Some properties of branching processes with random control functions and affected by viral infectivity in random environments.

In this paper, a model of branching processes with random control functions and affected by viral infectivity in independent and identically distributed random environments is established, and the Markov property of the model and a sufficient condition for the model to be certainly extinct under some conditions are discussed. Then, the limit properties of the model are studied. Under the normalization factor {Sn:n∈N}, the normalization processes {Wˆn:n∈N} are studied, and the sufficient conditions of {Wˆn:n∈N} a.s., L1 and L2 convergence are given; A sufficient condition and a necessary condition for convergence to a nondegenerate at zero random variable are obtained. Under the normalization factor {In:n∈N}, the normalization processes {W¯n:n∈N} are studied, and the sufficient conditions of {W¯n:n∈N} a.s., and L1 convergence are obtained.

High-throughput screening of mutations affecting SARS-CoV-2 spike functions.

The spike (S) protein of SARS-CoV-2, which is undergoing rapid evolution, plays crucial roles in viral immune escape, infectivity, and transmissibility. To gain clinical insight, Dadonaite et al. developed a novel deep mutational scanning (DMS) platform for mapping the effects of S protein mutations on immune evasion and viral infectivity.

Role of cholesterol-recognition motifs in the infectivity of SARS-CoV-2 variants.

The presence of linear amino acid motifs with the capacity to recognize the neutral lipid cholesterol, known as Cholesterol Recognition/interaction Amino acid Consensus sequence (CRAC), and its inverse or mirror image, CARC, has recently been reported in the primary sequence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike S homotrimeric glycoprotein. These motifs also occur in the two other pathogenic coronaviruses, SARS-CoV, and Middle-East respiratory syndrome CoV (MERS-CoV), most conspicuously in the transmembrane domain, the fusion peptide, the amino-terminal domain, and the receptor binding domain of SARS-CoV-2 S protein. Here we analyze the presence of cholesterol-recognition motifs in these key regions of the spike glycoprotein in the pathogenic CoVs. We disclose the inherent pathophysiological implications of the cholesterol motifs in the virus-host cell interactions and variant infectivity.

The Role of Host Cell Glycans on Virus Infectivity: The SARS-CoV-2 Case.

Glycans are ubiquitously expressed sugars, coating the cell and protein surfaces. They are found on many proteins as either short and branched chains or long chains sticking out from special membrane proteins, known as proteoglycans. This sugar cushion, the glycocalyx, modulates specific interactions and protects the cell. Here it is shown that both the expression of proteoglycans and the glycans expressed on the surface of both the host and virus proteins have a critical role in modulating viral attachment to the cell. A mathematical model using SARS-Cov-2 as an archetypical virus to study the glycan role during infection is proposed. It is shown that this occurs via a tug-of-war of forces. On one side, the multivalent molecular recognition that viral proteins have toward specific host glycans and receptors. On the other side, the glycan steric repulsion that a virus must overcome to approach such specific receptors. By balancing both interactions, viral tropism can be predicted. In other words, the authors can map out the cells susceptible to virus infection in terms of receptors and proteoglycans compositions.

Evaporation of Water from Respiratory Droplets: A Review

Airborne transmission of virus-laden respiratory droplets has drawn a great deal of attention since the outbreak of COVID-19. Ambient relative humidity is the key factor controlling the evaporation of water from airborne droplets. The evaporation processes of respiratory droplets due to decreases in relative humidity are not well understood because of the complexity of the chemical compositions of the droplets. This article reviews the current understanding of the effects of relative humidity on the physicochemical and aerodynamic properties of respiratory droplets and on the infectivity of enveloped viruses. © 2021 The authors.

V367F Mutation in SARS-CoV-2 Spike RBD Emerging during the Early Transmission Phase Enhances Viral Infectivity through Increased Human ACE2 Receptor Binding Affinity.

The current pandemic of COVID-19 is caused by a novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The SARS-CoV-2 spike protein receptor-binding domain (RBD) is the critical determinant of viral tropism and infectivity. To investigate whether naturally occurring RBD mutations during the early transmission phase have altered the receptor binding affinity and infectivity, we first analyzed in silico the binding dynamics between SARS-CoV-2 RBD mutants and the human angiotensin-converting enzyme 2 (ACE2) receptor. Among 32,123 genomes of SARS-CoV-2 isolates (December 2019 through March 2020), 302 nonsynonymous RBD mutants were identified and clustered into 96 mutant types. The six dominant mutations were analyzed applying molecular dynamics simulations (MDS). The mutant type V367F continuously circulating worldwide displayed higher binding affinity to human ACE2 due to the enhanced structural stabilization of the RBD beta-sheet scaffold. The MDS also indicated that it would be difficult for bat SARS-like CoV to infect humans. However, the pangolin CoV is potentially infectious to humans. The increased infectivity of V367 mutants was further validated by performing receptor-ligand binding enzyme-linked immunosorbent assay (ELISA), surface plasmon resonance, and pseudotyped virus assays. Phylogenetic analysis of the genomes of V367F mutants showed that during the early transmission phase, most V367F mutants clustered more closely with the SARS-CoV-2 prototype strain than the dual-mutation variants (V367F+D614G), which may derivate from recombination. The analysis of critical RBD mutations provides further insights into the evolutionary trajectory of early SARS-CoV-2 variants of zoonotic origin under negative selection pressure and supports the continuing surveillance of spike mutations to aid in the development of new COVID-19 drugs and vaccines. IMPORTANCE A novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has caused the pandemic of COVID-19. The origin of SARS-CoV-2 was associated with zoonotic infections. The spike protein receptor-binding domain (RBD) is identified as the critical determinant of viral tropism and infectivity. Thus, whether mutations in the RBD of the circulating SARS-CoV-2 isolates have altered the receptor binding affinity and made them more infectious has been the research hot spot. Given that SARS-CoV-2 is a novel coronavirus, the significance of our research is in identifying and validating the RBD mutant types emerging during the early transmission phase and increasing human angiotensin-converting enzyme 2 (ACE2) receptor binding affinity and infectivity. Our study provides insights into the evolutionary trajectory of early SARS-CoV-2 variants of zoonotic origin. The continuing surveillance of RBD mutations with increased human ACE2 affinity in human or other animals is critical to the development of new COVID-19 drugs and vaccines against these variants during the sustained COVID-19 pandemic.

SARS-CoV-2 in hospital indoor environments is predominantly non-infectious.

BACKGROUND: The ongoing SARS-CoV-2 pandemic has spread rapidly worldwide and disease prevention is more important than ever. In the absence of a vaccine, knowledge of the transmission routes and risk areas of infection remain the most important existing tools to prevent further spread. METHODS: Here we investigated the presence of the SARS-CoV-2 virus in the hospital environment at the Uppsala University Hospital Infectious Disease ward by RT-qPCR and determined the infectivity of the detected virus in vitro on Vero E6 cells. RESULTS: SARS-CoV-2 RNA was detected in several areas, although attempts to infect Vero E6 cells with positive samples were unsuccessful. However, RNase A treatment of positive samples prior to RNA extraction did not degrade viral RNA, indicating the presence of SARS-CoV-2 nucleocapsids or complete virus particles protecting the RNA as opposed to free viral RNA. CONCLUSION: Our results show that even in places where a moderate concentration (Ct values between 30 and 38) of SARS-CoV-2 RNA was found; no infectious virus could be detected. This suggests that the SARS-CoV-2 virus in the hospital environment subsides in two states; as infectious and as non-infectious. Future work should investigate the reasons for the non-infectivity of SARS-CoV-2 virions.

An in vitro model for assessment of SARS-CoV-2 infectivity by defining the correlation between virus isolation and quantitative PCR value: isolation success of SARS-CoV-2 from oropharyngeal swabs correlates negatively with Cq value.

BACKGROUND: At the beginning of the pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), little was known about its actual rate of infectivity and any COVID-19 patient positive in laboratory testing was supposed to be highly infective and a public health risk factor. METHODS: One hundred oropharyngeal samples were obtained during routine work flow of testing symptomatic persons by quantitative polymerase chain reaction (qPCR) and were inoculated onto cell culture of VeroB4 cells to study the degree of infectivity of SARS-CoV-2 in vitro. Quantification by virus titration and an external standard using synthetic RNA gave the breaking point of infectivity in SARS-CoV-2 in vitro. RESULTS: A clear negative correlation (r = - 0.76; p < 0.05) could be asserted between the viral load in quantitative polymerase chain reaction (qPCR) and the probability of a successful isolation in serial isolation experiments of specific oropharyngeal samples positive in qPCR. Quantification by virus titration and an external standard using synthetic RNA indicate a Cq between 27 and 30 in E-gene screening PCR as a breaking point in vitro, where infectivity decreases significantly and isolations become less probable. CONCLUSIONS: This study showed that only the 21% of samples with the highest viral load were infectious enough to transmit the virus in vitro and determined that the dispersion rate in vitro is surprisingly close to those calculated in large retrospective epidemiological studies for SARS-CoV-2. This raises the question of whether this simple in vitro model is suitable to give first insights in dispersion characters of novel or neglected viral pathogens. The statement that SARS-CoV-2 needs at least 40,000 copies to reliably induce infection in vitro is an indication of its transmissibility in Public Health decisions. Applying quantitative PCR systems in diagnosis of SARS-CoV2 can distinguish between patients providing a high risk of transmission and those, where the risk of transmission is probably limited to close and long-lasting contacts.

Real-time PCR assay as a simple and efficient tool for viral stability study.

Aim: For oncolytic virus trials, regulatory agencies often require pharmaceutical industry to evaluate risks of released viruses from patients to environment. This study was to establish a real-time PCR method to assess viral shedding and viral stability in human urine. Results/methodology: Herein, we describe an incubation of viral drug product in human urine and use of real-time PCR as a simple, efficient and high throughput assay to assess the level and stability of a nonenveloped and single stranded RNA virus. The viral stability issue is critical to the collection, transport, storage and testing of clinical samples. Discussion/conclusion: In summary, this simple method provides useful viral stability information at various temperatures and detergents. A similar approach may apply to other RNA viruses (including SARS-CoV-2).

Retrospective study, full-length genome characterization and evaluation of viral infectivity and pathogenicity of chimeric porcine deltacoronavirus detected in Vietnam.

Increased evidence of porcine deltacoronavirus (PDCoV) causing diarrhoea in pigs has been reported in several countries worldwide. The virus has currently evolved into three separated groups including US, China and Southeast Asia (SEA) groups. In Vietnam, PDCoV was first reported in 2015. Based on phylogenetic analyses of spike, membrane and nucleocapsid genes, it is suggested that Vietnam PDCoV is chimeric virus. In the present study, we retrospectively investigated the presence of PDCoV in Vietnam and the full-length genomes of six PDCoV isolates identified in 2014-2016 were further characterized. The results demonstrated that Vietnam PDCoV was first detected as early as 2014. All six Vietnam PDCoV are in the SEA group and further divided into two separated subgroups including SEA-1 and SEA-2. Vietnam PDCoV in SEA-2 was closely related to Thai and Lao PDCoV. Recombination analysis demonstrated that three isolates in SEA-1 were a chimeric virus of which P12_14_VN_0814, the first Vietnam isolate, and US PDCoV isolates were major and minor parents, respectively. The recombination was further evaluated by phylogenetic construction based on 3 recombinant fragments. The first and third fragments, closely related to P12_14_VN_0814, were associated with ORF1a/1b and N genes, respectively. The second fragment, associated with S, E, and M genes, was closely related to US PDCoV isolates. High antigenic and hydrophobic variations were detected in S1 protein. Three-day-old pigs challenged with the chimeric virus displayed clinical diseases and villus atrophy. In conclusion, Vietnam PDCoV is genetically diverse influenced by an external introduction from neighbouring countries. The chimeric Vietnam PDCoV can induce a disease similar to Thai PDCoV.

The impact of thermal pasteurization on viral load and detectable live viruses in human milk and other matrices: a rapid review.

Holder pasteurization (62.5 °C, 30 min) of human milk is thought to reduce the risk of transmitting viruses to an infant. Some viruses may be secreted into milk - others may be contaminants. The effect of thermal pasteurization on viruses in human milk has yet to be rigorously reviewed. The objective of this study is to characterize the effect of common pasteurization techniques on viruses in human milk and non-human milk matrices. Databases (MEDLINE, Embase, Web of Science) were searched from inception to April 20th, 2020, for primary research articles assessing the impact of pasteurization on viral load or detection of live virus. Reviews were excluded, as were studies lacking quantitative measurements or those assessing pasteurization as a component of a larger process. Overall, of 65 131 reports identified, 109 studies were included. Pasteurization of human milk at a minimum temperature of 56-60 °C is effective at reducing detectable live virus. In cell culture media or plasma, coronaviruses (e.g., SARS-CoV, SARS-CoV-2, MERS-CoV) are highly susceptible to heating at ≥56 °C. Although pasteurization parameters and matrices reported vary, all viruses studied, except parvoviruses, were susceptible to thermal killing. Future research important for the study of novel viruses should standardize pasteurization protocols and should test inactivation in human milk. Novelty In all matrices, including human milk, pasteurization at 62.5 °C was generally sufficient to reduce surviving viral load by several logs or to below the limit of detection. Holder pasteurization (62.5 °C, 30 min) of human milk should be sufficient to inactivate nonheat resistant viruses, including coronaviruses, if present.

Mutations Strengthened SARS-CoV-2 Infectivity.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infectivity is a major concern in coronavirus disease 2019 (COVID-19) prevention and economic reopening. However, rigorous determination of SARS-CoV-2 infectivity is very difficult owing to its continuous evolution with over 10,000 single nucleotide polymorphisms (SNP) variants in many subtypes. We employ an algebraic topology-based machine learning model to quantitatively evaluate the binding free energy changes of SARS-CoV-2 spike glycoprotein (S protein) and host angiotensin-converting enzyme 2 receptor following mutations. We reveal that the SARS-CoV-2 virus becomes more infectious. Three out of six SARS-CoV-2 subtypes have become slightly more infectious, while the other three subtypes have significantly strengthened their infectivity. We also find that SARS-CoV-2 is slightly more infectious than SARS-CoV according to computed S protein-angiotensin-converting enzyme 2 binding free energy changes. Based on a systematic evaluation of all possible 3686 future mutations on the S protein receptor-binding domain, we show that most likely future mutations will make SARS-CoV-2 more infectious. Combining sequence alignment, probability analysis, and binding free energy calculation, we predict that a few residues on the receptor-binding motif, i.e., 452, 489, 500, 501, and 505, have high chances to mutate into significantly more infectious COVID-19 strains.

Factors determining the diffusion of COVID-19 and suggested strategy to prevent future accelerated viral infectivity similar to COVID.

This study has two goals. The first is to explain the geo-environmental determinants of the accelerated diffusion of COVID-19 that is generating a high level of deaths. The second is to suggest a strategy to cope with future epidemic threats similar to COVID-19 having an accelerated viral infectivity in society. Using data on sample of N = 55 Italian province capitals, and data of infected individuals at as of April 7th, 2020, results reveal that the accelerate and vast diffusion of COVID-19 in North Italy has a high association with air pollution of cities measured with days exceeding the limits set for PM10 (particulate matter 10 µm or less in diameter) or ozone. In particular, hinterland cities with average high number of days exceeding the limits set for PM10 (and also having a low wind speed) have a very high number of infected people on 7th April 2020 (arithmetic mean is about 2200 infected individuals, with average polluted days greater than 80 days per year), whereas coastal cities also having days exceeding the limits set for PM10 or ozone but with high wind speed have about 944.70 average infected individuals, with about 60 average polluted days per year; moreover, cities having more than 100 days of air pollution (exceeding the limits set for PM10), they have a very high average number of infected people (about 3350 infected individuals, 7th April 2020), whereas cities having less than 100 days of air pollution per year, they have a lower average number of infected people (about 1014 individuals). The findings here also suggest that to minimize the impact of future epidemics similar to COVID-19, the max number of days per year that Italian provincial capitals or similar industrialized cities can exceed the limits set for PM10 or for ozone, considering their meteorological conditions, is about 48 days. Moreover, results here reveal that the explanatory variable of air pollution in cities seems to be a more important predictor in the initial phase of diffusion of viral infectivity (on 17th March 2020, b1 = 1.27, p < 0.001) than interpersonal contacts (b2 = 0.31, p < 0.05). In the second phase of maturity of the transmission dynamics of COVID-19, air pollution reduces intensity (on 7th April 2020 with b'1 = 0.81, p < 0.001) also because of the indirect effect of lockdown, whereas regression coefficient of transmission based on interpersonal contacts has a stable level (b'2 = 0.31, p < 0.01). This result reveals that accelerated transmission dynamics of COVID-19 is due to mainly to the mechanism of "air pollution-to-human transmission" (airborne viral infectivity) rather than "human-to-human transmission". Overall, then, transmission dynamics of viral infectivity, such as COVID-19, is due to systemic causes: general factors that are the same for all regions (e.g., biological characteristics of virus, incubation period, etc.) and specific factors which are different for each region and/or city (e.g., complex interaction between air pollution, meteorological conditions and biological characteristics of viral infectivity) and health level of individuals (habits, immune system, age, sex, etc.). Lessons learned for COVID-19 in the case study here suggest that a proactive strategy to cope with future epidemics is also to apply especially an environmental and sustainable policy based on reduction of levels of air pollution mainly in hinterland and polluting cities- (having low wind speed, high percentage of moisture and number of fog days) -that seem to have an environment that foster a fast transmission dynamics of viral infectivity in society. Hence, in the presence of polluting industrialization in regions that can trigger the mechanism of air pollution-to-human transmission dynamics of viral infectivity, this study must conclude that a comprehensive strategy to prevent future epidemics similar to COVID-19 has to be also designed in environmental and socioeconomic terms, that is also based on sustainability science and environmental science, and not only in terms of biology, medicine, healthcare and health sector.