of human coronaviruses (hCoV-229E, hCoV-Oc43) and murine hepatitis virus (MHV-A59) at non-cytotoxic concentrations. The results demonstrated that 2-BP strongly suppressed MHV-A59 replication, while cerulenin and 2-FPA only moderately inhibited viral replication. Palmostatin B significantly enhanced viral replication. Notably, 2-BP exhibited superior efficacy. Interestingly, palmostatin B failed to rescue the inhibitory effects of 2-BP but effectively rescued cerulenin and 2-FPA, suggesting additional biological activities of 2-BP beyond palmitoylation inhibition. Furthermore, we discovered that 2-BP specifically disrupted lipid droplets (LDs), and this LD disruption was correlated with viral replication inhibition. Based on our findings, we conclude that the inhibitory effects of 2-BP on viral replication primarily stem from LD disruption rather than palmitoylation inhibition. Therefore, we revealed the crucial role of LDs in the viral replication. Our study provides insights into the development of wide-spectrum antiviral strategies.

microbiology↗

bioRxiv2023

Infection of equine bronchial epithelial cells with a SARS-CoV-2 pseudovirus

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causal agent of COVID-19, can infect animals by binding to the angiotensin-converting enzyme 2 (ACE2). Equine infection appears possible due to high homology ({approx}97%) between human and equine ACE2, evidence of in vitro infection in cell lines expressing equine ACE2, and evidence of seroconversion in horses after exposure to persons infected with SARS-CoV-2. Our objective was to examine susceptibility of cultured primary equine bronchial epithelial cells (EBECs) to a SARS-CoV-2 pseudovirus relative to human bronchial epithelial cells (HBECs; positive control). ACE2 expression in EBECs detected by immunofluorescence, western immunoblotting, and flow cytometry was lower in EBECs than in HBECs. EBECs were transduced with a lentivirus pseudotyped with the SARS-CoV-2 spike protein that binds to ACE2 and expresses the enhanced green fluorescent protein (eGFP) as a reporter. Cells were co-cultivated with the pseudovirus at a multiplicity of infection of 0.1 for 6 hours, washed, and maintained in media. After 96 hours, eGFP expression in EBECs was demonstrated by fluorescence microscopy, and mean {Delta} Ct values from quantitative PCR were significantly (P < 0.0001) higher in HBECs (8.78) than HBECs (3.24) indicating lower infectivity in EBECs. Equine respiratory tract cells were susceptible to infection with a SARS-CoV-2 pseudovirus. Lower replication efficiency in EBECs suggests that horses are unlikely to be an important zoonotic host of SARS-CoV-2, but viral mutations could render some strains more infectious to horses. Serological and virological monitoring of horses in contact with persons shedding SARS-CoV-2 is warranted. IMPORTANCEThis study provides the first published evidence for SARS-CoV-2 pseudovirus infection in equine airway epithelial cells, which were less susceptible to infection than cells of human origin. This was presumably due to lower ACE2 expression in equine cells, lower viral affinity for equine ACE2, or both. Our results are important considering recent evidence for asymptomatic seroconversion in horses following exposure to COVID-19 positive humans, despite this lower susceptibility, and increased affinity of viral variants of concern for equine ACE2 compared to ancestral strains. Thus, there is great need to better characterize SARS-CoV-2 susceptibility in horses for the benefit of veterinary and human health.

microbiology↗

bioRxiv2020

Sub-second heat inactivation of coronavirus

Heat treatment denatures viral proteins that comprise the virion, making virus incapable of infecting a host. Coronavirus (CoV) virions contain single-stranded RNA genomes with a lipid envelope and 4 proteins, 3 of which are associated with the lipid envelope and thus are thought to be easily denatured by heat or surfactant-type chemicals. Prior studies have shown that a temperature of as low as 75 {degrees}C and treatment duration of 15 min can effectively inactivate CoV. The applicability of a CoV heat inactivation method greatly depends on the length of time of a heat treatment and the temperature needed to inactivate the virus. With the goal of finding conditions where sub-second heat exposure of CoV can sufficiently inactivate CoV, we designed and developed a simple system that can measure sub-second heat inactivation of CoV. The system is composed of capillary stainless-steel tubing immersed in a temperature-controlled oil bath followed by an ice bath, through which virus solution can be flowed at various speeds. Flowing virus solution at different speeds, along with a real-time temperature monitoring system, allows the virus to be accurately exposed to a desired temperature for various durations of time. Using mouse hepatitis virus (MHV), a beta-coronavirus, as a model system, we identified that 85.2 {degrees}C for 0.48 s exposure is sufficient to obtain > 5 Log10 reduction in viral titer (starting titer: 5 x 107 PFU/mL), and that when exposed to 83.4 {degrees}C for 0.95 s, the virus was completely inactivated (zero titer, > 6 Log10 reduction). IMPORTANCEThree coronaviruses (CoVs) have now caused global outbreaks within the past 20 years, with the COVID19 pandemic caused by SARS-CoV-2 still ongoing. Methods that can rapidly inactivate viruses, especially CoVs, can play critical roles in ensuring public safety and safeguarding personal health. Heat treatment of viruses to inactive them can be an efficient and inexpensive method, with the potential to be incorporated into various human-occupied spaces. In this work, a simple system that can heat-treat viruses for extremely short period was developed and utilized to show that sub-second exposure of CoV to heat is sufficient to inactivate CoV. This opens up the possibility of developing instruments and methods of disinfecting CoV in diverse settings, including rapid liquid disinfection and airborne virus disinfection. The developed method can also be broadly utilized to assess heat sensitivity of viruses other viral pathogens of interest and develop sub-second rapid heat inactivation approaches.

bioengineering↗

bioRxiv2019

A single dose of dietary nitrate increases maximal muscle speed and power in healthy older men and women

BackgroundAging results in reductions in maximal muscular strength, speed, and power, which often lead to functional limitations highly predictive of disability, institutionalization, and mortality in the elderly. This may be partially due to reduced nitric oxide (NO) bioavailability. We therefore hypothesized that dietary nitrate (NO3-), a source of NO via the NO3- [->] nitrite (NO2-) [->] NO enterosalivary pathway, could increase muscle contractile function in older subjects. MethodsTwelve healthy older (age 71{+/-}1 y) subjects were studied using a randomized, doubleblind, placebo-controlled, crossover design. After fasting overnight, subjects were tested 2 h after ingesting beetroot juice containing or devoid of 13.4{+/-}0.5 mmol NO3-. Plasma NO3- and NO2- and breath NO were measured periodically, and muscle function was determined using isokinetic dynamometry. ResultsNO3- ingestion increased (P<0.001) plasma NO3-, plasma NO2-, and breath NO by 1050%, 140%, and 110%, respectively. Maximal velocity of knee extension increased (P<0.01) by 1.1{+/-}0.3 rad/s, or by 10.9{+/-}3.5%. Maximal knee extensor power increased (P<0.05) by 0.14{+/-}0.06 W/kg, or by 4.4{+/-}2.3%. ConclusionsAcute dietary NO3- intake improves muscle contractility in older individuals. These findings have important implications for this population, in whom diminished muscle function can lead to functional limitations, dependence, and even premature death.

physiology↗