Viral Dose and Immunosuppression Modulate the Progression of Acute BVDV-1 Infection in Calves: Evidence of Long Term Persistence after Intra-Nasal Infection

Bovine viral diarrhoea virus (BVDV) infection of cattle causes a diverse range of clinical outcomes from being asymptomatic, or a transient mild disease, to producing severe cases of acute disease leading to death. Four groups of calves were challenged with a type 1 BVDV strain, originating from a severe outbreak of BVDV in England, to study the effect of viral dose and immunosuppression on the viral replication and transmission of BVDV. Three groups received increasing amounts of virus: Group A received 10^2.55TCID₅₀/ml, group B 10^5.25TCID₅₀/ml and group C 10^6.7TCID ₅₀/ml. A fourth group (D) was inoculated with a medium dose (10^5.25TCID₅₀/ml) and concomitantly treated with dexamethasone (DMS) to assess the effects of chemically induced immunosuppression. Naïve calves were added as sentinel animals to assess virus transmission. The outcome of infection was dose dependent with animals given a higher dose developing severe disease and more pronounced viral replication. Despite virus being shed by the low-dose infection group, BVD was not transmitted to sentinel calves. Administration of dexamethasone (DMS) resulted in more severe clinical signs, prolonged viraemia and virus shedding. Using PCR techniques, viral RNA was detected in blood, several weeks after the limit of infectious virus recovery. Finally, a recently developed strand-specific RT-PCR detected negative strand viral RNA, indicative of actively replicating virus, in blood samples from convalescent animals, as late as 85 days post inoculation. This detection of long term replicating virus may indicate the way in which the virus persists and/or is reintroduced within herds.     

Effect of Porcine Epidemic Diarrhea Virus Infectious Doses on Infection Outcomes in Na?ve Conventional Neonatal and Weaned Pigs

Porcine epidemic diarrhea virus (PEDV) was identified in the United States (U.S.) swine population for the first time in April 2013 and rapidly spread nationwide. However, no information has been published regarding the minimum infectious dose (MID) of PEDV in different pig models. The main objective of this study was to determine the oral minimum infectious dose of PEDV in naïve conventional neonatal piglets and weaned pigs. A U.S. virulent PEDV prototype isolate (USA/IN19338/2013) with known infectious titer was serially ten-fold diluted in virus-negative cell culture medium. Dilutions with theoretical infectious titers from 560 to 0.0056 TCID₅₀/ml together with a medium control were orogastrically inoculated (10ml/pig) into 7 groups of 5-day-old neonatal pigs (n = 4 per group) and 7 groups of 21-day-old weaned pigs (n = 6 per group). In 5-day-old pigs, 10ml of inoculum having titers 560–0.056 TCID₅₀/ml, corresponding to polymerase chain reaction (PCR) cycle threshold (Ct) values 24.2–37.6, resulted in 100% infection in each group; 10ml of inoculum with titer 0.0056 TCID₅₀/ml (Ct>45) caused infection in 25% of the inoculated pigs. In 21-day-old pigs, 10ml of inoculum with titers 560–5.6 TCID₅₀/ml (Ct 24.2–31.4) resulted in 100% infection in each group while 10ml of inoculum with titers 0.56–0.0056 TCID₅₀/ml (Ct values 35.3 –>45) did not establish infection in any pigs under study conditions as determined by clinical signs, PCR, histopathology, immunohistochemistry, and antibody response. These data reveal that PEDV infectious dose is age-dependent with a significantly lower MID for neonatal pigs compared to weaned pigs. This information should be taken into consideration when interpreting clinical relevance of PEDV PCR results and when designing a PEDV bioassay model. The observation of such a low MID in neonates also emphasizes the importance of strict biosecurity and thorough cleaning/disinfection on sow farms.     

The effect of climbing chalk powder on the infectivity of human coronavirus OC43

There does not appear to be any studies in the published literature on the stability of SARS-CoV-2 in climbing chalk powder (magnesium carbonate and/or calcium carbonate), which has been hypothesized to pose a potential risk of fomite transmission of coronavirus disease 2019 (COVID-19) within climbing gyms. The aim of this study was to determine the infectivity of a model human coronavirus HCoV-OC43 in the presence of climbing chalk powder on a dry plastic surface. The stability of HCoV-OC43 on a plastic surface dusted with climbing chalk powders (magnesium carbonate, calcium carbonate or a blended chalk) was determined by titration on BHK-21 fibroblast cells. No chalk and no virus controls were included. HCoV-OC43 was stable on the plastic surface for 48 h. The stability of HCoV-OC43 was significantly (P ≤ 0·05) reduced in the presence of magnesium carbonate, calcium carbonate and the chalk blend; the infectivity was reduced by ≥2·29 log₁₀ 50% tissue culture infective dose (TCID₅₀) immediately upon on contact and by ≥2·46 log₁₀ TCID₅₀ within 1 h of contact. These findings suggest that the infectivity of coronaviruses is reduced by climbing chalk, limiting the risk of potential fomite transmission.     

Tea crude extracts effectively inactivate severe acute respiratory syndrome coronavirus 2

It is well known that black and green tea extracts, particularly polyphenols, have antimicrobial activity against various pathogenic microbes including viruses. However, there is limited data on the antiviral activity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which emerged rapidly in China in late 2019 and which has been responsible for coronavirus disease 2019 (COVID-19) pandemic globally. In this study, 20 compounds and three extracts were obtained from black and green tea and found that three tea extracts showed significant antiviral activity against SARS-CoV-2, whereby the viral titre decreased about 5 logs TCID₅₀ per ml by 1·375 mg ml⁻¹ black tea extract and two-fold diluted tea bag infusion obtained from black tea when incubated at 25°C for 10 s. However, when concentrations of black and green tea extracts were equally adjusted to 344 µg ml⁻¹, green tea extracts showed more antiviral activity against SARS-CoV-2. This simple and highly respected beverage may be a cheap and widely acceptable means to reduce SARS-CoV-2 viral burden in the mouth and upper gastrointestinal and respiratory tracts in developed as well as developing countries.     

High-throughput,single-copy sequencing reveals SARS-CoV-2 spike variants coincident with mounting humoral immunity during acute COVID-19

Tracking evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) within infected individuals will help elucidate coronavirus disease 2019 (COVID-19) pathogenesis and inform use of antiviral interventions. In this study, we developed an approach for sequencing the region encoding the SARS-CoV-2 virion surface proteins from large numbers of individual virus RNA genomes per sample. We applied this approach to the WA-1 reference clinical isolate of SARS-CoV-2 passaged in vitro and to upper respiratory samples from 7 study participants with COVID-19. SARS-CoV-2 genomes from cell culture were diverse, including 18 haplotypes with non-synonymous mutations clustered in the spike NH₂-terminal domain (NTD) and furin cleavage site regions. By contrast, cross-sectional analysis of samples from participants with COVID-19 showed fewer virus variants, without structural clustering of mutations. However, longitudinal analysis in one individual revealed 4 virus haplotypes bearing 3 independent mutations in a spike NTD epitope targeted by autologous antibodies. These mutations arose coincident with a 6.2-fold rise in serum binding to spike and a transient increase in virus burden. We conclude that SARS-CoV-2 exhibits a capacity for rapid genetic adaptation that becomes detectable in vivo with the onset of humoral immunity, with the potential to contribute to delayed virologic clearance in the acute setting.     

A cohort autopsy study defines COVID-19 systemic pathogenesis

Severe COVID-19 disease caused by SARS-CoV-2 is frequently accompanied by dysfunction of the lungs and extrapulmonary organs. However, the organotropism of SARS-CoV-2 and the port of virus entry for systemic dissemination remain largely unknown. We profiled 26 COVID-19 autopsy cases from four cohorts in Wuhan, China, and determined the systemic distribution of SARS-CoV-2. SARS-CoV-2 was detected in the lungs and multiple extrapulmonary organs of critically ill COVID-19 patients up to 67 days after symptom onset. Based on organotropism and pathological features of the patients, COVID-19 was divided into viral intrapulmonary and systemic subtypes. In patients with systemic viral distribution, SARS-CoV-2 was detected in monocytes, macrophages, and vascular endothelia at blood–air barrier, blood–testis barrier, and filtration barrier. Critically ill patients with long disease duration showed decreased pulmonary cell proliferation, reduced viral RNA, and marked fibrosis in the lungs. Permanent SARS-CoV-2 presence and tissue injuries in the lungs and extrapulmonary organs suggest direct viral invasion as a mechanism of pathogenicity in critically ill patients. SARS-CoV-2 may hijack monocytes, macrophages, and vascular endothelia at physiological barriers as the ports of entry for systemic dissemination. Our study thus delineates systemic pathological features of SARS-CoV-2 infection, which sheds light on the development of novel COVID-19 treatment.Subject terms: Mechanisms of disease, Immunology     

Epidemiological,Clinical and Serological Characteristics of Children with Coronavirus Disease 2019 in Wuhan: A Single-centered,Retrospective Study

In December 2019, SARS-CoV-2 was first detected in the samples obtained from three adult patients who suffered from an unknown viral pneumonia in Wuhan (Li et al. 2020). This unknown viral pneumonia is further named as coronavirus disease 2019 (COVID-19) by the World Health Organization. To date, the number of new COVID-19 cases has continued to skyrocket and the impact of SARS-CoV-2 on humans is far greater than any pathogen of this century in both breadth and depth. Previous studies have shown that adults with COVID-19 have symptoms of fever, dry cough, dyspnea, fatigue and lymphocytopenia. Moreover, COVID-19 is more likely to cause death in the elderly, especially those with chronic comorbidities (Huang et al. 2020). In Wuhan, more than 50, 000 COVID-19 cases have been confirmed, including over 780 pediatric patients, and only one child death case (Lu et al. 2020). Although the number of children cases was far fewer than that of adults, COVID-19 might endanger children's health and the information on children remains limited, especially in serological study. In the retrospective study, the investigators analyzed the epidemiological, clinical and serological characteristics of children with COVID-19 in Wuhan in the early stages of the outbreak, which might provide theoretical and practical help in controlling COVID-19 and similar emerging infectious diseases in the future.     

Gut Microbiome Alterations in COVID-19

Since the outset of the coronavirus disease 2019 (COVID-19) pandemic, the gut microbiome in COVID-19 has garnered substantial interest, given its significant roles in human health and pathophysiology. Accumulating evidence is unveiling that the gut microbiome is broadly altered in COVID-19, including the bacterial microbiome, mycobiome, and virome. Overall, the gut microbial ecological network is significantly weakened and becomes sparse in patients with COVID-19, together with a decrease in gut microbiome diversity. Beyond the existence of severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), the gut microbiome of patients with COVID-19 is also characterized by enrichment of opportunistic bacteria, fungi, and eukaryotic viruses, which are also associated with disease severity and presentation. Meanwhile, a multitude of symbiotic bacteria and bacteriophages are decreased in abundance in patients with COVID-19. Such gut microbiome features persist in a significant subset of patients with COVID-19 even after disease resolution, coinciding with ‘long COVID’ (also known as post-acute sequelae of COVID-19). The broadly-altered gut microbiome is largely a consequence of SARS-CoV-2 infection and its downstream detrimental effects on the systemic host immunity and the gut milieu. The impaired host immunity and distorted gut microbial ecology, particularly loss of low-abundance beneficial bacteria and blooms of opportunistic fungi including Candida, may hinder the reassembly of the gut microbiome post COVID-19. Future investigation is necessary to fully understand the role of the gut microbiome in host immunity against SARS-CoV-2 infection, as well as the long-term effect of COVID-19 on the gut microbiome in relation to the host health after the pandemic.     

COVID-19, leprosy,and neutrophils

Coronavirus Disease 2019 (COVID-19), a disease caused by the betacoronavirus Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), has only recently emerged, while Mycobacterium leprae, the etiological agent of leprosy, has endured for more than 2,000 years. As soon as the initial reports of COVID-19 became public, several entities, including the Brazilian Leprosy Society, warned about the possible impact of COVID-19 on leprosy patients. It has been verified that COVID-19 carriers can be either asymptomatic or present varying degrees of severe respiratory failure in association with cytokine storm and death, among other diseases. Severe COVID-19 patients show increased numbers of neutrophils and serum neutrophil extracellular trap (NET) markers, in addition to alterations in the neutrophil-to-lymphocyte ratio (NLR). The absence of antiviral drugs and the speed of COVID-19 transmission have had a major impact on public health systems worldwide, leading to the almost total collapse of many national and local healthcare services. Leprosy, an infectious neurological and dermatological illness, is widely considered to be the most frequent cause of physical disabilities globally. The chronic clinical course of the disease may be interrupted by acute inflammatory episodes, named leprosy reactions. These serious immunological complications, characterized by cytokine storms, are responsible for amplifying peripheral nerve damage. From 30% to 40% of all multibacillary leprosy (MB) patients experience erythema nodosum leprosum (ENL), a neutrophilic immune-mediated condition. ENL patients often present these same COVID-19-like symptoms, including high levels of serum NET markers, altered NLR, and neutrophilia. Moreover, the consequences of a M. leprae–SARS-CoV-2 coinfection have yet to be fully investigated. The goal of the present viewpoint is to describe some of the similarities that may be found between COVID-19 and leprosy disease in the context of neutrophilic biology.     

Phenotypes and Functions of SARS-CoV-2-Reactive T Cells

Infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19), which is an ongoing pandemic disease. SARS-CoV-2-specific CD4⁺ and CD8⁺ T-cell responses have been detected and characterized not only in COVID-19 patients and convalescents, but also unexposed individuals. Here, we review the phenotypes and functions of SARS-CoV-2-specific T cells in COVID-19 patients and the relationships between SARS-CoV-2-specific T-cell responses and COVID-19 severity. In addition, we describe the phenotypes and functions of SARS-CoV-2-specific memory T cells after recovery from COVID-19 and discuss the presence of SARS-CoV-2-reactive T cells in unexposed individuals and SARS-CoV-2-specific T-cell responses elicited by COVID-19 vaccines. A better understanding of T-cell responses is important for effective control of the current COVID-19 pandemic.     

Vaccine effectiveness against SARS-CoV-2 infection,hospitalization, and death when combining a first dose ChAdOx1 vaccine with a subsequent mRNA vaccine in Denmark: A nationwide population-based cohort study

BackgroundThe recommendations in several countries to stop using the ChAdOx1 vaccine has led to vaccine programs combining different Coronavirus Disease 2019 (COVID-19) vaccine types, which necessitates knowledge on vaccine effectiveness (VE) of heterologous vaccine schedules. The aim of this Danish nationwide population-based cohort study was therefore to estimate the VE against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection and COVID-19–related hospitalization and death following the first dose of the ChAdOx1 vaccine and the combination of the ChAdOx1/mRNA vaccines.Methods and findingsAll individuals alive in or immigrating to Denmark from 9 February 2021 to 23 June 2021 were identified in the Danish Civil Registration System. Information on exposure, outcomes, and covariates was obtained from Danish national registries. Poisson and Cox regression models were used to calculate crude and adjusted VE, respectively, along with 95% confidence intervals (CIs) against SARS-CoV-2 infection and COVID-19–related hospitalization or death comparing vaccinated versus unvaccinated individuals. The VE estimates were adjusted for calendar time as underlying time and for sex, age, comorbidity, country of origin, and hospital admission. The analyses included 5,542,079 individuals (97.6% of the total Danish population). A total of 144,360 individuals were vaccinated with the ChAdOx1 vaccine as the first dose, and of these, 136,551 individuals received an mRNA vaccine as the second dose. A total of 1,691,464 person-years and 83,034 SARS-CoV-2 infections were included. The individuals vaccinated with the first dose of the ChAdOx1 vaccine dose had a median age of 45 years. The study population was characterized by an equal distribution of males and females; 6.7% and 9.2% originated from high-income and other countries, respectively. The VE against SARS-CoV-2 infection when combining the ChAdOx1 and an mRNA vaccine was 88% (95% CI: 83; 92) 14 days after the second dose and onwards. There were no COVID-19–related hospitalizations or deaths among the individuals vaccinated with the combined vaccine schedule during the study period. Study limitations including unmeasured confounders such as risk behavior and increasing overall vaccine coverage in the general population creating herd immunity are important to take into consideration when interpreting the results.ConclusionsIn this study, we observed a large reduction in the risk of SARS-CoV-2 infection when combining the ChAdOx1 and an mRNA vaccine, compared with unvaccinated individuals.

Mie Agermose Gram and co-workers study the effectiveness of heterologous SARS-CoV-2 vaccine combinations in the Danish population.     

新冠病毒中和单克隆抗体及纳米抗体研究进展北大核心CSCD

由严重急性呼吸系统综合征冠状病毒2型(severe acute respiratory syndrome coronavirus-2,SARS-CoV-2)引起的疾病被命名为新型冠状病毒肺炎(coronavirus disease 2019,COVID-19),是一种具有强传染性、高易感性、长潜伏期的传染病。病毒刺突蛋白受体结合结构域(receptor binding domain,RBD)和细胞血管紧张素转换酶2(angiotensin-converting enzyme 2,ACE2)之间的相互作用使得SARS-CoV-2顺利进入细胞。本文对SARS-CoV-2与ACE2的相关作用机制进行了简单概述,对目前针对SARS-CoV-2中和单克隆抗体、纳米抗体的最新研究进展进行了总结,探讨了新冠肺炎的发展过程和抗体药物的研究方向,以期为包括新冠肺炎在内的新发、突发传染病中和抗体药物的研发提供参考。     

Modeling the onset of symptoms of COVID-19: Effects of SARS-CoV-2 variant

Identifying order of symptom onset of infectious diseases might aid in differentiating symptomatic infections earlier in a population thereby enabling non-pharmaceutical interventions and reducing disease spread. Previously, we developed a mathematical model predicting the order of symptoms based on data from the initial outbreak of SARS-CoV-2 in China using symptom occurrence at diagnosis and found that the order of COVID-19 symptoms differed from that of other infectious diseases including influenza. Whether this order of COVID-19 symptoms holds in the USA under changing conditions is unclear. Here, we use modeling to predict the order of symptoms using data from both the initial outbreaks in China and in the USA. Whereas patients in China were more likely to have fever before cough and then nausea/vomiting before diarrhea, patients in the USA were more likely to have cough before fever and then diarrhea before nausea/vomiting. Given that the D614G SARS-CoV-2 variant that rapidly spread from Europe to predominate in the USA during the first wave of the outbreak was not present in the initial China outbreak, we hypothesized that this mutation might affect symptom order. Supporting this notion, we found that as SARS-CoV-2 in Japan shifted from the original Wuhan reference strain to the D614G variant, symptom order shifted to the USA pattern. Google Trends analyses supported these findings, while weather, age, and comorbidities did not affect our model’s predictions of symptom order. These findings indicate that symptom order can change with mutation in viral disease and raise the possibility that D614G variant is more transmissible because infected people are more likely to cough in public before being incapacitated with fever.     

Modeling insights into SARS-CoV-2 respiratory tract infections prior to immune protection

Mechanistic insights into human respiratory tract (RT) infections from SARS-CoV-2 can inform public awareness as well as guide medical prevention and treatment for COVID-19 disease. Yet the complexity of the RT and the inability to access diverse regions pose fundamental roadblocks to evaluation of potential mechanisms for the onset and progression of infection (and transmission). We present a model that incorporates detailed RT anatomy and physiology, including airway geometry, physical dimensions, thicknesses of airway surface liquids (ASLs), and mucus layer transport by cilia. The model further incorporates SARS-CoV-2 diffusivity in ASLs and best-known data for epithelial cell infection probabilities, and, once infected, duration of eclipse and replication phases, and replication rate of infectious virions. We apply this baseline model in the absence of immune protection to explore immediate, short-term outcomes from novel SARS-CoV-2 depositions onto the air-ASL interface. For each RT location, we compute probability to clear versus infect; per infected cell, we compute dynamics of viral load and cell infection. Results reveal that nasal infections are highly likely within 1–2 days from minimal exposure, and alveolar pneumonia occurs only if infectious virions are deposited directly into alveolar ducts and sacs, not via retrograde propagation to the deep lung. Furthermore, to infect just 1% of the 140 m² of alveolar surface area within 1 week, either 10³ boluses each with 10⁶ infectious virions or 10⁶ aerosols with one infectious virion, all physically separated, must be directly deposited. These results strongly suggest that COVID-19 disease occurs in stages: a nasal/upper RT infection, followed by self-transmission of infection to the deep lung. Two mechanisms of self-transmission are persistent aspiration of infected nasal boluses that drain to the deep lung and repeated rupture of nasal aerosols from infected mucosal membranes by speaking, singing, or cheering that are partially inhaled, exhaled, and re-inhaled, to the deep lung.     

On the Calculation of TCID50 for Quantitation of Virus Infectivity

正Dear Editor The most important property of a virus is its infectivity. To measure infectivity, one can assay viral replication in cells to obtain a titer for a given virus stock. A titer is defined as a given number of infectious viral units per unit volume,and an infectious unit is the smallest amount of virus that produces recognizable effects [e.g., cytopathic effect(CPE), dot blot immunoreactivity]. The median tissue culture infectious dose (TCID_(50)) is defined as the dilution of a virus required to infect 50%of a given cell culture.     

Severe Acute Respiratory Syndrome Coronavirus 2: Manifestations of Disease and Approaches to Treatment and Prevention in Humans

The coronavirus disease 2019 (COVID-19) pandemic was caused by a novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). This virus has challenged civilization and modern science in ways that few infectious diseases and natural disasters have previously, causing globally significant human morbidity and mortality and triggering economic downturns across financial markets that will be dealt with for generations. Despite this, the pandemic has also brought an opportunity for humanity to come together and participate in a shared scientific investigation. Clinically, SARS-CoV-2 is associated with lower mortality rates than other recently emerged coronaviruses, such as SARS-CoV and the Middle East respiratory syndrome coronavirus (MERS-CoV). However, SARS-CoV-2 exhibits efficient human-to-human spread, with transmission often occurring before symptom recognition; this feature averts containment strategies that had worked previously for SARS-CoV and MERS-CoV. Severe COVID-19 disease is characterized by dysregulated inflammatory responses associated with pulmonary congestion and intravascular coagulopathy leading to pneumonia, vascular insults, and multiorgan disease. Approaches to treatment have combined supportive care with antivirals, such as remdesivir, with immunomodulatory medications, including corticosteroids and cytokine-blocking antibody therapies; these treatments have advanced rapidly through clinical trials. Innovative approaches to vaccine development have facilitated rapid advances in design, testing, and distribution. Much remains to be learned about SARS-CoV-2 and COVID-19, and further biomedical research is necessary, including comparative medicine studies in animal models. This overview of COVID-19 in humans will highlight important aspects of disease, relevant pathophysiology, underlying immunology, and therapeutics that have been developed to date.

In December 2019, a cluster of cases of pneumonia without a clear etiology occurred in Wuhan, China. With remarkable speed and efficiency, the etiology of this illness was soon identified as a novel coronavirus; the complete viral genome was sequenced and published on January 10, 2020.¹⁸² These events introduced the world to coronavirus disease 2019 (COVID-19). The disease, now known to be caused by a novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has developed into the most significant pandemic of recent times. In less than a year since the virus was first recognized, multiple candidate vaccines were developed worldwide, and some of them rapidly progressed to clinical trials and widespread administration. As the pandemic continues, a number of sequence variants of the virus have emerged around the world. This continued viral evolution highlights the need for continued biomedical research to facilitate understanding of the pathogenesis of COVID-19, seeking innovative therapeutic and preventative strategies for the current and possibly future pandemics. This article will review aspects of SARS-CoV-2 infection of humans and COVID-19, focusing on important aspects of clinical disease, pathophysiology, immunology, and the development of therapeutic and preventative measures to provide context for discussion of the animal models used to study SARS-CoV-2 and COVID-19.