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SARS Immunity and Vaccination
Abstract
Severe acute respiratory syndrome (SARS) is a serious and fatal infectious disease caused by SARS coronavirus (SARS-Cov), a novel human coronavirus. SARS-Cov infection stimulates cytokines (e.g., IL-10, IFN-gamma, IL-1, etc.) expression dramatically, and T lymphocytes and their subsets CD4(+) and CD8(+) T cells are decreased after onset of the disease. SARS-specific IgG antibody is generated in the second week and persists for a long time, whereas IgM is expressed transiently. The spike protein and neucleocapsid protein are most abundant in SARS-Cov and contribute dominantly to the antibody production during the course of disease. Spike protein, especially the ACE-2 binding region (318-510aa) is capable of producing neutralizing antibody to SARS-Cov. Neucleocapsid protein induces protective specific CTL to SARS-Cov. Therefore, applications with spike subunit, neucleocapsid subunit as well as inactivated SARS-Cov are three prospective vaccination strategies for SARS.
... [ 12] Four main sub-groupings of coronaviruses, alpha, beta, gamma, and delta, are known . [13] The viruses that cause the most serious infections are severe acute respiratory syndrome (SARS), which was caused by SARS-CoV in 2002, [14,15], Middle East Respiratory Syndrome (MERS), which was caused by MERS-CoV in 2012. [15] and SARS-CoV-2 [17,18] (which causes the disease COVID-19) In Figure.1 a schematic representation of the genome organization and functional domains of S protein for SARS-CoV, SARS-CoV-2 and MERS-CoV is shown. ...
... [13] The viruses that cause the most serious infections are severe acute respiratory syndrome (SARS), which was caused by SARS-CoV in 2002, [14,15], Middle East Respiratory Syndrome (MERS), which was caused by MERS-CoV in 2012. [15] and SARS-CoV-2 [17,18] (which causes the disease COVID-19) In Figure.1 a schematic representation of the genome organization and functional domains of S protein for SARS-CoV, SARS-CoV-2 and MERS-CoV is shown. The structural genes encode the structural proteins, spike (S), envelope (E), membrane (M), and nucleocapsid (N), which are common features to all coronaviruses, [14 ] Another important SARS-COV-2 protein is the viral 3chymotrypsin-like cysteine protease (3CLpro), also termed M-Pro, is a polypeptide of 306 amino acids, which plays a key role in the replication of coronavirus. ...
In this study, our approach was to conduct a complete investigation of molecular docking analysis with the major SARS-CoV-2 proteins and SARS CoV-1 proteins. We analyzed more than 6000 drugs, downloaded by the Pubchem database. Particular attention, we have focused on 3CLpro Covid-19 protein, by "Blind docking" method and "Selective docking" procedure, in Ligand Binding site, with Autodock Vina using Pyrx, a small Virtual Screen Library, and with Autodock 4 using AMDock Software. We have selected about 30 drugs with a high-affinity Binding score (between-9.7 and-12 kcal/mol) from our results. Next, carrying out docking Validation methods, using Human Serum Albumin, as a docking control protein, has led us to select only the three best drugs (Ubrogepant, Atogepant, and Paritaprevir), potentially active in the Ligand Binding site pocket of 3CL-pro SARS-CoV-2 protein. Ubrogepant and Atogepant, have Ki values around, 20-25 nM and Binding Energy of approximately-10 kcal/mol, except for Paritaprevir which reports exceptional Ki values of approximately 1nM and Binding Energy of approximately-12.5 kcal/mol. This has led us to conclude, that they could be excellent candidates against SARS-COV-2, even though further in vitro and in vivo studies are needed.
... 16 Levels of acute-response cytokines -TNF Alpha, IL-6, ferritin, C reactive protein (CRP), High Sensitive C reactive protein (hsCRP) and pro-coagulant factors D-Dimer, Prothrombin time (PT) and activated Partial thromboplastin time (aPTT), many of them measurable are elevated. 17 Cytokine storm leads to a critical life-threatening condition requiring intensive care admission and having a quite high mortality in many conditions including viral infections, cancer, sepsis, rheumatic diseases and systemic juvenile idiopathic arthritis. 18 Lung injury is also one of the consequences of the cytokine storm that can progress into acute lung injury or it is a more severe form Acute Respiratory Distress Syndrome (ARDS). ...
... 18 Lung injury is also one of the consequences of the cytokine storm that can progress into acute lung injury or it is a more severe form Acute Respiratory Distress Syndrome (ARDS). 17 Cytokine storm has been reported in several viral infections including influenza H5N1 virus, influenza H1N1 virus, and the two corona viruses highly related to COVID-19; severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East Respiratory Syndrome coronavirus (MERS-CoV). ...
Severe Acute Respiratory Syndrome (SARS) novel coronavirus 2 (nCoV2) has been declared as a pandemic
by the WHO and it is implicated in varying degree of mortality and morbidity among different nations.
Several studies have shown that susceptibility to cytokine storm is responsible for differences in mortality
and morbidity. Interleukin-6 (IL-6) has been implicated strongly for the cause of cytokine storm in
COVID-19 infections. But it is unclear about the cause for the severity of COVID-19. Recent studies have
shown that bradykinins, one of the peptides are also implicated in storm leading to severe complications among
the infected. Hence, this review article is focused on role of cytokines and bradykinins in causing storms in
COVID-19.
Keywords: Bradykinin, COVID -19, Cytokine storm, Inflammation, Interleukin - 6
... 16 Levels of acute-response cytokines -TNF Alpha, IL-6, ferritin, C reactive protein (CRP), High Sensitive C reactive protein (hsCRP) and pro-coagulant factors D-Dimer, Prothrombin time (PT) and activated Partial thromboplastin time (aPTT), many of them measurable are elevated. 17 Cytokine storm leads to a critical life-threatening condition requiring intensive care admission and having a quite high mortality in many conditions including viral infections, cancer, sepsis, rheumatic diseases and systemic juvenile idiopathic arthritis. 18 Lung injury is also one of the consequences of the cytokine storm that can progress into acute lung injury or it is a more severe form Acute Respiratory Distress Syndrome (ARDS). ...
... 18 Lung injury is also one of the consequences of the cytokine storm that can progress into acute lung injury or it is a more severe form Acute Respiratory Distress Syndrome (ARDS). 17 Cytokine storm has been reported in several viral infections including influenza H5N1 virus, influenza H1N1 virus, and the two corona viruses highly related to COVID-19; severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East Respiratory Syndrome coronavirus (MERS-CoV). ...
Abstract Severe Acute Respiratory Syndrome (SARS) novel coronavirus 2 (nCoV2) has been declared as a pandemic by the WHO and it is implicated in varying degree of mortality and morbidity among different nations. Several studies have shown that susceptibility to cytokine storm is responsible for differences in mortality and morbidity. Interleukin-6 (IL-6) has been implicated strongly for the cause of cytokine storm in COVID-19 infections. But it is unclear about the cause for the severity of COVID-19. Recent studies have shown that bradykinins, one of the peptides are also implicated in storm leading to severe complications among the infected. Hence, this review article is focused on role of cytokines and bradykinins in causing storms in COVID-19. Keywords: Bradykinin, COVID -19, Cytokine storm, Inflammation, Interleukin - 6
... T helper cells (CD4 + ) are simulated by antigens and differentiate into different subtypes of T cells to participate in cell-mediated and/or humoral immunity, while CD8 + cytotoxic T lymphocytes play a pivotal role in recognition of endogenous antigen peptides and elimination of infected cells [35]. Our study showed the immunological changes in COVID-19 patients with DM, who presented a higher percentage of CD4 + , but a lower percentage of CD8 + than those in nondiabetic patients. ...
... But how the T helper cells and suppressor T cells act in the pathogenesis of COVID-19 needs more researches to elucidate. Activated CD4 + T cells secrete either inflammatory cytokines (TNF-α, INF-γ, IL-6, and IL-2) or cytokines with antiinflammatory cytokines (Th2: IL-4, IL-10) [35]. The imbalance of the immune system leads to inflammation, which is a response of an organism to virus infection. ...
The induction of inflammation and cytokine storm was proposed to play a critical role in COVID-19. This study is aimed at investigating the relationship between glucose metabolism and the inflammatory state of inpatients with COVID-19. 71 inpatients with COVID-19 were classified into nondiabetes mellitus (NDM) group, impaired fasting glucose (IFG) group, and diabetes mellitus (DM) group. The average hospitalization days were significantly shorter in DM patients when compared with patients in the IFG group and NDM group. CD4+ T cell percentage was higher while CD8+ T cells percentage was lower in the DM group than those in the NDM group. The serum levels of IL-6, IL-2, IL-10, and INF-γ in the DM group were upregulated when compared with those in the NDM group. The serum levels of TNF-α, IL-4, IL-2, IL-10, and INF-γ were significantly higher in the DM group than those in the IFG group. A significant difference was observed in CD4+ T cell, CD4+/CD8+ ratio percentage, IL-6, and IL-10 between the NDM group and DM group with adjusted BMI. In conclusion, COVID-19 patients with elevated glucose levels have promoted cytokine profiles and immune response.
... Those >10% patients who had no IgG antibodies after 21 days post symptom onset most likely lost those IgG antibodies after the infection was resolved.Why antibodies to SARS-CoV-2 in infected persons fade so quickly unlike antibodies to SARS-CoV-1? After infection with SARS-CoV-1, patients start to produce SARS--speci c IgG antibody in the second week, which persists for a long time36,37 . Even after 210 days after symptom onset, neutralizing viral antibodies (anti-viral IgG) are still detectable in recovered SARS patients38 . ...
... Even after 210 days after symptom onset, neutralizing viral antibodies (anti-viral IgG) are still detectable in recovered SARS patients38 . It is believed that the Spike protein and nucleocapsid protein play a central role in the antibody production36,37,39 . Research has focused on developing vaccines and therapeutics targeting the Spike protein40,41 and the nucleocapsid protein42 . ...
Background It is to be determined whether people infected with SARS-CoV-2 will develop long-term immunity against SARS-CoV-2 and retain long-lasting antibodies after the infection is resolved. This study was to explore the outcomes of IgG antibodies to SARS-CoV-2 in four groups of individuals in Wuhan, China.Methods
We conducted a cross-sectional study on the following four groups who received both COVID-19 IgM/IgG tests and RT-PCR tests for SARS-CoV-2 from February 29, 2020 to April 29, 2020: 1470 hospitalized patients with COVID-19 from Leishenshan Hospital, Zhongnan Hospital of Wuhan University, and Wuhan No. 7 Hospital, 3832 healthcare providers without COVID-19 diagnosis, 19555 general workers, and 1616 other patients to be admitted to the hospital (N=26473). COVID-19 patients who received IgM/IgG tests <21 days after symptom onset were excluded. Results IgG prevalence was 89.8% (95% CI 88.2-91.3%) in COVID-19 patients, 4.0% (95% CI 3.4-4.7%) in healthcare providers, 4.6 (95% CI 4.3-4.9 %) in general workers, and 1.0% in other patients (p all <0.001 for comparisons with COVID-19 patients). IgG prevalence increased significantly by age among healthcare workers and general workers. Among hospitalized COVID-19 patients, presence of IgG antibodies to SARS-CoV-2 was not associated with most disease severity, presence of comorbidities, treatment received, and clinical characteristics. We identified 24 hospitalized patients with COVID-19 and multiple COVID-19 antibody tests who lost previously detected IgG antibodies to SARS-CoV-2Conclusions
IgG antibodies to SARS-CoV-2 in infected people may become undetectable overtime.
... This rises rapidly to 92.3% between days 11 and 24, and the IgG component of the test offers a sensitivity of 98.6% during this phase of the infection or even longer [13]. IgG antibodies are the main antibodies produced as an immune response, indicating that the disease has entered a recovery period or that there is a prior infection [14,15]. Therefore, combined tests of immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies can not only provide the diagnosis of infectious diseases but also help to evaluate the stages of infection in the body [12,13]. ...
... However, a low viral load in patients' throat and the limitation of RT-PCR result in a significant number of false-negative reports, which should not be ignored [25]. In general, the coronavirus stimulates the immune response and IgM antibodies are produced after 4-24 days upon infection and then quickly decline until disappear, while on the other hand, IgG antibodies are usually produced after IgM and continue to rise and remain high in the body for long periods [14,15]. For treatment monitoring and status of the disease, the decrease or even disappearance of the concentration of IgM and the increase in the concentration of IgG indicates the severity of the patient and the immunity to the pathogenicity of SARS-CoV-2. ...
Background
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused coronavirus disease 2019 (COVID-19) epidemic in China, December 2019. The clinical features and treatment of COVID-19 patients remain largely elusive. However, accurate detection is required for SARS-CoV-2 infection diagnosis. We aimed to evaluate the antibodies-based test and nucleic acid-based test for SARS-CoV-2-infected patients.
Methods
We retrospectively studied 133 patients diagnosed with SARS-CoV-2 and admitted to Renmin Hospital of Wuhan University, China, from January 23 to March 1, 2020. Demographic data, clinical records, laboratory tests, and outcomes were collected. Data were accessed by SARS-CoV-2 IgM-IgG antibody test and real-time reverse transcriptase PCR (RT-PCR) detection for SARS-CoV-2 nucleic acid in COVID-19 patients.
Results
Of 133 COVID-19 patients, there were 44 moderate cases, 52 severe cases, and 37 critical cases with no differences in gender and age among three subgroups. In RT-PCR detection, the positive rate was 65.9%, 71.2%, and 67.6% in moderate, severe, and critical cases, respectively. Whereas the positive rate of IgM/IgG antibody detection in patients was 79.5%/93.2%, 82.7%/100%, and 73.0%/97.3% in moderate, severe, and critical cases, respectively. Moreover, the IgM and IgG antibodies concentrations were also examined with no differences among three subgroups.
Conclusion
The IgM-IgG antibody test exhibited a useful adjunct to RT-PCR detection, and improved the accuracy in COVID-19 diagnosis regardless of the severity of illness, which provides an effective complement to the false-negative results from a nucleic acid test for SARS-CoV-2 infection diagnosis after onsets.
... Противовирусный иммунный ответ на коронавирусную инфекцию стал изучаться многими исследовательскими группами после вспышки SARS-CoV в 2002 году [10,11,12]. Нейтрализующие антитела и Т-клеточный ответ, главным образом, были направлены против S-белка, который участвует в распознавании рецептора и проникновении вируса в клетку-мишень и играет важную роль в адаптивной эволюции вируса [4]. ...
The pandemic of coronavirus infection (COVID-19) has stimulated the development, testing and widespread use of preventive vaccines based on various platforms. Our aim was to perform a direct comparison of immunogenicity of various vaccines within a single study in small groups within six months of SARS-CoV-2 vaccination and revaccination. The stdy group included subjects vaccinated with Sputnik V adenovirus vaccine, mRNA vaccines, and CoviVac whole-virion vaccine. Their immune status was assessed by enzyme immunoassay as specific antibody levels. Moreover, the neutralizing ability of detected antibodies was assessed using a cell test system based on pseudoviral technology. All of the mentioned vaccines were shown to elicit an immune response against SARS-CoV-2 RBD antigen, however, appropriate antibody titers and neutralizing capacities differed depending on the type of vaccine. The mRNA vaccines proved to be the most immunogenic, the effectiveness of the immune response to the Sputnik V adenovirus-based vaccine was lower. However, 6 months after vaccination, the effectiveness of virus neutralizing antibodies induced by these vaccines did not differ. The whole-virion CoviVac vaccine with proven efficiency by independent epidemiological studies, induced an antibody response against the RBD protein to a lesser extent. The seropositive participants of the study, both previously exposed to COVID-19 disease or vaccinated, exhibited high-titer production of antibodies already after the first dose of the Sputnik V vaccine, and a significantly higher antibody titer 6 months after the booster immunization as compared with initial level of antibodies, along with direct correlation between the antibody titers and their neutralizing activity.
... As with other pathogenic human coronaviruses, the most established correlate of protection for SARS-CoV-2 is virus-specific neutralizing antibodies (Khoury et al., 2021). Albeit with distinct kinetics, naturally acquired immunity to severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East respiratory syndrome coronavirus (MERS-CoV) generates antigen-specific and virus neutralizing antibodies that can last more than 18 months (Zhu, 2004;Liu et al., 2006;Alshukairi et al., 2016;Payne et al., 2016;Mubarak et al., 2019;Memish et al., 2020). Due to the short duration of the SARS-CoV outbreak, and the lack of observational cohort studies to assess MERS-CoV reinfection, questions remain as to whether these long-lasting antibodies protect against reinfection (Memish et al., 2020;Al-Tawfiq et al., 2021). ...
The ongoing evolution of SARS-CoV-2 continues to raise new questions regarding the duration of immunity to reinfection with emerging variants. To address these knowledge gaps, controlled investigations in established animal models are needed to assess duration of immunity induced by each SARS-CoV-2 lineage and precisely evaluate the extent of cross-reactivity and cross-protection afforded. Using the Syrian hamster model, we specifically investigated duration of infection acquired immunity to SARS-CoV-2 ancestral Wuhan strain over 12 months. Plasma spike- and RBD-specific IgG titers against ancestral SARS-CoV-2 peaked at 4 months post-infection and showed a modest decline by 12 months. Similar kinetics were observed with plasma virus neutralizing antibody titers which peaked at 2 months post-infection and showed a modest decline by 12 months. Reinfection with ancestral SARS-CoV-2 at regular intervals demonstrated that prior infection provides long-lasting immunity as hamsters were protected against severe disease when rechallenged at 2, 4, 6, and 12 months after primary infection, and this coincided with the induction of high virus neutralizing antibody titers. Cross-neutralizing antibody titers against the B.1.617.2 variant (Delta) progressively waned in blood over 12 months, however, re-infection boosted these titers to levels equivalent to ancestral SARS-CoV-2. Conversely, cross-neutralizing antibodies to the BA.1 variant (Omicron) were virtually undetectable at all time-points after primary infection and were only detected following reinfection at 6 and 12 months. Collectively, these data demonstrate that infection with ancestral SARS-CoV-2 strains generates antibody responses that continue to evolve long after resolution of infection with distinct kinetics and emergence of cross-reactive and cross-neutralizing antibodies to Delta and Omicron variants and their specific spike antigens.
... Thanks to past experiences in vaccine design and manufacturing, the pipeline for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was rapidly established to tackle this outbreak situation [5,6]. Lessons learned from severe acute respiratory syndrome coronavirus (SARS-CoV) [7], Middle East respiratory syndrome coronavirus (MERS-CoV) [8], and influenza virus [9], etc. made researchers design vaccines empirically, which shorted trial and error time while an effective SARS-CoV-2 vaccine was urgently important. Additionally, the pandemic boosted the development of vaccine technologies, and an increasing number of novel strategies were exploited to meet the urgent need for effective vaccines. ...
Vaccination is the most cost-effective means in the fight against infectious diseases. Various kinds of vaccines have been developed since the outbreak of COVID-19, some of which have been approved for clinical application. Though vaccines available achieved partial success in protecting vaccinated subjects from infection or hospitalization, numerous efforts are still needed to end the global pandemic, especially in the case of emerging new variants. Safe and efficient vaccines are the key elements to stop the pandemic from attacking the world now; novel and evolving vaccine technologies are urged in the course of fighting (re)-emerging infectious diseases. Advances in biotechnology offered the progress of vaccinology in the past few years, and lots of innovative approaches have been applied to the vaccine design during the ongoing pandemic. In this review, we summarize the state-of-the-art vaccine strategies involved in controlling the transmission of SARS-CoV-2 and its variants. In addition, challenges and future directions for rational vaccine design are discussed.
... 2 Very high levels of INF-g, IL-1b, IL-6, IL-8, IL-12, and TNF-a cytokines were found in SARS patients who developed ARDS. [3][4][5] High mortality rates associated with epidemics of SARS (9.6%) and Middle East respiratory syndrome (34.4%) posed a major global threat in recent past. 6 Till date, globally, 22 June 2021, there have been 178,360,849 confirmed cases of COVID-19, including 3,869,384 deaths, reported to WHO. ...
Background: COVID-19 is a highly contagious viral infection resulting in severe acute respiratory syndrome Coronavirus (SARS-COV-2). The objective of this study is to determine the risk factors affecting the outcome of moderate to severe COVID 19 patients admitted in HDU of Sir Ganga Ram Hospital, Lahore. Study Design: Prospective cohort study. Place and Duration of Study: COVID HDU of Sir Ganga Ram Hospital, Lahore from 1st June 2020 to 30th November 2020. Methodology: One hundred and eight patients with positive PCR for COVID having moderate to severe COVID disease were enrolled. After written consent, all the demographic and clinical information was obtained. All the patients were followed till their discharge or death. Results: Sixty eight were discharged, 26 were died and14 got left against medical advice. The median age of all patients was53.6 years and the median age of patients who died was significantly older than those who survived. Conclusion: The potential risk factors for high mortality were older age, female sex, those presented with within 1 week from onset of symptoms that need ventilation at admission and comorbidities like Diabetes, Hypertension, and ischemic heart disease. This study could help clinicians to identify patients with poor prognosis at an early stage. Keywords: Covid-19, Outcome, HDU admission criteria
... IgM is mainly produced in the primary immune response to infectious factors or antigens, as IgG mainly produced in the secondary. Studies have shown that SARS specific antibodies are produced in the second week of infection and last for a long time, while IgM is transiently expression (27). IgM is an antibody in the acute phase, which generally appears 3-5 days after the onset of the disease, remains positive for about one month, and then gradually decreases. ...
Objective
To analyze the macular structure of age-related cataract (ARC) patients with different antibody levels after COVID-19 vaccine injection, in order to obtain the effect of COVID-19 vaccine on the macular structure, and speculate whether the COVID-19 vaccine has adverse effects on the macular structure.
Methods
This retrospective study is conducted to analysis on the status of COVID-19 vaccine and the thickness of different layers at different positions in the macular area of ARC patients. In the age, sex and eye axial length matched population, in the un-injection, no-antibody, IgM and IgG positive groups after vaccination, the choroid, ganglion cell complex, nerve fiber layer and retinal thickness at different positions of ETDRS zoning in the macular area were discussed.
Results
A total of 164 patients (164 eyes) were included in the analysis. There were 63 males and 101 females. The average age was 65.99 ± 8.43 years. There was no significant difference in age and sex among the groups (p>0.05). The average axial length of 164 eyes was 23.56 ± 1.46mm, and no significant difference between the groups (p>0.05). Non parametric test and ANOVA test for the thickness of choroid, retina, ganglion cell complex and retinal nerve fiber layer in each division of ETDRS showed no significant difference in the four groups of un-injection, no-antibody, IgM and IgG (p>0.05). There was no correlation between the antibody concentration and the thickness of macular structure (p>0.05).
Conclusion
There was no significant difference in the thickness of choroid, retina, ganglion cell complex and retinal fiber layer in different macular areas after COVID-19 vaccine injection. There was no linear correlation between the thickness of choroid, retina, ganglion cell complex and retinal fiber layer and the antibody concentration produced after COVID-19 vaccine injection. It suggests that the injection of COVID-19 vaccine might have no significant effect on the macular structure of eye.
... The titers of SARS-CoV-2 antibody measured indicates that most antibodies generated against S protein and N protein belonged to subtype IgG; there were only low levels of antibodies with subtype IgM (21). This is not surprising; IgG antibody is typically produced for several weeks and lasts for a long time (more than 13 weeks), whereas IgM antibody is transiently expressed (22). This pattern was reproduced here: specific-IgG antibody responses peaked at 28 days after the second vaccine dose and decreased to a low level by 6 months post-vaccination. ...
Background
Owing to the coronavirus disease 2019 (COVID-19) pandemic and the emergency use of different types of COVID-19 vaccines, there is an urgent need to consider the effectiveness and persistence of different COVID-19 vaccines.
Methods
We investigated the immunogenicity of CoronaVac and Covilo, two inactivated vaccines against COVID-19 that each contain inactivated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The levels of neutralizing antibodies to live SARS-CoV-2 and the inhibition rates of neutralizing antibodies to pseudovirus, as well as the immunoglobulin (Ig)G and IgM responses towards the spike (S) and nucleocapsid (N) protein of SARS-CoV-2 at 180 days after two-dose vaccination were detected.
Results
The CoronaVac and Covilo vaccines induced similar antibody responses. Regarding neutralizing antibodies to live SARS-CoV-2, 77.9% of the CoronaVac vaccine recipients and 78.3% of the Covilo vaccine recipients (aged 18–59 years) seroconverted by 28 days after the second vaccine dose. Regarding SARS-CoV-2-specific antibodies, 97.1% of the CoronaVac vaccine recipients and 95.7% of the Covilo vaccine recipients seroconverted by 28 days after the second vaccine dose. The inhibition rates of neutralizing antibody against a pseudovirus of the SARS-CoV-2 Delta variant were significantly lower compared with those against a pseudovirus of wildtype SARS-CoV-2. Associated with participant characteristics and antibody levels, persons in the older age group and with basic disease, especially a chronic respiratory disease, tended to have lower anti-SARS-CoV-2 antibody seroconversion rates.
Conclusion
Antibodies that were elicited by these two inactivated COVID-19 vaccines appeared to wane following their peak after the second vaccine dose, but they persisted at detectable levels through 6 months after the second vaccine dose, and the effectiveness of these antibodies against the Delta variant of SARS-CoV-2 was lower than their effectiveness against wildtype SARS-CoV-2, which suggests that attention must be paid to the protective effectiveness, and its persistence, of COVID-19 vaccines on SARS-CoV-2 variants.
... Importantly, S protein is dominantly contributed to SARS- [59] proved that the constructed recombinant forms of S-DNA gene vaccine may elicit protective immunity by a high antibody production anti-SARS-CoV-1-S protein target for vaccination [59]. Recent data demonstrated that ACE-2 is SARS-CoV-1receptor and its ligand is located at region of S1/S2, results indicated that this region contributed to neutralizing antibodies production [57]. Therefore, it is proposed that ACE-2 binding region of SARS-CoV-1spike fragment might contain two antigenic epitopes for neutralizing antibody production and these epitopes might be useful for SARS-CoV-1 vaccination. ...
... Furthermore, anti-S antibody titers as measured by antibody capture ELISA indicates that a majority of antibodies against the S protein is of the IgG subtype, with lower levels of the IgA and IgM subtypes. This is not surprising, as individuals recovering from SARS-CoV-1 have been shown to have strong IgG and IgM antibody responses with IgM titers spiking shortly after infection and steadily decreasing thereafter and IgG titers spiking and then remaining at higher concentrations for upwards of 13 weeks [49]. The role of these isotypes in neutralization have also been extensively studied, with several high-affinity IgG antibodies isolated from acutely infected patients [31,33,[35][36][37][38]. ...
The novel coronavirus, SARS-CoV-2 that causes COVID-19 has resulted in the death of nearly 4 million people within the last 18 months. While preventive vaccination, and monoclonal antibody therapies have been rapidly developed and deployed, early in the pandemic the use of COVID-19 convalescent plasma (CCP) was a common means of passive immunization with a theoretical risk of antibody-dependent enhancement (ADE) of viral infection. Though vaccines elicit a strong and protective immune response and transfusion of CCP with high titers of neutralization activity are correlated with better clinical outcomes, the question of whether antibodies in CCP can enhance infection of SARS-CoV-2 has not been directly addressed. In this study, we analyzed for and observed passive transfer of neutralization activity with CCP transfusion. Furthermore, to specifically understand if antibodies against the spike protein (S) enhance infection, we measured the anti-S IgG, IgA, and IgM responses and adapted retroviral-pseudotypes to measure virus neutralization with target cells expressing the ACE2 virus receptor and the Fc alpha receptor (FcαR) or Fc gamma receptor IIA (FcγRIIA). Whereas neutralizing activity of CCP correlated best with higher titers of anti-S IgG antibodies, the neutralizing titer was not affected when Fc receptors were present on target cells. These observations support the absence of antibody-dependent enhancement of infection (ADE) by IgG and IgA isotypes found in CCP. The results presented, therefore, not only supports the therapeutic use of currently available antibody-based treatment, including the continuation of CCP transfusion strategies, but also the use of various vaccine platforms in a prophylactic approach.
... Importantly, S protein is dominantly contributed to SARS- [59] proved that the constructed recombinant forms of S-DNA gene vaccine may elicit protective immunity by a high antibody production anti-SARS-CoV-1-S protein target for vaccination [59]. Recent data demonstrated that ACE-2 is SARS-CoV-1receptor and its ligand is located at region of S1/S2, results indicated that this region contributed to neutralizing antibodies production [57]. Therefore, it is proposed that ACE-2 binding region of SARS-CoV-1spike fragment might contain two antigenic epitopes for neutralizing antibody production and these epitopes might be useful for SARS-CoV-1 vaccination. ...
COVID-19 is a new public health crisis caused by the novel respiratory pathogen SARS-CoV-2. It is one of the most significant pandemic events in recent history. The SARS-CoV-2 Beta corona virus was transmitted to humans in the end of 2019 by unknown intermediary host from bats in Wuhan, Hubei province (China). It marked the third major coronavirus source of disaster in the 21stcentury.The three last severe respiratory tract infections caused by the SARS-CoV-1, MERS-CoV and SARS-CoV-2 caused high human mortality. Viral genomic sequencing and investigations and the development of advanced vaccine strategies are expected to give us more information on these emerging pathogens and controlling them in the future. The aim of this review is to summarize updated information regarding these emerging human coronaviruses to understand their molecular and structural biology, transmissions and potential vaccine approaches actually developed against the SARS-CoV-2.
... Previous studies have shown that marked elevation of proinflammatory cytokines, such as IL-1, TNF-α, IL-6, TGF-β, and IL-10 were correlated with pulmonary inflammation and severe lung impairment in MERS-CoV patients (20). SARS-CoV infection was also reported to induce ARDS associated with highly expressed proinflammatory cytokines, such as IL-1β, IL-6, IL-8, IL-12, INF-γ, and TNF-α (21)(22)(23). ...
Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection may trigger a cytokine storm, which is characterized by uncontrolled overproduction of proinflammatory cytokines. Objectives: We aimed to investigate the association between circulating levels of inflammatory cytokines and severity of coronavirus disease 2019 (COVID-19). Methods: This cross-sectional study included 46 severe and 32 mildly symptomatic COVID-19 patients. The serum levels of cytokines and chemokines were determined using the Bio-Plex ProTM Human Cytokine Screening Panel. Results: Out of a total of 78 patients with confirmed COVID-19, 54 (69.2%) were males, and 24 (30.8%) were females. The mean age was 43.1 ± 13.3 and 58.2 ± 15 in mild and severe patients, respectively. Severe patients were characterized by significant laboratory abnormalities, such as increased WBC (P = 0.002) and neutrophil counts (P = 0.001), higher levels of ALT (P = 0.03), AST (P = 0.002), LDH (P < 0.001), urea (P = 0.013), ferritin (P < 0.001), D-dimer (P = 0.042), CRP (P < 0.001), and decreased lymphocyte (P < 0.001) and platelet (P = 0.045) counts. The levels of IL-6, IL-8, IL-13, TNF-α, IFN-γ, MIP-1β, and MCP-1 increased in the severe group compared to the mild group. However, significant differences were observed only for IL-6 (P < 0.001) and IL-8 (P < 0.001) levels. Conclusions: Serum IL-6 and IL-8 levels can be used as potential prognostic biomarkers of disease severity in COVID-19 patients.
... However, under the situation of the COVID-19 outbreak, the medical community is facing tremendous pressure to rapidly develop effective vaccines (261). In past pandemics such as those involving Ebola, H1N1, SARS, and MERS, vaccine development was unable to be completed owing to the end of the pandemic and the reallocation of scientific funds (262)(263)(264)(265). Since July 2 nd , 2020, there has been 158 vaccine candidates for COVID-19, 135 of which are in the preclinical or the developing stage. ...
The coronaviruses that cause notable diseases, namely, severe acute respiratory syndrome (SARS), middle east respiratory syndrome (MERS) and coronavirus disease 2019 (COVID-19), exhibit remarkable similarities in genomic components and pathogenetic mechanisms. Although coronaviruses have widely been studied as respiratory tract pathogens, their effects on the hepatobiliary system have seldom been reported. Overall, the manifestations of liver injury caused by coronaviruses typically involve decreased albumin and elevated aminotransferase and bilirubin levels. Several pathophysiological hypotheses have been proposed, including direct damage, immune-mediated injury, ischemia and hypoxia, thrombosis and drug hepatotoxicity. The interaction between pre-existing liver disease and coronavirus infection has been illustrated, whereby coronaviruses influence the occurrence, severity, prognosis and treatment of liver diseases. Drugs and vaccines used for treating and preventing coronavirus infection also have hepatotoxicity. Currently, the establishment of optimized therapy for coronavirus infection and liver disease comorbidity is of significance, warranting further safety tests, animal trials and clinical trials.
... Furthermore, anti-S antibody titers, as measured by antibody capture ELISA, indicate that a majority of antibodies against the S protein are of the IgG subtype, with lower levels of the IgA and IgM subtypes. This is not surprising, as individuals recovering from SARS-CoV-1 have been shown to have strong IgG and IgM antibody responses, with IgM titers spiking shortly after infection and steadily decreasing thereafter and IgG titers spiking and then remaining at higher concentrations for upwards of 13 weeks [49]. The role of these isotypes in neutralization have also been extensively studied, with several high-affinity IgG antibodies isolated from acutely infected patients [31,33,[36][37][38][39]. ...
The novel coronavirus SARS-CoV2, which causes COVID-19, has resulted in the death of nearly 4 million people within the last 18 months. While preventive vaccination and monoclonal antibody therapies have been rapidly developed and deployed, early in the pandemic the use of COVID-19 convalescent plasma (CCP) was a common means of passive immunization, with the theoretical risk of antibody-dependent enhancement (ADE) of viral infection remaining undetermined. Though vaccines elicit a strong and protective immune response, and transfusion of CCP with high titers of neutralization activity are correlated with better clinical outcomes, the question of whether antibodies in CCP can enhance infection of SARS-CoV2 has not been directly addressed. In this study, we analyzed for and observed passive transfer of neutralization activity with CCP transfusion. Furthermore, to specifically understand if antibodies against the spike protein (S) enhance infection, we measured the anti-S IgG, IgA, and IgM responses and adapted retroviral-pseudotypes to measure virus neutralization with target cells expressing the ACE2 virus receptor and the Fc alpha receptor (FcaR) or Fc gamma receptor IIA (FcgRIIA). Whereas neutralizing activity of CCP correlated best with higher titers of anti-S IgG antibodies, the neutralizing titer was not affected when Fc receptors were present on target cells. These observations support the absence of antibody-dependent enhancement of infection (ADE) by IgG and IgA isotypes found in CCP. The results presented, therefore, support the clinical use of currently available antibody-based treatment including the continued study of CCP transfusion strategies.
... High NLR are commonly presented and associated with more severe viral infection. 23,24 Recent data indicated that the NLR, an indicator of innate immunity, was identified as a powerful predictive and prognostic factor for severe COVID-19. 18 Previous data showed severe SARS was associated with more robust serological responses including early seroconversion and higher IgG levels. ...
Purpose
The coronavirus disease 2019 (COVID-19) has quickly became a global threat to public health, and it is difficult to predict severe patients and their prognosis. Here we intend to develop effective models for late identification of patients at disease progression and outcome.
Methods
A total of 197 patients were included with 20-day median follow-up time. We first developed a nomogram for disease severity discrimination, then created a prognostic nomogram for severe patients.
Results
40.6% of patients were severe and 59.4% were non-severe. The multivariate logistic analysis indicated that IgG, neutrophil-to-lymphocyte ratio (NLR), lactate dehydrogenase, platelet, albumin, and blood urea nitrogen were significant factors associated with the severity of COVID-19. Using immune response phenotyping based on NLR and IgG level, the logistic model showed patients with NLRhiIgGhi phenotype are most likely to have severe disease, especially compared to those with NLRloIgGlo phenotype. The C-indices of the two discriminative nomogram was 0.86 and 0.87 respectively, which indicated sufficient discriminative power. As for predicting clinical outcome for severe patients, IgG, NLR, age, lactate dehydrogenase, platelet, monocytes, and procalcitonin were significant predictors. The prognosis of severe patients with NLRhiIgGhi phenotype were significantly worse than NLRloIgGhi group. The two prognostic nomograms also showed good performance in estimating the risk of progression.
Conclusion
The present nomogram models are useful to identify COVID-19 patients with progression based on individual characteristics and immune response-related indicators. Patients at high-risk for severe illness and poor outcome from COVID-19 should be managed with intensive supportive care and appropriate therapeutic strategies.
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... In this study, patients with SARS-CoV-2 infection developed a weak innate immune-cell ability to produce cytokines, such as IFN-γ, and showed decreased CD4 + and CD8 + T cells. SARS-CoV infection increases cytokine expression (e.g., IFN-γ, IL-1, IL-6, IL-10, IL-12, and IL-16) dramatically, and T lymphocytes and their CD4 + and CD8 + T cells subsets are decreased after the onset of infection [36]. In contrast, HIV-infected patients showed increased mean absolute CD3 + T-cell numbers and absolute CD8 + T-cell numbers [37]. ...
To simultaneously determine clinical and immunological responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in young and old females and males, 681 coronavirus disease 2019 (COVID-19) patients and 369 normal controls (NCs) were analyzed based on age and sex classifications using multiple linear regression analysis. Compared to the age-matched NCs, both young and old male and female non-comorbid COVID-19 patients had lower lymphocyte counts and alanine aminotransferase (ALT) concentration, and only young male and female patients had lower neutrophil counts. Compared to young patients, both old males and females had significantly higher plasma ALT and AST concentrations. Compared to young and old females, age-matched males had higher plasma ALT and AST concentrations, but only young males had higher C-reactive protein (CRP) concentration. Compared to females, old males, but not young males, showed higher incidence of critical illness. Compared to young patients, old females had more leukocyte and neutrophil counts above the normal upper limit and B cell count below the normal lower limit (NLL), while old males had more lymphocyte and natural killer (NK) cell counts below the NLL. No sex or age associations with B cell and NK cell counts were observed. However, there were age-dependent decreases in CD8+ T-cell counts in both male and female COVID-19 patients. Age was negatively associated with CD8+ T cell counts but positively associated with neutrophil count, CRP, ALT, and AST concentrations, and sex (females) was negatively associated with neutrophil count, CRP, ALT, and AST concentrations. The present study suggests that SARS-CoV-2 infection mainly induced 1) beneficial sex (female)-related differences regarding reduced COVID-19 disease severity and negative associations with inflammatory responses and liver damage, and 2) harmful age-related differences relating to negative associations with CD8+ T cell count and positive associations with inflammatory responses and liver damage. Thus, sex and age are biological variables that should be considered in the prevention and treatment of COVID-19.
... Its clinical manifestations include persistent fever, headache, muscle soreness, and decreased white blood cell count; severe cases could experience acute respiratory distress syndrome (ARDS) and multiple organ failure 9 . SARS-CoV infection stimulates cytokines dramatically, including IL-10, and decreases T lymphocytes and their subsets CD4 + and CD8 + T cells after disease onset 10 . It has been previously revealed the effect of GCS on pulmonary oxygenation function in the treatment of SARS. ...
Objective:
Coronavirus disease 2019 (COVID-19) has elevated mortality in severe and critical patients globally. This study examined the effect of glucocorticoids (GCS) on the time of virus clearance and absorption of lung lesions in severe and critical COVID-19 patients.
Patients and methods:
Severe and critical COVID-19 cases diagnosed in Wuhan Pulmonary Hospital from January 7 to February 10, 2020 were analyzed. The generalized linear model was utilized to assess the effects of GCS therapy on the times of nucleic acid test turning negative and improved pulmonary imaging, respectively.
Results:
Of 66 patients, 51 (77.3%) and 15 (22.7%) were severe and critical cases, respectively, and aged 62 ± 11 years. A total of 58 patients (87.9%) tested negative, and 56 (84.8%) showed improved lung imaging. Age, thrombocytopenia, CD8 + T cell count, course of GCS therapy, and total dose were correlated with the time of nucleic acid test turning negative (p < 0.05), and sex was correlated with the time of initial pulmonary imaging improvement (p < 0.05). The time of nucleic acid test turning negative in individuals with GCS therapy course ≤ 10 days was shorter than that of the GCS therapy course > 10 days group (p=0.001). No statistical difference was found in the dose, course of GCS, and initial time of improved lung imaging.
Conclusions:
Increasing the dose of GCS and prolonging the course of treatment do not shorten the time of nucleic acid test turning negative or improved absorption of pulmonary lesions. Thus, the rational use of GCS is particularly important.
... Coronaviruses are similar to the influenza virus, due to its contagion and its high transmissibility and they have triggered epidemics, such as SARS-CoV in 2002-2004 [5] and the MERS-CoV in 2015 [2,6]. The experience of these previous epidemics shows that a modification in behavior to adopt protective measures is required, such as the use of masks, washing hands, and isolation, principally among the affected populations [7]. ...
Objective:
To investigate the association among social media exposure, risk perception, preventive behaviors, and attitudes toward the COVID-19 epidemic in Bolivia.
Methods:
We launched an online survey in La Paz and El Alto, Bolivia, during April and May 2020. The questionnaire examined: Socio-demographic factors, Social media use, Risk Perception, Preventive behaviors, attitudes and the willingness to use a vaccine if it were available in the context of the COVID-19 epidemic. A logistic regression was used to evaluate factors associated with risk perception and a structural equation model (SEM) was performed to explore the pathway of the relationship among social media exposure, risk perception and preventive behaviors and attitudes.
Results:
Among 886 participants, the most were young adults, between 18-25 years old (73.4%) and 577 (65.1%) were female. During the the week before the survey 387 (43.7%) reported be exposure to social media Covid-19 information almost always or always. Moreover 304 (34.3%) were categorized as with a high risk perception. The multivariable analyses show that being female (aOR = 1.5, CI 95% 1.1-2.1) and having high exposure to Covid-19 information on social media (aOR = 2.5, CI 95% 1.3-5.3) were associated with a higher risk perception for Covid-19. Furthermore, SEM results indicated that risk perception is associated with the adoption of preventive behaviors and attitudes (β = 0.605, p < 0.001) including the acceptance of a vaccine if one were available (β = 0.388, p < 0.001).
Conclusion:
Social media exposure to COVID-19 information influences the adoption of preventive attitudes and behaviors through shaping risk perception. Understanding the role of social media during the pandemic could help policymakers and communicators to develop better communication strategies that enable the population to adopt appropriate attitudes and behaviors.
... ACE2 is a surface metalloproteinase that is encoded in the brain, liver, kidney, spleen, respiratory system, and small intestine [127][128][129]. Studies showed that ACE2 is used as an entry receptor by SARS-CoV and SAR-CoV-2 and other related viruses [38,112,113,128,130,131]. Comparing the genomic and protein structural features of human ACE2 [132] and the resulting complex of human ACE2 with SARS S protein [133] revealed that 17 out 19 human ACE2 codons correlate to residues in both structures. ...
One year after its first outbreak reported in China, coronavirus disease 2019 (COVID-19) pandemic is still sweeping the World causing serious infections and claiming more fatalities. COVID-19 is caused by the novel corona virus SARS-CoV-2, which belongs to the genus Betacoronavirus (β-CoVs) which is of greatest clinical importance since it contains many other viruses that cause respiratory disease in humans including OC43, HKU1, SARS-CoV and MERS. The spike (S) glycoprotein of β-CoVs is a key virulence factor determining disease pathogenesis and host tropism, and it also mediates virus binding to host’s receptors to allow viral entry into host cells, i.e., the first step in virus lifecycle. This, viral entry inhibitors are considered promising putative drugs for COVID-19. Herein, we mined the biomedical literature for viral entry inhibitors of other corona viruses, with special emphasis on β-CoVs entry inhibitors. We also outlined the structural features of SARS-CoV-2 S protein and how it differs from other β-CoVs to better understand the structural determinants of S protein binding to its human receptor (ACE2). This review highlighted several promising viral entry inhibitors as potential treatments for COVID-19.
... In response to the SARS-CoV outbreak in 2003, efforts targeting the ACE2−spike interaction emphasized antibodybased therapeutics. 21,22 Phage display libraries 10,11 and surface plasmon resonance 23 were used to develop neutralizing peptides. However, reported small-molecule screens for SARS-CoV at the time focused on target-agnostic cellular approaches. ...
The COVID-19 pandemic, caused by SARS-CoV-2, is a pressing public health emergency garnering a rapid response from scientists across the globe. Host cell invasion is initiated through direct binding of the viral spike protein to the host receptor angiotensin-converting enzyme 2 (ACE2). Disrupting the spike protein–ACE2 interaction is a potential therapeutic target for treating COVID-19. We have developed a proximity-based AlphaLISA assay to measure the binding of SARS-CoV-2 spike protein receptor binding domain (RBD) to ACE2. Utilizing this assay platform, a drug-repurposing screen against 3384 small-molecule drugs and preclinical compounds was carried out, yielding 25 high-quality primary hits, of which only corilagin was validated in cherry-picking. This established AlphaLISA RBD–ACE2 platform can facilitate evaluation of biologics or small molecules that can perturb this essential viral–host interaction to further the development of interventions to address the global health pandemic.
... Sin embargo, los estudios nos indicaron su mayor utilidad después del día 7 de infección (8) ; por lo cual, hoy sabemos que la presencia de IgM pudiera prolongarse y la IgG empezar a disminuir a partir del tercer mes (9) . La epidemia previa en el 2003 al SARS-CoV mostró la persistencia de IgM e IgG hasta 13 semanas posinfección, sugiriendo una respuesta inmune humoral contra el SARS (10) . Asimismo, se sabe que los anticuerpos de otros tipos de coronavirus van disminuyendo con el tiempo y han durado entre 12 y 52 meses (11) . ...
La infección por el virus SARS-CoV-2 que nos tiene en medio de una pandemia que cambió el curso de la humanidad, no solo afectó y consumió la vida de muchos pacientes, sino que también se encargóde destruir y quitar la vida principalmente a los responsables de defenderla: personal médico, de enfermería y salud en general. Cuando nos formamos en la escuela de medicina, lo hicimos por convicción y pensando siempre en proteger la salud y la vida de los demás, pero nunca lo hicimos pensando en sacrifcar nuestrapropia vida. Esta pandemia nos ha demostrado que, a pesar de los grandes adelantos en medicina, la era de los antibióticos, la biología molecular, la capacidad de crear vida en el laboratorio, los billones de dólares gastados en tecnología médica y la inmensa cantidad de tinta empleada en publicar artículos en revistas de ciencias médicas, aún no estamos preparados para enfrentar a un virus nuevo e invisible al ojo humano, pero que ya acabó con la vida de más gente que en las dos bombas atómicas de Hiroshima y Nagasaki
... MERS-CoV strains with mutations in the NSP3 and ORF4a display differential STAT3 activation and different inflammatory cytokine profiles (Selinger et al., 2014). In SARS-CoV infection, a reduction of IL-4 is observed, which participates in humoral protection, an increase of IFN-γ, that participates in a potent cell-mediated immune response and also the elevation of IL-10 that plays a part in disease susceptibility (Zhu, 2004). ...
SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2), as well as SARS-CoV from 2003 along with
MERS-CoV from 2012, is a member of the Betacoronavirus genus of the Nidovirales order and is currently the cause
of the pandemic called COVID-19 (or Coronavirus disease 2019). COVID-19, which is characterized by cough,
fever, fatigue, and severe cases of pneumonia, has affected more than 23 million people worldwide until August
25th, 2020. Here, we present a review of the cellular mechanisms associated with human coronavirus replication,
including the unique molecular events related to the replication transcription complex (RTC) of coronaviruses. We
also present information regarding the interactions between each viral protein and cellular proteins associated
to known host-pathogen implications for the coronavirus biology. Finally, a specific topic addresses the current
attempts for pharmacological interventions against COVID-19, highlighting the possible effects of each drug on the
molecular events of viral replication. This review intends to aid future studies for a better understanding of the SARSCoV-
2 replication cycle and the development of pharmacological approaches targeting COVID-19.
... 33 Previous reports demonstrated that the CoV N protein induced protective specific CTLs. [34][35][36][37] Moreover, NAbs titers significantly correlated with the number of N proteinspecific T cells, suggesting that the production of NAbs might be linked with the activation of antiviral T cells. 28,38 Another study reported that antisera to M proteins exhibited high neutralizing titers toward SARS-CoV infection, indicative of the importance of M protein for developing an effective protein-based vaccine. ...
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an emerging virus that is highly pathogenic and has caused the recent worldwide pandemic officially named coronavirus disease (COVID-19). Currently, considerable efforts have been put into developing effective and safe drugs and vaccines against SARS-CoV-2. Vaccines, such as inactivated vaccines, nucleic acid-based vaccines, and vector vaccines, have already entered clinical trials. In this review, we provide an overview of the experimental and clinical data obtained from recent SARS-CoV-2 vaccines trials, and highlight certain potential safety issues that require consideration when developing vaccines. Furthermore, we summarize several strategies utilized in the development of vaccines against other infectious viruses, such as severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East respiratory syndrome coronavirus (MERS-CoV), with the aim of aiding in the design of effective therapeutic approaches against SARS-CoV-2.
... However, false-negative results can occur in tests based on antibody detection. The IgM antibodies are produced as part of the early immune response during the initial stage of the infection, while the IgG antibodies indicate that the disease has entered a recovery period, or may be present if there has been prior infection (89,90). The antibody tests are used in cases where RT-PCR is negative and there is an epidemiological bond to SARS-CoV-2 infection and during the period when symptons are first presented and the viral load is high (91). ...
The emergence of a new coronavirus (SARS-CoV-2) outbreak represents a challenge for the diagnostic laboratories responsible for developing test kits to identify those infected with SARS-CoV-2. Methods with rapid and accurate detection are essential to control the sources of infection, to prevent the spread of the disease and to assist decision-making by public health managers. Currently, there is a wide variety of tests available with different detection methodologies, levels of specificity and sensitivity, detection time, and with an extensive range of prices. This review therefore aimed to conduct a patent search in relation to tests for the detection of SARS-CoV, MERS-CoV, and SARS-CoV-2. The greatest number of patents identified in the search were registered between 2003 and 2011, being mainly deposited by China, the Republic of Korea, and the United States. Most of the patents used the existing RT-PCR, ELISA, and isothermal amplification methods to develop simple, sensitive, precise, easy to use, low-cost tests that reduced false-negative or false-positive results. The findings of this patent search show that an increasing number of materials and diagnostic tests for the coronavirus are being produced to identify infected individuals and combat the growth of the current pandemic; however, there is still a question in relation to the reliability of the results of these tests.
... Severe morbidity and mortality associated with SARS make it crucial to develop a safe and successful vaccine to prevent reemergence and spread of disease (36). It is vital to develop protective immune responses, including neutralization antibody and cytotoxic T lymphocytes generation (66). ...
Human coronavirus infections have been known to cause mild respiratory illness. It changed in the last two decades as three global outbreaks by coronaviruses led to significant mortality and morbidity. SARS CoV-1 led to the first epidemic of the 21st century due to coronavirus. SARS COV-1 infection had a broad array of symptoms with respiratory and gastrointestinal as most frequent. The last known case was reported in 2004. Middle East respiratory syndrome coronavirus (MERS-CoV) led to the second outbreak in 2012, and case fatality was much higher than SARS. MERS-CoV has a wide array of clinical presentations from mild, moderate to severe, and some patients end up with acute respiratory distress syndrome (ARDS). The third and recent outbreak by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) started in December 2019, which lead to a global pandemic. Patients with SARS-CoV2 infection can be asymptomatic or have a range of symptoms with fever, cough, and shortness of breath being most common. Reverse transcriptase-Polymerase chain reaction (RT-PCR) is a diagnostic test of choice for SARS CoV-1, MERS-CoV, and SARS CoV-2 infections.
This review aims to discuss epidemiological, clinical features, diagnosis, and management of human coronaviruses with a focus on SARS CoV-1, MERS-CoV, and SARS CoV-2 infections.
... A previous study [11] described that IgG and IgM can be positive before the clinical diagnosis, and that this may provide a quick, simple and accurate aided detection method for suspected COVID-19 patients, when compared to RNA detection. In a previous study in SARS, the level of IgG and IgM antibodies increased to detectable levels at the third week of illness [12]. Therefore, the investigators speculated that a similar phenomenon may occur in the course of the SARS-CoV-2 infection. ...
COVID-19 is caused by SARS-CoV-2 infection and was initially discovered in Wuhan. This outbreak quickly spread all over China and then to more than 20 other countries. SARS-CoV-2 fluorescent microsphere immunochromatographic test strips were prepared by the combination of time-resolved fluorescence immunoassay with a lateral flow assay. The analytical performance and clinical evaluation of this testing method was done and the clinical significance of the testing method was verified. The LLOD of SARS-CoV-2 antibody IgG and IgM was 0.121U/L and 0.366U/L. The specificity of IgM and IgG strips in healthy people and in patients with non-COVID-19 disease was 94%, 96.72% and 95.50%, 99.49%, respectively; and sensitivity of IgM and IgG strips for patients during treatment and follow-up was 63.02%, 37.61% and 87.28%, 90.17%, respectively. The SARS-CoV-2 antibody test strip can provide rapid, flexible and accurate testing, and is able to meet the clinical requirement for rapid on-site testing of virus. The ability to detect IgM and IgG provided a significant benefit for the detection and prediction of clinical course with COVID-19 patients.
... 8,13 Lymphopenia occurred within the first two weeks of illness in SARS patients. 18 Likewise, in severe cases with middle east respiratory syndrome (MERS), which was induced by another coronavirus (MERS-CoV), serum levels of cytokine and chemokine also elevated in the early two weeks. 19 In the present study, most of the ALT elevations were begin within 1-3 weeks after symptoms onset. ...
Background
The profiles of liver function abnormalities in COVID‐19 patients need to be clarified.
Methods
In this retrospective study, consecutive COVID‐19 patients over 60 years old in Renmin Hospital of Wuhan University from Jan 1 to Feb 6 were included. Data of demographics, clinical characteristics, comorbidities, laboratory tests, medications and outcomes were collected and analyzed. Sequential alterations of serum alanine aminotransferase (ALT) were monitored.
Results
330 patients were included and classified into two groups with normal (n=234) or elevated ALT (n=96). There were fewer females (40.6% vs 54.7%, P =0.020) and more critical cases (30.2% vs. 19.2%, P =0.026) in patients with elevated ALT compared to the normal group. Higher levels of bacterial infection indices (e.g. white blood cell count, neutrophil count, C‐reactive protein and procalcitonin) were observed in the elevated group. Spearman correlation showed that both ALT and AST levels were positively correlated with those indices of bacterial infection. No obvious effects of medications on ALT abnormalities were found. In patients with elevated ALT, most ALT elevations were mild and transient. 59.4% of the patients had ALT concentrations of 41–100 U/L, while only a few patients (5.2%) had high serum ALT concentrations above 300 U/L. ALT elevations occurred at 13 (10–17) days and recovered at 28 (18–35) days from disease onset. For most patients, the elevation of serum ALT levels occurred at 6–20 days after disease onset and reached their peak values within a similar time frame. The recovery of serum ALT levels to normal frequently occurred at 16–20 days or 31–35 days after disease onset.
Conclusions
Liver function abnormalities were observed in 29.1% of the elderly COVID‐19 patients, which were slightly and transient in most cases. Liver function abnormalities in COVID‐19 may be correlated with bacterial infection.
... Although the patient did not have a contact history with other COVID-19 patients after discharge, the possibility of reinfection could not be completely ruled out during his stay in triple room of isolation ward. Insufficient production of protective antibodies might be responsible for reinfection [11] and for failure to clear the virus completely in this patient. We tracked the antibody levels of 5 patients with good recovery of the disease (detected on day 24 to day 37 after symptom onset) and found that levels of IgM and IgG were higher than that of this patient, especially IgG levels were all higher than 100 AU/ml (supplementary Table 1). ...
Background:
The rapid spread of coronavirus disease 2019 (COVID-19) was declared as an emerging public health threat by the World Health Organization. As various measures have been taken successfully to combat the epidemic caused by SARS-CoV-2, a growing number of fully recovered patients have been discharged from hospitals. However, some of them have relapsed. Little is known about the causes that triggered the relapse.
Case presentation:
We report a case of a 40 years old man who suffered from recurrent pulmonary infection with progression of lesions on chest computed tomography (CT), elevated levels of ferritin and IL2R, reduced lymphocyte count and positive oropharyngeal swab test for SARS-CoV-2 again after 5 days discharge from hospital. The anti-SARS-CoV-2 antibody level of this patient was very low at the time of relapse, suggesting a weak humoral immune response to the virus. Total exon sequencing revealed mutations in TRNT1 gene, which may be responsible for B cell immunodeficiency. Therefore, uncleared SARS-CoV-2 at his first discharge was likely to lead to his recurrence. However, viral superinfection and non-infectious organizing pneumonia could not be completely excluded.
Conclusion:
COVID-19 relapse may occur in a part of discharged patients with low titers of anti-SARS-CoV-2 antibodies. These patients should be maintained in isolation for longer time even after discharge. A more sensitive method to detect SARS-CoV-2 needs to be established and serological testing for specific antibodies may be used as a reference to determine the duration of isolation.
... Lymphopenia, neutrophilia, and high NLR are commonly presented and associated with more severe viral infection (Zhu, 2004;Liu et al., 2020b;Wang et al., 2020). Recent data indicated that the NLR was identified as a powerful predictive and prognostic factor for severe COVID-19 (Liu et al., 2020a). ...
Introduction: A recently emerging respiratory disease named coronavirus disease 2019 (COVID-19) has quickly spread across the world. This disease is initiated by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and uncontrolled cytokine storm, but it remains unknown as to whether a robust antibody response is related to clinical deterioration and poor outcome in COVID-19 patients. Methods: Anti-SARS-CoV-2 IgG and IgM antibodies were determined by chemiluminescence analysis (CLIA) in COVID-19 patients at a single center in Wuhan. Median IgG and IgM levels in acute and convalescent-phase sera (within 35 days) for all included patients were calculated and compared between severe and non-severe patients. Immune response phenotyping based on the late IgG levels and neutrophil-to-lymphocyte ratio (NLR) was characterized to stratified patients into different disease severities and outcomes. Results: A total of 222 patients were included in this study. IgG was first detected on day 4 of illness, and its peak levels occurred in the fourth week. Severe cases were more frequently found in patients with high IgG levels, compared to those with low IgG levels (51.8 vs. 32.3%; p = 0.008). Severity rates for patients with NLRhiIgGhi, NLRhiIgGlo, NLRloIgGhi, and NLRloIgGlo phenotype were 72.3, 48.5, 33.3, and 15.6%, respectively (p < 0.0001). Furthermore, severe patients with NLRhiIgGhi, NLRhiIgGlo had higher inflammatory cytokines levels including IL-2, IL-6 and IL-10, and decreased CD4+ T cell count compared to those with NLRloIgGlo phenotype (p < 0.05). Recovery rates for severe patients with NLRhiIgGhi, NLRhiIgGlo, NLRloIgGhi, and NLRloIgGlo phenotype were 58.8% (20/34), 68.8% (11/16), 80.0% (4/5), and 100% (12/12), respectively (p = 0.0592). Dead cases only occurred in NLRhiIgGhi and NLRhiIgGlo phenotypes. Conclusions: COVID-19 severity is associated with increased IgG response, and an immune response phenotyping based on the late IgG response and NLR could act as a simple complementary tool to discriminate between severe and non-severe COVID-19 patients, and further predict their clinical outcome.
... A considerable number of studies have demonstrated that infection with SARS-CoV-2 initiates an antibody response. On average, seroconversion is observed between 10 and 14 days post infection, a timeline similar to that observed for the previous SARS-CoV epidemic (58,(119)(120)(121)(122)(123). Serological investigations of 262 hospitalized COVID-19 patients across the Chongqing region, China, reported simultaneous or sequential seroconversion (IgM and IgG) against recombinant antigens containing the nucleoprotein and a peptide from the spike protein of SARS-CoV-2 (124). ...
Coronavirus disease-19 (COVID-19) describes a set of symptoms that develop following infection by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Whilst COVID-19 disease is most serious in patients with significant co-morbidities, the reason for healthy individuals succumbing to fulminant infection is largely unexplained. In this review, we discuss the most recent findings in terms of clinical features and the host immune response, and suggest candidate immune pathways that may be compromised in otherwise healthy individuals with fulminating COVID-19. On the basis of this early knowledge we reason a potential genetic effect on host immune response pathways leading to increased susceptibility to SARS-CoV-2 infection. Understanding these pathways may help not only in unraveling disease pathogenesis, but also in suggesting targets for therapy and prophylaxis. Importantly such insight should instruct efforts to identify those at increased risk in order to institute preventative measures, such as prophylactic medication and/or vaccination, when such opportunities arise in the later phases of the current pandemic or during future similar pandemics.
Vaksin DNA telah menjadi bidang penelitian yang menarik dalam upaya melawan penyakit infeksius pada hewan. Vaksin DNA adalah formulasi vaksin yang menggunakan fragmen DNA sebagai antigen. DNA pada vaksin mengandung informasi genetik yang diambil dari agen penyakit, seperti virus atau bakteri, yang ingin ditargetkan. Penelitian ini bertujuan untuk mengembangkan vaksin DNA pada hewan dan manusia. Metode penelitian menggunakan klon plasmid DNA dengan gen nukleoprotein virus Covid-19 yang ditransformasikan pada bakteri E. coli stellar. Pengujian titer antibodi vaksin pada mencit dilakukan dengan uji Enzyme-linked immunosorbent assay. Hasil penelitian menunukkan klon plasmid DNA dengan gen nukleoprotein virus SARS-Cov-2 dapat dikembangkan dan dibuktikan. Pemberian klon plasmid DNA dengan gen nukleoprotein pada mencit dapat menginduksi terbentuknya antibodi spesifik terhadap nukleoprotein virus SARS-Cov-2. Pemberian plasmid rekombinan pcDNA-Nukleoprotein terbukti menginduksi terbentuknya antibodi terhadap nukleoprotein yang meningkat dari minggu ke-1 sampai minggu ke-4 dan ke-5 yang kemudian menurun pada minggu ke 6 pasca vaksinasi.
Plant-derived extracellular vesicles (EVs) may represent a platform for the delivery of RNA-based vaccines, exploiting their natural membrane envelope to protect and deliver nucleic acids. Here, EVs extracted from orange (Citrus sinensis) juice (oEVs) were investigated as carriers for oral and intranasal SARS-CoV-2 mRNA vaccine. oEVs were efficiently loaded with different mRNA molecules (coding N, subunit 1 and full S proteins) and the mRNA was protected from degrading stress (including RNase and simulated gastric fluid), delivered to target cells and translated into protein. APC cells stimulated with oEVs loaded with mRNAs induced T lymphocyte activation in vitro. The immunization of mice with oEVs loaded with S1 mRNA via different routes of administration including intramuscular, oral and intranasal stimulated a humoral immune response with production of specific IgM and IgG blocking antibodies and a T cell immune response, as suggested by IFN-γ production by spleen lymphocytes stimulated with S peptide. Oral and intranasal administration also triggered the production of specific IgA, the mucosal barrier in the adaptive immune response. In conclusion, plant-derived EVs represent a useful platform for mRNA-based vaccines administered not only parentally but also orally and intranasally.
SARS-CoV-2-specific T cell response has been proven essential for viral clearance, COVID-19 outcome and long-term memory. Impaired early T cell-driven immunity leads to a severe form of the disease associated with lymphopenia, hyperinflammation and imbalanced humoral response. Analyses of acute SARS-CoV-2 infection have revealed that mild COVID-19 course is characterized by an early induction of specific T cells within the first 7 days of symptoms, coordinately followed by antibody production for an effective control of viral infection. In contrast, patients who do not develop an early specific cellular response and initiate a humoral immune response with subsequent production of high levels of antibodies, develop severe symptoms. Yet, delayed and persistent bystander CD8+ T cell activation has been also reported in hospitalized patients and could be a driver of lung pathology. Literature supports that long-term maintenance of T cell response appears more stable than antibody titters. Up to date, virus-specific T cell memory has been detected 22 months post-symptom onset, with a predominant IL-2 memory response compared to IFN-γ. Furthermore, T cell responses are conserved against the emerging variants of concern (VoCs) while these variants are mostly able to evade humoral responses. This could be partly explained by the high HLA polymorphism whereby the viral epitope repertoire recognized could differ among individuals, greatly decreasing the likelihood of immune escape. Current COVID-19-vaccination has been shown to elicit Th1-driven spike-specific T cell response, as does natural infection, which provides substantial protection against severe COVID-19 and death. In addition, mucosal vaccination has been reported to induce strong adaptive responses both locally and systemically and to protect against VoCs in animal models. The optimization of vaccine formulations by including a variety of viral regions, innovative adjuvants or diverse administration routes could result in a desirable enhanced cellular response and memory, and help to prevent breakthrough infections. In summary, the increasing evidence highlights the relevance of monitoring SARS-CoV-2-specific cellular immune response, and not only antibody levels, as a correlate for protection after infection and/or vaccination. Moreover, it may help to better identify target populations that could benefit most from booster doses and to personalize vaccination strategies.
In this study, our approach was to carry out a complete investigation of molecular docking analysis with the major SARS-CoV-2 proteins. We analysed more than 6000 drugs, downloaded by the PubChem database. Particular attention, we have focused on Spike Glycoprotein and Main protease 3CL pro Covid-19 proteins, by-Blind docking‖ method and-Selective docking‖ procedure, in Ligand Binding site, with AutoDock Vina using Pyrx software. From our results, we have selected Tirilazad against COVID-19. In fact, it reported having an excellent ability to bind both with the-Native Spike Glycoprotein‖, with a Binding Energy of-11.8 kcal mol-1 , and with the South African (B.1.351) SARS-CoV-2 spike protein variant, with a Binding Energy-10 kcal mol-1. Indeed, in the second case, the docking analysis was evaluated in the active area of three key amino acids belonging to the Spike Protein RBD, responsible for a higher binding with the ACE2 receptor. They are ASN 417, Lys 484, and Tyr 501 respectively. In addition, Tirilazad has shown a Binding energy score of approximately-10.5 kcal mol-1 against SARS-COV-2 Main protease. This has led us to conclude that this drug could be an excellent candidate against Coronavirus (COVID-19) pandemic, even though further in vitro and in vivo studies are needed.
Acute respiratory distress syndrome (ARDS) is a major complication of the respiratory illness coronavirus disease 2019, with a death rate reaching up to 40%. The main underlying cause of ARDS is a cytokine storm that results in a dysregulated immune response. This review discusses the role of cytokines and chemokines in SARS-CoV-2 and its predecessors SARS-CoV and MERS-CoV, with particular emphasis on the elevated levels of inflammatory mediators that are shown to be correlated with disease severity. For this purpose, we reviewed and analyzed clinical studies, research articles, and reviews published on PubMed, EMBASE, and Web of Science. This review illustrates the role of the innate and adaptive immune responses in SARS, MERS, and COVID-19 and identifies the general cytokine and chemokine profile in each of the three infections, focusing on the most prominent inflammatory mediators primarily responsible for the COVID-19 pathogenesis. The current treatment protocols or medications in clinical trials were reviewed while focusing on those targeting cytokines and chemokines. Altogether, the identified cytokines and chemokines profiles in SARS-CoV, MERS-CoV, and SARS-CoV-2 provide important information to better understand SARS-CoV-2 pathogenesis and highlight the importance of using prominent inflammatory mediators as markers for disease diagnosis and management. Our findings recommend that the use of immunosuppression cocktails provided to patients should be closely monitored and continuously assessed to maintain the desirable effects of cytokines and chemokines needed to fight the SARS, MERS, and COVID-19. The current gap in evidence is the lack of large clinical trials to determine the optimal and effective dosage and timing for a therapeutic regimen.
With the exception of inactivated vaccines, all SARS-CoV-2 vaccines currently used for clinical application focus on the spike envelope glycoprotein as a virus-specific antigen. Compared to other SARS-CoV-2 genes, mutations in the spike protein gene are more rapidly selected and spread within the population, which carries the risk of impairing the efficacy of spike-based vaccines. It is unclear to what extent the loss of neutralizing antibody epitopes can be compensated by cellular immune responses, and whether the use of other SARS-CoV-2 antigens might cause a more diverse immune response and better long-term protection, particularly in light of the continued evolution towards new SARS-CoV-2 variants. To address this question, we explored immunogenicity and protective effects of adenoviral vectors encoding either the full-length spike protein (S), the nucleocapsid protein (N), the receptor binding domain (RBD) or a hybrid construct of RBD and the membrane protein (M) in a highly susceptible COVID-19 hamster model. All adenoviral vaccines provided life-saving protection against SARS-CoV-2-infection. The most efficient protection was achieved after exposure to full-length spike. However, the nucleocapsid protein, which triggered a robust T-cell response but did not facilitate the formation of neutralizing antibodies, controlled early virus replication efficiently and prevented severe pneumonia. Although the full-length spike protein is an excellent target for vaccines, it does not appear to be the only option for future vaccine design.
Objective:
To determine the antibody levels (IgM and IgG), using ELISA in suspected patients of COVID-19.
Study design:
Descriptive cross-sectional study. Place & Duration of Study: Real Time PCR Diagnostic and Research Laboratory, Peshawar, Khyber Pakhtunkhwa, Pakistan, from May to July 2020.
Methodology:
A total of 94 blood specimens were collected from suspected COVID-19 patients. The antibody levels (IgG and IgM) were determined, using a COVID-19 ELISA IgG and IgM kit.
Results:
Out of a total 94 serum specimens, specimens were predominantly collected from males (70.2%, n=66) as compared to females (29.8%, n=28). Amongst six different age groups, the majority of the samples were found in the 31-45 years, 16-30 years, and 46-60 years groups, 42.6% (n=40), 23.4% (n=22) and 22.3% (n=21), respectively. Of the 94 suspected COVID-19 patients' serum specimens, IgG and IgM were detected in 29.8% (n=28) and 39.4% (n=37), specimens, respectively. The IgG antibodies were detected more in males (60.0%, n=18) than females (40.0%, n=12) samples. Similarly, IgM antibodies were also found more frequently in males (61.1%, n=22) as compared to females (38.9%, n=14).
Conclusion:
Detection of antibodies in COVID-19 infected patients provides vital clinical information for clinicians and could be used for the identification of suspected cases. Moreover, males were more prone to disease compared to females, and the 31-45 years age group was also more affected. Key Words: Serological assays, IgG, IgM, Peshawar, Pakistan.
Cytokines play pleiotropic, antagonistic, and collaborative in viral disease. The high morbidity and mortality of coronavirus disease 2019 (COVID-19) make it a significant threat to global public health. Elucidating its pathogenesis is essential to finding effective therapy. A retrospective study was conducted on 71 patients hospitalized with COVID-19. Data on cytokines, T lymphocytes, and other clinical and laboratory characteristics were collected from patients with variable disease severity. The effects of cytokines on the overall survival (OS) and event-free survival (EFS) of patients were analyzed. The critically severe and severe patients had higher infection indexes and significant multiple organ function abnormalities than the mild patients (P<0.05). IL-6 and IL-10 were significantly higher in the critically severe patients than in the severe and mild patients (P<0.05). IL-6 and IL-10 were closely associated with white blood cells, neutrophils, T lymphocyte subsets, D-D dimer, blood urea nitrogen, complement C1q, procalcitonin C-reactive protein. Moreover, the IL-6 and IL-10 levels were closely correlated to dyspnea and dizziness (P<0.05). The patients with higher IL-10 levels had shorter OS than the group with lower levels (P<0.05). The older patients with higher levels of single IL-6 or IL-10 tended to have shorter EFS (P<0.05), while the patients who had more elevated IL-6 and IL-10 had shorter OS (P<0.05). The Cox proportional hazard model revealed that IL-6 was the independent factor affecting EFS. IL-6 and IL-10 play crucial roles in COVID-19 prognosis.
A novel beta-coronavirus was reported in Wuhan, Hubei Province, China, which in December 2019, named SARS-CoV-2. It causes Coronavirus Disease 2019 (COVID-19) that can affect lung tissue and airways. The immune system can respond to SARS-CoV-2 infection via various mechanisms. Cytokines play crucial roles in COVID-19. In the present study, the latest information on the immune responses, transmission, symptoms, diagnosis, and treatment of COVID-19 is reviewed.Journal of Cellular & Molecular Anesthesia
O tratamento ultraconservador vem se destacando cientificamente como uma técnica que tem capacidade de controlar a lesão de cárie por meio da desorganização do biofilme dentário sem a restauração da cavidade. Assim, o objetivo deste estudo foi mostrar que uma lesão de cárie nem sempre necessita de intervenção invasiva desde que seja realizada a remoção das retenções da cavidade com o controle da dieta e higiene bucal pelo paciente. Abordamos dois casos de pa- cientes com lesões cariosas extensas, sem comprometimento pulpar e alterações patológicas no periápice. O primeiro caso relacionado a um paciente que apresentava lesão de cárie no dente 75, na qual a princípio a remoção das retenções da cavidade foram realizadas, mas por intervenção de outro profissional, restauração com cimento de ionômero de vidro foi realizada; e o segundo caso de um paciente que apresentava lesão de cárie inativa nos dentes 74 e 75, nos quais foi efetuado o acompanhamento clínico e radiográfico. Nos dois casos, os pais foram devidamente instruídos sobre a importância da higiene bucal e da dieta. Ambos os pacientes retornaram após 6 meses. No primeiro caso, a restauração não estava mais presente, enquanto no segundo caso não foi obser- vado progressão de cárie. Concluímos que o tratamento ultraconservador parece ser uma opção adequada para o controle de lesões de cárie, sem necessidade restauradora.
Context, Covert severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections could be seeding new outbreaks. How to identify asymptomatic SARS-CoV-2 infections early has become a global focus.
Objective, To explore the roles of immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies detection, nucleic acid tests and computed tomography (CT) scanning to identify asymptomatic SARS-CoV-2 infection.
Design, The clinical data of 389 individuals with close contacts including in general characteristics, SARS-CoV-2 etiology, serum-specific IgM and IgG antibody detection and CT imaging results, were systematically analyzed.
Results, The present study showed that only 89 of the 389 individuals with close contacts were positive after the first nucleic acid test, while 300 individuals were still negative after two nucleic acid tests. Among the 300 individuals, 75 did not have pneumonia, and the other 225 individuals had pulmonary imaging changes. A total of 143 individuals were eventually diagnosed as having asymptomatic infection through IgM antibody and IgG antibody detection. The sensitivity, specificity and false-negative rate of IgM and IgG antibody detection were approximately 97.1% (347/357), 95.3% (204/214) and 4.67% (10/214), respectively. It also indicated that over approximately 2 weeks, most individuals were both IgM positive and IgG positive, accounting for 68.57% (72/105). Over approximately 3 weeks, the proportion of IgM-positive and IgG-positive individuals decreased to 8.57% (9/105), and the proportion of IgM-negative and IgG-positive individuals increased to 76.19% (80/105).
Conclusions, There are highlighted prospects of IgM/IgG antibody detection as a preferred method in identifying the individuals with asymptomatic SARS CoV-2 infection, especially combined with nucleic acid tests and pulmonary CT scanning.
Since its emergence in December 2019, severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) has developed into a global pandemic within a matter of months. While subunit vaccines are one of the prominent options for combating coronavirus disease 2019 (COVID‐19), the immunogenicity of spike protein‐based antigens remains unknown. When immunized in mice, the S1 domain induced much higher IgG and IgA antibody levels than the RBD and more efficiently neutralized SARS‐CoV‐2 when adjuvanted with alum. It is inferred that a large proportion of these neutralization epitopes are located in the S1 domain but outside the RBD and that some of these are spatial epitopes. This finding indicates that expression systems with posttranslational modification abilities are important to maintain the natural configurations of recombinant spike protein antigens and are critical for effective COVID‐19 vaccines. Further, adjuvants prone to a Th1 response should be considered for S1‐based subunit COVID‐19 vaccines to reduce the potential risk of antibody‐dependent enhancement (ADE) of infection.
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We still know very little about how the human immune system responds to SARS-CoV-2. Here we construct a SARS-CoV-2 proteome microarray containing 18 out of the 28 predicted proteins and apply it to the characterization of the IgG and IgM antibodies responses in the sera from 29 convalescent patients. We find that all these patients had IgG and IgM antibodies that specifically bind SARS-CoV-2 proteins, particularly the N protein and S1 protein. Besides these proteins, significant antibody responses to ORF9b and NSP5 are also identified. We show that the S1 specific IgG signal positively correlates with age and the level of lactate dehydrogenase (LDH) and negatively correlates with lymphocyte percentage. Overall, this study presents a systemic view of the SARS-CoV-2 specific IgG and IgM responses and provides insights to aid the development of effective diagnostic, therapeutic and vaccination strategies. Currently very little is known about how our immune system responds to SARS-CoV-2 infection. Here the authors generate a SARS-CoV-2 proteome microarray for profiling of IgG and IgM responses to COVID-19 in patients and find significant responses to ORF9b and NSP5, as well as the S1 and N proteins.
The COVID-19 pandemic, caused by SARS-CoV-2, is a pressing public health emergency garnering rapid response from scientists across the globe. Host cell invasion is initiated through direct binding of the viral spike protein to the host receptor angiotensin-converting enzyme 2 (ACE2). Disrupting the spike-ACE2 interaction is a potential therapeutic target for treating COVID-19. We have developed a proximity-based AlphaLISA assay to measure binding of SARS-CoV-2 spike protein Receptor Binding Domain (RBD) to ACE2. Utilizing this assay platform, a drug-repurposing screen against 3,384 small molecule drugs and pre-clinical compounds was performed, yielding 25 high-quality, small-molecule hits that can be evaluated in cell-based models. This established AlphaLISA RBD-ACE2 platform can facilitate evaluation of biologics or small molecules that can perturb this essential viral-host interaction to further the development of interventions to address the global health pandemic.
In March 2003, a novel coronavirus (SARS-CoV) was discovered in association with cases of severe acute respiratory syndrome (SARS). The sequence of the complete genome of SARS-CoV was determined, and the initial characterization of the viral genome is presented in this report. The genome of SARS-CoV is 29,727 nucleotides in length and has 11 open reading frames, and its genome organization is similar to that of other coronaviruses. Phylogenetic analyses and sequence comparisons showed that SARS-CoV is not closely related to any of the previously characterized coronaviruses.
Cats that have succumbed to feline infectious peritonitis, an immune-mediated disease caused by variants of feline coronaviruses, show apoptosis and T-cell depletion in their lymphoid organs. The ascitic fluid that develops in the course of the condition causes apoptosis in vitro but only in activated T cells. Since feline infectious peritonitis virus does not infect T cells, and viral proteins did not inhibit T-cell proliferation, we postulate that soluble mediators released during the infection cause apoptosis and T-cell depletion.
DNA vaccination is an effective means of eliciting both humoral and cellular immunity, including cytotoxic T lymphocytes (CTL). Using an influenza virus model, we previously demonstrated that injection of DNA encoding influenza virus nucleoprotein (NP) induced major histocompatibility complex class I-restricted CTL and cross-strain protection from lethal virus challenge in mice (J. B. Ulmer et al., Science 259:1745-1749, 1993). In the present study, we have characterized in more detail the cellular immune responses induced by NP DNA, which included robust lymphoproliferation and Th1-type cytokine secretion (high levels of gamma interferon and interleukin-2 [IL-2], with little IL-4 or IL-10) in response to antigen-specific restimulation of splenocytes in vitro. These responses were mediated by CD4+ T cells, as shown by in vitro depletion of T-cell subsets. Taken together, these results indicate that immunization with NP DNA primes both cytolytic CD8+ T cells and cytokine-secreting CD4+ T cells. Further, we demonstrate by adoptive transfer and in vivo depletion of T-cell subsets that both of these types of T cells act as effectors in protective immunity against influenza virus challenge conferred by NP DNA.
Cytotoxic T lymphocytes (CTLs) are proposed to be critical for protection from intracellular pathogens such as Ebola virus.
However, there have been no demonstrations that protection against Ebola virus is mediated by Ebola virus-specific CTLs. Here,
we report that C57BL/6 mice vaccinated with Venezuelan equine encephalitis virus replicons encoding the Ebola virus nucleoprotein
(NP) survived lethal challenge with Ebola virus. Vaccination induced both antibodies to the NP and a major histocompatibility
complex class I-restricted CTL response to an 11-amino-acid sequence in the amino-terminal portion of the Ebola virus NP.
Passive transfer of polyclonal NP-specific antiserum did not protect recipient mice. In contrast, adoptive transfer of CTLs
specific for the Ebola virus NP protected unvaccinated mice from lethal Ebola virus challenge. The protective CTLs were CD8+, restricted to the Db class I molecule, and recognized an epitope within amino acids 43 to 53 (VYQVNNLEEIC) in the Ebola virus NP. The demonstration
that CTLs can prevent lethal Ebola virus infection affects vaccine development in that protective cellular immune responses
may be required for optimal protection from Ebola virus.
In 1997, the first documented instance of human respiratory disease and death associated with a purely avian H5N1 influenza virus resulted in an overall case-fatality rate of 33%. The biological basis for the severity of human H5N1 disease has remained unclear. We tested the hypothesis that virus-induced cytokine dysregulation has a role.
We used cDNA arrays and quantitative RT-PCR to compare the profile of cytokine gene expression induced by viruses A/HK/486/97 and A/HK/483/97 (both H5N1/97) with that of human H3N2 and H1N1 viruses in human primary monocyte-derived macrophages in vitro. Secretion of tumour necrosis factor alpha (TNF alpha) from macrophages infected with the viruses was compared by ELISA. By use of naturally occurring viral reassortants and recombinant viruses generated by reverse genetic techniques, we investigated the viral genes associated with the TNF-alpha response.
The H5N1/97 viruses induced much higher gene transcription of proinflammatory cytokines than did H3N2 or H1N1 viruses, particularly TNF alpha and interferon beta. The concentration of TNF-alpha protein in culture supernatants of macrophages infected with these viruses was similar to that induced by stimulation with Escherichia coli lipopolysaccharide. The non-structural (NS) gene-segment of H5N1/97 viruses contributed to the increase in TNF alpha induced by the virus.
The H5N1/97 viruses are potent inducers of proinflammatory cytokines in macrophages, the most notable being TNF alpha. This characteristic may contribute to the unusual severity of human H5N1 disease.
Severe acute respiratory syndrome (SARS) is a condition of unknown cause that has recently been recognized in patients in Asia, North America, and Europe. This report summarizes the initial epidemiologic findings, clinical description, and diagnostic findings that followed the identification of SARS in Canada.
SARS was first identified in Canada in early March 2003. We collected epidemiologic, clinical, and diagnostic data from each of the first 10 cases prospectively as they were identified. Specimens from all cases were sent to local, provincial, national, and international laboratories for studies to identify an etiologic agent.
The patients ranged from 24 to 78 years old; 60 percent were men. Transmission occurred only after close contact. The most common presenting symptoms were fever (in 100 percent of cases) and malaise (in 70 percent), followed by nonproductive cough (in 100 percent) and dyspnea (in 80 percent) associated with infiltrates on chest radiography (in 100 percent). Lymphopenia (in 89 percent of those for whom data were available), elevated lactate dehydrogenase levels (in 80 percent), elevated aspartate aminotransferase levels (in 78 percent), and elevated creatinine kinase levels (in 56 percent) were common. Empirical therapy most commonly included antibiotics, oseltamivir, and intravenous ribavirin. Mechanical ventilation was required in five patients. Three patients died, and five have had clinical improvement. The results of laboratory investigations were negative or not clinically significant except for the amplification of human metapneumovirus from respiratory specimens from five of nine patients and the isolation and amplification of a novel coronavirus from five of nine patients. In four cases both pathogens were isolated.
SARS is a condition associated with substantial morbidity and mortality. It appears to be of viral origin, with patterns suggesting droplet or contact transmission. The role of human metapneumovirus, a novel coronavirus, or both requires further investigation.
We sequenced the 29,751-base genome of the severe acute respiratory syndrome (SARS)-associated coronavirus known as the Tor2 isolate. The genome sequence reveals that this coronavirus is only moderately related to other known coronaviruses, including two human coronaviruses, HCoV-OC43 and HCoV-229E. Phylogenetic analysis of the predicted viral proteins indicates that the virus does not closely resemble any of the three previously known groups of coronaviruses. The genome sequence will aid in the diagnosis of SARS virus infection in humans and potential animal hosts (using polymerase chain reaction and immunological tests), in the development of antivirals (including neutralizing antibodies), and in the identification of putative epitopes for vaccine development.
To evaluate the haematological findings of patients with severe acute respiratory syndrome (SARS).
Analysis of the demographic, clinical, and laboratory characteristics of patients with SARS.
Prince of Wales Hospital, Hong Kong. Subjects All patients with a diagnosis of SARS between 11 March and 29 March 2003 who had no pre-existing haematological disorders.
Clinical end points included the need for intensive care and death. Univariate and multivariate analyses were performed to examine factors associated with adverse outcome.
64 male and 93 female patients were included in this study. The most common findings included lymphopenia in 153 (98%) of the 157 patients, neutrophilia in 129 (82%), thrombocytopenia in 87 patients (55%), followed by thrombocytosis in 77 (49%), and isolated prolonged activated partial thromboplastin time in 96 patients (63%). The haemoglobin count dropped by more than 20 g/l from baseline in 95 (61%) patients. Four patients (2.5%) developed disseminated intravascular coagulation. Lymphopenia was shown in haemato-lymphoid organs at postmortem examination. Multivariate analysis showed that advanced age and a high concentration of lactate dehydrogenase at presentation were independent predictors of an adverse outcome. Subsets of peripheral blood lymphocytes were analysed in 31 patients. The counts of CD4 positive and CD8 positive T cells fell early in the course of illness. Low counts of CD4 and CD8 cells at presentation were associated with adverse outcomes.
Abnormal haematological variables were common among patients with SARS. Lymphopenia and the depletion of T lymphocyte subsets may be associated with disease activity.
Editor—In their review of the severe acute respiratory syndrome (SARS) Wong et al emphasise lymphopenia as a hallmark feature.1 Panesar suggested that glucocorticoids or stimulation of the hypothalamic-pituitary-adrenal axis leads to lymphocyte margination and that patients without lymphopenia may have adrenal insufficiency.2
Apoptosis may also explain the lymphopenia of SARS. In severe paramyxovirus infections in humans such as measles, lymphopenia is commonly present and associated with more severe disease. One of us with Carrington recently reported that lymphopenia is also seen with another paramyxovirus infection: respiratory syncytial virus, which causes bronchiolitis in young children.3
Children with more severe bronchiolitis from respiratory syncytial virus infection have significantly lower absolute lymphocyte counts than those with mild disease. Bronchiolitis is ubiquitous and, in the developed world, the commonest reason a child under 1 year of age is admitted to hospital. Studies in mice show that not only is the lymphocyte immune response to virus essential in controlling the virus but it also causes disease.4 The fact the immune response is both saint and sinner is believed to be why the use of ribavirin has proved less effective in bronchiolitis than was first hoped. hoped.
Figure 1
SARS, respiratory syncytial virus disease, measles, and sepsis show parallels in the occurrence of lymphopenia. In sepsis and measles apoptosis is believed to be the mechanism of lymphopenia. In models of sepsis, for example, inhibitors of apoptosis ameliorate illness and prevent death.5 There may be important therapeutic implications for patients with SARS from these new research areas.
The widespread threat of severe acute respiratory syndrome (SARS) to human life has spawned challenges to develop fast and accurate analytical methods for its early diagnosis and to create a safe antiviral vaccine for preventive use. Consequently, we thoroughly investigated the immunoreactivities with patient sera of a series of synthesized peptides from SARS-coronavirus structural proteins.
We synthesized 41 peptides ranging in size from 16 to 25 amino acid residues of relatively high hydrophilicity. The immunoreactivities of the peptides with SARS patient sera were determined by ELISA.
Four epitopic sites, S599, M137, N66, and N371-404, located in the SARS-coronavirus S, M, and N proteins, respectively, were detected by screening synthesized peptides. Notably, N371 and N385, located at the COOH terminus of the N protein, inhibited binding of antibodies to SARS-coronavirus lysate and bound to antibodies in >94% of samples from SARS study patients. N385 had the highest affinity for forming peptide-antibody complexes with SARS serum.
Five peptides from SARS structural proteins, especially two from the COOH terminus of the N protein, appear to be highly immunogenic and may be useful for serologic assays. The identification of these antigenic peptides contributes to the understanding of the immunogenicity and persistence of SARS coronavirus.
Spike (S) proteins of coronaviruses, including the coronavirus that causes severe acute respiratory syndrome (SARS), associate with cellular receptors to mediate infection of their target cells. Here we identify a metallopeptidase, angiotensin-converting enzyme 2 (ACE2), isolated from SARS coronavirus (SARS-CoV)-permissive Vero E6 cells, that efficiently binds the S1 domain of the SARS-CoV S protein. We found that a soluble form of ACE2, but not of the related enzyme ACE1, blocked association of the S1 domain with Vero E6 cells. 293T cells transfected with ACE2, but not those transfected with human immunodeficiency virus-1 receptors, formed multinucleated syncytia with cells expressing S protein. Furthermore, SARS-CoV replicated efficiently on ACE2-transfected but not mock-transfected 293T cells. Finally, anti-ACE2 but not anti-ACE1 antibody blocked viral replication on Vero E6 cells. Together our data indicate that ACE2 is a functional receptor for SARS-CoV.
To study the inflammatory cytokine profile in children with severe acute respiratory syndrome (SARS) and to investigate whether monoclonal antibody to tumor necrosis factor-alpha (TNF-alpha) could be considered for treatment of these patients.
Plasma inflammatory cytokine concentrations (interleukin [IL]-1beta, IL-6, IL-8, IL-10, IL-12p70, and TNF-alpha) were monitored longitudinally on admission, immediately before corticosteroids, and 1 to 2 days and 7 to 10 days after the drug treatment in a cohort of pediatric patients (n = 8) with virologic confirmed SARS-associated coronavirus infection. None of the patients required mechanical ventilation or intensive care treatment. All children except 1 (patient 3) received corticosteroids.
Plasma IL-1beta levels (excluding patient 3) were substantially elevated immediately before (range: 7-721 ng/L) and 7 to 10 days after (range: 7-664 ng/L) corticosteroid treatment. In contrast, the plasma concentrations of other key proinflammatory cytokines, including IL-6 and TNF-alpha, were not overtly increased in any of the patients throughout the course of illness. In addition, plasma IL-10 concentration was significantly lower 1 to 2 days and 7 to 10 days after corticosteroid treatment, compared with the immediate pretreatment level. Similarly, plasma IL-6 and IL-8 concentrations were significantly decreased 7 to 10 days after the drug treatment.
Pediatric SARS patients have markedly elevated circulating IL-1beta levels, which suggests selective activation of the caspase-1-dependent pathway. Other key proinflammatory cytokines, IL-6 and TNF-alpha, showed only mildly elevated levels at the initial phase of the illness. The current evidence does not support the use of TNF-alpha monoclonal antibody in this group of children.
The angiotensin-converting enzyme (ACE)-related carboxypeptidase, ACE2, is a type I integral membrane protein of 805 amino acids that contains one HEXXH + E zinc-binding consensus sequence. ACE2 has been implicated in the regulation of heart function and also as a functional receptor for the coronavirus that causes the severe acute respiratory syndrome (SARS). To gain further insights into this enzyme, the first crystal structures of the native and inhibitor-bound forms of the ACE2 extracellular domains were solved to 2.2- and 3.0-A resolution, respectively. Comparison of these structures revealed a large inhibitor-dependent hinge-bending movement of one catalytic subdomain relative to the other ( approximately 16 degrees ) that brings important residues into position for catalysis. The potent inhibitor MLN-4760 ((S,S)-2-[1-carboxy-2-[3-(3,5-dichlorobenzyl)-3H-imidazol4-yl]-ethylamino]-4-methylpentanoic acid) makes key binding interactions within the active site and offers insights regarding the action of residues involved in catalysis and substrate specificity. A few active site residue substitutions in ACE2 relative to ACE appear to eliminate the S(2)' substrate-binding subsite and account for the observed reactivity change from the peptidyl dipeptidase activity of ACE to the carboxypeptidase activity of ACE2.
Public health measures have successfully identified and contained outbreaks of the severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV), but concerns remain over the possibility of future recurrences. Finding a vaccine for this virus therefore remains a high priority. Here, we show that a DNA vaccine encoding the spike (S) glycoprotein of the SARS-CoV induces T cell and neutralizing antibody responses, as well as protective immunity, in a mouse model. Alternative forms of S were analysed by DNA immunization. These expression vectors induced robust immune responses mediated by CD4 and CD8 cells, as well as significant antibody titres, measured by enzyme-linked immunosorbent assay. Moreover, antibody responses in mice vaccinated with an expression vector encoding a form of S that includes its transmembrane domain elicited neutralizing antibodies. Viral replication was reduced by more than six orders of magnitude in the lungs of mice vaccinated with these S plasmid DNA expression vectors, and protection was mediated by a humoral but not a T-cell-dependent immune mechanism. Gene-based vaccination for the SARS-CoV elicits effective immune responses that generate protective immunity in an animal model.
Severe acute respiratory syndrome (SARS) is a recently emerged infectious disease caused by a novel coronavirus, but its immunopathological mechanisms have not yet been fully elucidated. We investigated changes in plasma T helper (Th) cell cytokines, inflammatory cytokines and chemokines in 20 patients diagnosed with SARS. Cytokine profile of SARS patients showed marked elevation of Th1 cytokine interferon (IFN)-gamma, inflammatory cytokines interleukin (IL)-1, IL-6 and IL-12 for at least 2 weeks after disease onset, but there was no significant elevation of inflammatory cytokine tumour necrosis factor (TNF)-alpha, anti-inflammatory cytokine IL-10, Th1 cytokine IL-2 and Th2 cytokine IL-4. The chemokine profile demonstrated significant elevation of neutrophil chemokine IL-8, monocyte chemoattractant protein-1 (MCP-1), and Th1 chemokine IFN-gamma-inducible protein-10 (IP-10). Corticosteroid reduced significantly IL-8, MCP-1 and IP-10 concentrations from 5 to 8 days after treatment (all P < 0.001). Together, the elevation of Th1 cytokine IFN-gamma, inflammatory cytokines IL-1, IL-6 and IL-12 and chemokines IL-8, MCP-1 and IP-10 confirmed the activation of Th1 cell-mediated immunity and hyperinnate inflammatory response in SARS through the accumulation of monocytes/macrophages and neutrophils.
Current treatment schemes for severe acute respiratory syndrome (SARS) include broad-spectrum antibiotics, glucocorticoids,
and ribavirin. We evaluated the susceptibility of the SARS-related coronavirus (SARS CoV) to ribavirin and interferon (IFN)-α
in vitro by use of cytopathic effect, plaque assay, and immunoblot analysis. Ribavirin did not inhibit viral growth at concentrations
attainable in human serum. In contrast, IFN-α showed an in vitro inhibitory effect starting at concentrations of 1000 IU/mL.
In conclusion, ribavirin alone is unlikely to be beneficial in the prophylaxis or treatment of SARS CoV infections. Clinical
trials with IFN-α might be justified to determine a beneficial effect on the outcome of SARS.
Severe acute respiratory syndrome (SARS) is a novel infectious disease caused by the SARS-associated coronavirus (SARS-CoV).
There are four major structural proteins in the SARS-CoV, including the nucleocapsid, spike, membrane, and small envelope
proteins. In this study, two sets of truncated fragments of spike protein were generated, the first were approximately 210-bp
nonoverlapping fragments and the second were overlapping segments of 750 to 900 bp. From these 23 fragments, we identified
a fragment of 259 amino acids (amino acids 441 to 700) that is a major immunodominant epitope. This fragment was highly expressed,
and the purified fragment C could detect all 33 SARS patient serum samples tested, collected from 7 to 60 days after the onset
of fever, but had no reactivity with all 66 healthy human serum samples tested. Thus, fragment C of spike protein was identified
as an immunodominant antigen and could be used for serological detection of SARS-CoV infection.
Severe acute respiratory syndrome (SARS) is a serious threat to public health and the economy on a global scale. The SARS
coronavirus (SARS-CoV) has been identified as the etiological agent for SARS. Thus, vaccination against SARS-CoV may represent
an effective approach to controlling SARS. DNA vaccines are an attractive approach for SARS vaccine development, as they offer
many advantages over conventional vaccines, including stability, simplicity, and safety. Our investigators have previously
shown that DNA vaccination with antigen linked to calreticulin (CRT) dramatically enhances major histocompatibility complex
class I presentation of linked antigen to CD8+ T cells. In this study, we have employed this CRT-based enhancement strategy to create effective DNA vaccines using SARS-CoV
nucleocapsid (N) protein as a target antigen. Vaccination with naked CRT/N DNA generated the most potent N-specific humoral
and T-cell-mediated immune responses in vaccinated C57BL/6 mice among all of the DNA constructs tested. Furthermore, mice
vaccinated with CRT/N DNA were capable of significantly reducing the titer of challenging vaccinia virus expressing the N
protein of the SARS virus. These results show that a DNA vaccine encoding CRT linked to a SARS-CoV antigen is capable of generating
strong N-specific humoral and cellular immunity and may potentially be useful for control of infection with SARS-CoV.
The spike protein (S), a membrane component of severe acute respiratory syndrome coronavirus (SARS-CoV) is anticipated to be an important component of candidate vaccines. We constructed recombinant forms of the highly attenuated modified vaccinia virus Ankara (MVA) containing the gene encoding full-length SARS-CoV S with and without a C-terminal epitope tag called MVA/S-HA and MVA/S, respectively. Cells infected with MVA/Sor MVA/S-HA synthesized a 200-kDa protein, which was recognized by antibody raised against a synthetic peptide of SARS-CoV S or the epitope tag in Western blot analyses. Further studies indicated that S was N-glycosylated and migrated in SDS polyacrylamide gels with an apparent mass of approximately 160 kDa after treatment with peptide N-glycosidase F. The acquisition of resistance to endoglycosidase H indicated trafficking of S to the medial Golgi compartment, and confocal microscopy showed that S was transported to the cell surface. Intranasal or intramuscular inoculations of BALB/c mice with MVA/S produced serum antibodies that recognized the SARS S in ELISA and neutralized SARS-CoV in vitro. Moreover, MVA/S administered by either route elicited protective immunity, as shown by reduced titers of SARS-CoV in the upper and lower respiratory tracts of mice after challenge. Passive transfer of serum from mice immunized with MVA/S to naïve mice also reduced the replication of SARS-CoV in the respiratory tract after challenge, demonstrating a role for antibody to S in protection. The attenuated nature of MVA and the ability of MVA/S to induce neutralizing antibody that protects mice support further development of this candidate vaccine.
We report the evaluation of recombinant severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) nucleocapsid protein
enzyme-linked immunosorbent assay (ELISA)-based antibody tests for serodiagnosis of SARS-CoV pneumonia and compare the sensitivities
and specificities of this ELISA for detection of immunoglobulin G (IgG), IgM, IgA, and their combinations with serum samples
from 149 healthy blood donors who donated blood 3 years ago as controls and 106 SARS-CoV pneumonia patients in Hong Kong.
The specificities of the ELISA for IgG, IgM, and IgA detection were 95.3, 96.6, and 96.6%, respectively, with corresponding
sensitivities of 94.3, 59.4, and 60.4%, respectively. The present ELISA appears to be a sensitive test for serodiagnosis of
SARS-CoV pneumonia, is much more economical and less labor-intensive than the indirect immunofluorescence assay, and does
not require cultivation of SARS-CoV.
The immunogenicity of HLA-A2-restricted T-cell epitopes in the S protein of the Severe acute respiratory syndrome coronavirus (SARS-CoV) and of human coronavirus strain 229e (HCoV-229e) was analyzed for the elicitation of a T-cell immune response in donors who had fully recovered from SARS-CoV infection. We employed online database analysis to compare the differences in the amino acid sequences of the homologous T epitopes of HCoV-229e and SARS-CoV. The identified T-cell epitope peptides were synthesized, and their binding affinities for HLA-A2 were validated and compared in the T2 cell system. The immunogenicity of all these peptides was assessed by using T cells obtained from donors who had fully recovered from SARS-CoV infection and from healthy donors with no history of SARS-CoV infection. HLA-A2 typing by indirect immunofluorescent antibody staining showed that 51.6% of SARS-CoV-infected patients were HLA-A2 positive. Online database analysis and the T2 cell binding test disclosed that the number of HLA-A2-restricted immunogenic epitopes of the S protein of SARS-CoV was decreased or even lost in comparison with the homologous sequences of the S protein of HCoV-229e. Among the peptides used in the study, the affinity of peptides from HCoV-229e (H77 and H881) and peptides from SARS-CoV (S978 and S1203) for binding to HLA-A2 was higher than that of other sequences. The gamma interferon (IFN-gamma) release Elispot assay revealed that only SARS-CoV-specific peptides S1203 and S978 induced a high frequency of IFN-gamma-secreting T-cell response in HLA-A2(+) donors who had fully recovered from SARS-CoV infection; such a T-cell epitope-specific response was not observed in HLA-A2(+) healthy donors or in HLA-A2(-) donors who had been infected with SARS-CoV after full recovery. Thus, T-cell epitopes S1203 and S978 are immunogenic and elicit an overt specific T-cell response in HLA-A2(+) SARS-CoV-infected patients.
Objective. To investigate the dynamic changes observed in serum levels of interleukins (ILs), tumor necrosis factor-α (TNF-α and transforming growth factor-β1 (TGF-β1) in severe acute respiratory syndrome (SARS) patients. Methods. Sixty-one cases of SARS with positive antibodies to SARS coronavirus (SARS-CoV) were classified into the following categories: initial stage (3 - 7 days), peak stage (8 - 14 days), and remission and recovery stage ( 15 - 27 days). Forty-four healthy individuals were used as controls. Serum levels of ILs, TNF-α and TGF-β1 were measured in all subjects. Serum antibodies to SARS-CoV were detected only in SARS cases. Results. The mean concentration of serum IL -6 in SARS patients did not differ from that in the control group in initial and peak stages, but became significantly higher in remission and recovery stage compared with the control group, initial and peak stages ( P < 0.01). The mean concentration of serum IL-8 in SARS patients did not differ from that of the control group in initial stage, but was significantly higher than control group in peak stage and remission and recovery stage ( P < 0.05). And it was more significantly higher in remission and recovery stage than in peak stage ( P < 0.01). The mean concentrations of IL-16 and TNF-α in SARS patients were higher than those of the control group for every length of the clinical courses investigated, and were especially high in remission and recovery stage ( P <0.01 ). SARS patients experienced higher concentration of serum IL-13 compared with the controls in initial stage ( P < 0.01), but returned to normal levels in peak stage and in remission and recovery stage. The mean concentration of serum IL-18 in SARS patients was significantly lower than that of the control group during all clinical courses ( P < 0.05 ). The mean concentration of serum TGF-β1 in SARS patients was higher than that of the control group during all clinical courses. Although TGF-β1 in serum decreased in remission and recovery stage in SARS patients, the average was still higher than that of the control group ( P <0.01 ). Conclusions. Most proinflammatory cytokines and TGF-β1 were elevated during the early phase of SARS, which may be associated with lung infiltration and proliferation. Concurrently, the mean concentration of serum IL-13 decreased gradually, and the mean concentration of serum IL-18 level in SARS patients was lower than that of the control group during all the courses of SARS, suggesting that the immune state of the patients with SARS was obviously abnormal. Observing the dynamic changes in blood cytokine levels can provide a scientific basis to assess pathogenesis and efficacy of clinical treatment of SARS.
To understand the time-course of viraemia and antibody responses to severe acute respiratory syndrome-associated coronavirus (SARS-CoV), RT-PCR and ELISA were used to assay 376 blood samples from 135 SARS patients at various stages of the illness, including samples from patients who were in their early convalescent phase. The results showed that IgM antibodies decreased and became undetectable 11 weeks into the recovery phase. IgG antibodies, however, remained detectable for a period beyond 11 weeks and were found in 100 % of patients in the early convalescent phase. SARS-CoV viraemia mainly appeared 1 week after the onset of illness and then decreased over a period of 1 month, becoming undetectable in the blood samples of the convalescent patients. At the peak of viraemia, viral RNA was detectable in 75 % of blood samples from patients who were clinically diagnosed with SARS 1 or 2 weeks before the test.
Mollicute-Like Organisms (MLO) have been reported to be a cause of uveal tract and orbital chronic inflammatory disease. MLO are intracellular cytopathogenic cell wall deficient bacteria. No culture system exists for MLO, MLO disease diagnosis is based chiefly on direct detection of the organisms within diseased cells using a transmission electron microscope. Uveitis producing MLO are detectable within vitreous leucocytes as 0.005-0.01 micron filaments and undulating pleomorphic 0.01-1.0 micron tubulo-spherical bodies. Human uveitis producing MLO can be passed to laboratory animals. Inoculation into mouse eyelids produced intraocular, orbital, and lethal systemic chronic progressive inflammatory disease. MLO parasitised lesional leucocytes were found in all the disease sites. The MLO induced mouse chronic interstitial pneumonitis displayed 'sarcoid-like' granulomas. This report describes MLO parasitised vitreous leucocytes in the chronic uveitis of four sarcoidosis patients. The results indicate that MLO caused the uveitis. The implications of the results and Rifampin treatment of MLO disease are discussed.
Cytokine secretion is not confined to CD4+ T cells; rather, Type 1 and Type 2 populations of CD8+ and gamma delta T cells can also be generated in vitro and isolated from in vivo situations. These subsets and their physiological functions are significant.
Following infection of haplotype defined NIH-miniswine with virulent transmissible gastroenteritis coronavirus (TGEV), isolated mesenteric lymph node CD4+ T-cells mounted a specific proliferative response against infectious or inactivated purified virus in secondary in vitro stimulation. A specific, dose-dependent response to the three major recombinant viral proteins: spike (S), membrane (M), and nucleoprotein (N), purified by affinity chromatography, was characterized. Induction of in vitro antibody synthesis was analyzed. The purified recombinant viral proteins induced the in vitro synthesis of neutralizing TGEV-specific antibodies when porcine TGEV-immune cells were stimulated with each of the combinations made with two of the major structural proteins: S + N, S + M, and to a minor extent with M + N, but not by the individual proteins. S-protein was dissociated from purified virus using NP-40 detergent and then micellar S-protein oligomers (S-rosettes) were formed by removing the detergent. These occurred preferentially by the association of more than 10 S-protein trimmers. These S-rosettes in collaboration with either N or M-proteins elicited TGEV-specific antibodies with titers up to 84 and 60%, respectively, of those induced by the whole virus. N-protein could be partially substituted by a 15-mer peptide that represents a T helper epitope previously identified in N-protein (Antón et al. (1995)). These results indicate that the induction of high levels of TGEV-specific antibodies requires stimulation by at least two viral proteins, and that optimum responses are induced by a combination of S-rosettes and the nucleoprotein.
A recombinant baculovirus containing the S1 glycoprotein gene of the virulent nephropathogenic KM91 strain of infectious bronchitis virus (IBV) was constructed in order to investigate protective immunity in vaccinated chickens. Results from the protection test were evaluated by re-isolation of virus from the kidneys and tracheas of vaccinated chickens after challenge with strain KM91. After three immunizations, the recombinant S1 (rS1) glycoprotein induced 50% protection of the kidney, whilst inactivated KM91 induced 88% and 50% protection of the kidney and trachea, respectively. In chickens primed with the attenuated H120 vaccine strain, which is heterologous to KM91, the rS1 glycoprotein induced 83% protection of the kidney after two immunizations. Haemagglutination-inhibition titres were also increased in chickens immunized with the rS1 glycoprotein after three immunizations, and significantly higher titres were detected after challenge. These data indicate that the expressed rS1 glycoprotein alone can induce a protective immune response as well as an antibody response.
DNA vaccines expressing the envelope glycoprotein (GP) or nucleocapsid protein (NP) genes of Ebola virus were evaluated in adult, immunocompetent mice. The vaccines were delivered into the skin by particle bombardment of DNA-coated gold beads with the Powderject-XR gene gun. Both vaccines elicited antibody responses as measured by ELISA and elicited cytotoxic T cell responses as measured by chromium release assays. From one to four vaccinations with 0.5 microgram of the GP DNA vaccine resulted in a dose-dependent protection from Ebola virus challenge. Maximal protection (78% survival) was achieved after four vaccinations. Mice were completely protected with a priming dose of 0.5 microgram of GP DNA followed by three or four subsequent vaccinations with 1.5 micrograms of DNA. Partial protection could be observed for at least 9 months after three immunizations with 0.5 microgram of the GP DNA vaccine. Comparing the GP and NP vaccines indicated that approximately the same level of protection could be achieved with either vaccine.
Cytotoxic T-lymphocyte (CTL) response is an important component of anti-viral immunity. CTLs are specific to short peptides presented by MHC-I molecules and immunisation with the exact peptide sequence introduced in the cytosol is therefore a minimal approach, which potentially affords a high degree of controllability. We have examined the induction of murine CTL's by this approach using DNA plasmid minigene vaccines encoding known mouse K(k) minimal CTL epitopes (8 amino acids) from the influenza A virus hemagglutinin and nucleoprotein. We here report that such an approach is feasible and that wild type influenza virus flanking amino acid sequences can influence the CTL response but are not essential for optimal CTL induction. We also examined the effect of different new amino acid sequences flanking the CTL epitopes. In one version, two CTL epitopes were linked together as 'string of beads'. This did not improve CTL induction. In another version, one CTL epitope was inserted into a known T-helper protein (HBsAg). This did significantly augment the response probably due to immunological help from HBsAg Th epitopes. Finally, the CTL inducing minigene DNA vaccines were compared with Flu-induced CTL responses and tested for their protective effect against a lethal influenza A virus infection in mice and no effect was found. We conclude that a specific and highly directed CTL induction is possible by unlinked minigene DNA immunisation, but that CTL induction solely is not always sufficient to provide protection.
Cytotoxic T-lymphocyte (CTL) responses to measles virus (MV) play an important role in recovery from infection, with one of the major target proteins for CTL activity being the nucleoprotein (Np). In this report, a replication-deficient adenovirus-5 recombinant, expressing for MV Np (Rad68) was tested for in vivo priming of MV Np-specific CTL responses in BALB/c and CBA mice. In both strains of mice strong Np-specific CTL responses were induced and these responses were shown to be MHC class I restricted. Using overlapping 15mer peptides spanning residues 1-505 of MV Np a single epitope comprising residues 281-295 was identified in BALB/c mice whereas, in CBA mice two epitopes comprising residues 51-65 and 81-95, were identified. These epitopes were found to contain class I motifs for H-2L(d) and H-2K(k) MHC molecules, respectively. Immunization of BALB/c and CBA mice with the respective CTL epitopes resulted in the in vivo induction of peptide-and MV Np-specific CTL responses. In addition, the identified H-2K(k) restricted CTL epitopes conferred some protection against encephalitis induced following intracerebral challenge with a lethal dose of canine distemper virus (the Np of which shares 70% sequence homology with MV Np). These findings highlight the potential of using well-defined CTL epitopes to control virus infection.
Eighteen cases of human influenza A H5N1 infection were identified in Hong Kong from May to December 1997. Two of the six fatal cases had undergone a full post-mortem which showed reactive hemophagocytic syndrome as the most prominent feature. Other findings included organizing diffuse alveolar damage with interstitial fibrosis, extensive hepatic central lobular necrosis, acute renal tubular necrosis and lymphoid depletion. Elevation of soluble interleukin-2 receptor, interleukin-6 and interferon-gamma was demonstrated in both patients, whereas secondary bacterial pneumonia was not observed. Virus detection using isolation, reverse transcription-polymerase chain reaction and immunostaining were all negative. It is postulated that in fatal human infections with this avian subtype, initial virus replication in the respiratory tract triggers hypercytokinemia complicated by the reactive hemophagocytic syndrome. These findings suggest that the pathogenesis of influenza A H5N1 infection might be different from that of the usual human subtypes H1-H3.
Blood was collected from 55 cats with feline infectious peritonitis (FIP) and from 50 control cats in order to define whether differences in pathological findings and in distribution of feline coronaviruses (FCoV) can be associated with changes in haemograms, serum protein electrophoresis, and antibody titres. Compared to controls, the whole group of FIP-affected cats had blood changes consistent with FIP. Based on the pathological findings or on the immunohistochemical distribution of viral antigen, FIP-affected cats were divided in the following groups: subacute against acute lesions; low against strong intensity of positivity; intracellular against extracellular positivities; positive against negative lymph nodes. Lymphopenia was more evident in cats with acute forms, strong intensity of positivity, extracellular antigen and negative lymph nodes. Cats with positive lymph nodes had the most evident changes in the protein estimations. These results suggest that differences in pathological findings might depend on different reactive patterns to the FCoVs.
The spike 1 (S1) surface glycoprotein of infectious bronchitis virus (IBV) is the major inducer of the generation of virus neutralizing antibodies, and the administration of purified S1 has been shown to elicit a protective immune response against virulent virus challenge. On the basis of these observations, recombinant fowl poxvirus (rFPV) containing a cDNA copy of the S1 gene of IBV Mass 41 (rFPV-S1) was constructed and its immunogenicity and vaccine potential were evaluated. Initially, rFPV-S1 was shown to express the S1 in vito by indirect immunofluorescence staining and western blot analyses. Later, in vivo expression was demonstrated by the detection of IBV-specific serum immunoglobulin G and neutralization antibodies in the sera of chickens immunized with rFPV-S1. That the recombinant virus elicited anti-IBV protective immunity was indicated by the manifested, relatively mild clinical signs of disease, decreased titers of recovered challenge virus, and less severe histologic changes of the tracheas in virulent IBV Mass 41-challenged chickens previously receiving rFPV-S1 as compared with parental fowl poxvirus (FPV)-vaccinated control birds. In contrast, chickens immunized with either recombinant or parental FPV were resistant to a subsequent virulent FPV challenge. As to a preferred method of immunization, wing web administration appeared to be superior to the subcutaneous route because a greater percentage of birds vaccinated by the former protocol exhibited an anti-IBV humoral immune response. Thus, rFPV-S1 has potential as a poultry vaccine against both fowl pox and infectious bronchitis.
The cause of severe acute respiratory syndrome (SARS) has been identified as a new coronavirus. Whole genome sequence analysis of various isolates might provide an indication of potential strain differences of this new virus. Moreover, mutation analysis will help to develop effective vaccines.
We sequenced the entire SARS viral genome of cultured isolates from the index case (SIN2500) presenting in Singapore, from three primary contacts (SIN2774, SIN2748, and SIN2677), and one secondary contact (SIN2679). These sequences were compared with the isolates from Canada (TOR2), Hong Kong (CUHK-W1 and HKU39849), Hanoi (URBANI), Guangzhou (GZ01), and Beijing (BJ01, BJ02, BJ03, BJ04).
We identified 129 sequence variations among the 14 isolates, with 16 recurrent variant sequences. Common variant sequences at four loci define two distinct genotypes of the SARS virus. One genotype was linked with infections originating in Hotel M in Hong Kong, the second contained isolates from Hong Kong, Guangzhou, and Beijing with no association with Hotel M (p<0.0001). Moreover, other common sequence variants further distinguished the geographical origins of the isolates, especially between Singapore and Beijing.
Despite the recent onset of the SARS epidemic, genetic signatures are emerging that partition the worldwide SARS viral isolates into groups on the basis of contact source history and geography. These signatures can be used to trace sources of infection. In addition, a common variant associated with a non-conservative aminoacid change in the S1 region of the spike protein, suggests that immunological pressures might be starting to influence the evolution of the SARS virus in human populations.
The spike peplomer S1 subunit sequence from avian infectious bronchitis virus (IBV) Vic S strain was expressed in a plasmid under the control of the fowl adenovirus (FAV) major late promoter (MLP). Two recombinants were constructed in FAV serotype 8 (FAV 8) by inserting the expression cassette between the SnaBI and XbaI restriction enzyme sites (clone DA3) or between the SpeI sites (clone CA6-20). Expression of the S1 gene in the recombinants was confirmed by reverse transcription-polymerase chain reaction (RT-PCR) by 20h post-infection. Commercial broiler chickens were orally vaccinated at day 0 or day 6 post-hatch and challenged at day 35 post-hatch. FAV antibody ELISA confirmed that maternal antibody directed against inclusion body hepatitis (serotype 8) had decayed in control birds and that FAV specific serum IgG responses were produced in vaccinated birds at the time of challenge. Further, an S1 specific antibody response was detected prior to challenge. Birds were challenged with either Vic S (serotype B) or N1/62 (serotype C) strains of IBV. The tracheas of challenged birds were analyzed by RT-PCR and re-isolation of virus. In birds vaccinated at day 6, 90-100% protection at the trachea was induced against either homologous or heterologous challenge. The construction of a recombinant FAV expressing S1 of IBV demonstrates the potential of an alternative vaccination strategy against IBV.
To observe the changes of serum interleukins (IL), T-lymphocyte subsets, and white blood cell (WBC) count in patients with severe acute respiratory syndrome (SARS), and to investigate the relationship between injured immune function, immune response and disturbed immune adjustment in SARS patients.
The levels of serum IL-2, IL-10, IL-12 and T-lymphocyte subset counts were measured in 35 clinically diagnosed SARS patients by using enzyme linked immunosorbant assay (ELISA). The relationship between the measured results and WBC count was further analyzed.
The level of serum IL was increased to a great extent in the 35 SARS patients, and the levels of serum IL-2, IL-10 and IL-12 were 242.53 (92.69) pg/ml, 77.43 (63.37) pg/ml and 65.94 (43.21) pg/ml, respectively. The level of serum IL-2 increased markedly (P < 0.01). The peripheral blood CD(3)(+), CD(4)(+) and CD(8)(+) counts were lower than normal in 23 patients (67.7%), 26 patients (74.3%) and 15 patients (42.9%), respectively. The peripheral blood WBC counts were lower than 4.0 x 10(9)/L in 10 patients, and their CD(3)(+), CD(4)(+) and CD(8)(+) counts were 583.90 (315.58) x 10(6)/L, 272.00 (94.13) x 10(6)/L and 209.00 (72.21) x 10(6)/L, respectively. The peripheral blood WBC counts were (4.0 - 10.0) x 10(9)/L in 20 patients, and their CD(3)(+), CD(4)(+) and CD(8)(+) counts were 700.00 (502.96) x 10(6)/L, 347.00 (247.58) x 10(6)/L and 322.05 (228.47) x 10(6)/L, respectively. The peripheral blood WBC counts were higher than 10.0 x 10(9)/L in 5 patients, and their CD(3)(+), CD(4)(+) and CD(8)(+) counts were 1466.00 (630.86) x 10(6)/L, 783.00 (311.14) x 10(6)/L and 640.00 (294.40) x 10(6)/L, respectively. The decreased CD(3)(+), CD(4)(+) and CD(8)(+) counts were consistent with the decreased WBC counts. The level of IL in SARS patients was significantly higher than that in patients with chronic hepatitis B (P < 0.01).
The level of serum IL is closely related to cell immunity in SARS patients. The level of serum IL is increased evidently while CD(3)(+), CD(4)(+) and CD(8)(+) counts decrease. Both serum IL and CD are associated with injury of immune function, and thus they could be regarded as a monitoring index for judging the condition of SARS patients and prescribing immune therapy.
To the Editor: A novel coronavirus called the severe acute respiratory syndrome (SARS)–associated coronavirus (CoV) has been identified as the causal agent of SARS.1–3 To understand the humoral immunity to this virus, we studied the profile of IgM and IgG antibody responses to SARS-CoV. IgM and IgG antibodies were analyzed by an indirect enzyme-linked immunosorbent assay in 20 patients with SARS from week 1 of their illness to week 12 and in 103 healthy contacts. All 20 patients tested negative for IgM and IgG at week 1 after the onset of symptoms. Of these patients, 16 tested positive for . . .
To summarize the clinical features of liver injury in patients with severe acute respiratory syndrome (SARS), providing information for further mechanism and clinical study.
The clinical and some laboratory data of 154 patients suffered from SARS were collected and analyzed, who were admitted to the isolation wards of Beijing You-an Hospital from March 11 to June 3, 2003. The serum samples were taken from 46 patients to detect IL-1beta, IL-2, IL-4, IL-6, IL-8, IL-10, TNF-alpha, endotoxin and hepatitis related viral inclusions. In addition, 11 patients were detected ultrasonically, and 3 patients were described pathological features. Other two groups including 15 healthy care workers and 22 patients with chronic hepatitis in the same period were selected and analyzed as controls.
When being admitted to hospital, serum ALT and (or) AST levels were elevated in 37.7% SARS patients. Some of them (43.1%) were mild, and most of them (56.9%) were moderate. Abnormal liver function mainly resulted from ALT elevation (70.7%), then both ALT and AST elevation (22.4%). The aminotransferases in 75.9% SARS patients normalized within two weeks, while they elevated in four patients during the hospitalization. There was a significant difference in ALT/AST elevation rates between severe and mild clinical type (chi2=19.28, P<0.05). Serum total bilirubin values elevated in 8.4% patients. Serum albumin and prealbumin levels decreased in 24.0% and 28.6% patients, respectively. Creatine kinase (CK) and (or) creatine kinase MB (CK-MB) levels elevated in 72.7% patients when they hospitalized. The six kinds of interleukins and TNF-alpha levels during the first week of hospitalization were higher than those in the fourth week and in control groups (t>or=1.67, P<0.05). The levels of some factors, such as IL-1beta, IL-6 and IL-10 in patients with elevated ALT, were higher than those in ALT normal patients (t>or=2.36, P<0.05). In the first week, only 15.2% patients had elevated serum endotoxin level. Ultrasonic examination and pathological observation showed no special features, compared with those in common acute hepatitis patients.
All the results suggest strongly that there may be a systemic inflammatory response syndrome (SIRS) in most SARS patients in early stage, and the liver damage is only its partial signs. It may be beneficial to suppress cytokines storm in SARS patients in early stage, which will stop the progression of SIRS and release hepatic damage and improve the prognosis of SARS patients.
In a cohort of 38 patients with severe acute respiratory syndrome (SARS), we observed leukopenia in 47% of patients, lymphopenia
in 84%, and T lymphopenia in 95%. CD4+ T lymphocyte levels were reduced in 100% of patients, CD8+ T lymphocyte levels were reduced in 87%, B lymphocyte levels were reduced in 76%, and natural killer cell levels were reduced
in 55%. Our data suggested that these patients' immune systems were impaired during the course of SARS. The absolute counts
of lymphocyte subsets demonstrated a clinical significance for patients with SARS.
To elucidate the mechanism of strong inflammatory response in severe acute respiratory syndrome (SARS) patients.
24 patients managed with a standard corticosteroid protocol developed by Peking Union Medical College Hospital were followed up for 2 months and their clinical outcomes were documented. Plasma levels of interleukin (IL)-1 beta, IL-2, IL-4, IL-8, IL-10, IL-12 p70, interferon gamma (IFN gamma), and tumor necrosis factor alpha (TNF alpha) were determined by quantitative ELISA during the course of disease respectively. 12 healthy blood donors were used as normal controls.
All SARS patients had remarkably increased plasma IL-8 concentrations (median 31.23 ng/L) at the onset of disease compared with those of normal controls (6.28 ng/L, P < 0.01). Along with the course of disease IL-8 concentration kept going up and 75% (18/24) patients reached a peak median concentration of 149.65 ng/L (P < 0.01) at the third and fourth weeks. IL-8 came back to normal control levels one month after discharged from hospital. Meanwhile, SARS patients also showed high a TNF alpha level (median 23.12 ng/L) (P < 0.01) at the second week and reached a peak median of 136.35 ng/L (P < 0.05) at the third an fourth weeks. One month after discharged from hospital the plasma TNF alpha concentration fell down to a median of 94.88 ng/L, but it was still much higher than normal controls (3.77 ng/L, P < 0.01).
The SARS-associated coronovirus infection may cause complex cytokine cascade. IL-8 and TNF alpha probably play important roles in mediating strong inflammatory response, which are thought to be responsible for lung injury in SARS patients.
The causative agent of severe acute respiratory syndrome (SARS) has been identified as a new type of coronavirus. Here, we have investigated the ability of adenoviral delivery of codon-optimised SARS-CoV strain Urbani structural antigens spike protein S1 fragment, membrane protein, and nucleocapsid protein to induce virus-specific broad immunity in rhesus macaques. We immunised rhesus macaques intramuscularly with a combination of the three Ad5-SARS-CoV vectors or a control vector and gave a booster vaccination on day 28. The vaccinated animals all had antibody responses against spike protein S1 fragment and T-cell responses against the nucleocapsid protein. All vaccinated animals showed strong neutralising antibody responses to SARS-CoV infection in vitro. These results show that an adenoviral-based vaccine can induce strong SARS-CoV-specific immune responses in the monkey, and hold promise for development of a protective vaccine against the SARS causal agent.
Vaccines against infectious bronchitis of chickens (Gallus gallus domesticus) have arguably been the most successful, and certainly the most widely used, of vaccines for diseases caused by coronaviruses, the others being against bovine, canine, feline and porcine coronaviruses. Infectious bronchitis virus (IBV), together with the genetically related coronaviruses of turkey (Meleagris gallopovo) and ring-necked pheasant (Phasianus colchicus), is a group 3 coronavirus, severe acute respiratory syndrome (SARS) coronavirus being tentatively in group 4, the other known mammalian coronaviruses being in groups 1 and 2. IBV replicates not only in respiratory tissues (including the nose, trachea, lungs and airsacs, causing respiratory disease), but also in the kidney (associated with minor or major nephritis), oviduct, and in many parts of the alimentary tract--the oesophagus, proventriculus, duodenum, jejunum, bursa of Fabricius, caecal tonsils (near the distal end of the tract), rectum and cloaca (the common opening for release of eggs and faeces), usually without clinical effects. The virus can persist, being re-excreted at the onset of egg laying (4 to 5 months of age), believed to be a consequence of the stress of coming into lay. Genetic lines of chickens differ in the extent to which IBV causes mortality in chicks, and in respect of clearance of the virus after the acute phase. Live attenuated (by passage in chicken embryonated eggs) IBV strains were introduced as vaccines in the 1950s, followed a couple of decades later by inactivated vaccines for boosting protection in egg-laying birds. Live vaccines are usually applied to meat-type chickens at 1 day of age. In experimental situations this can result in sterile immunity when challenged by virulent homologous virus. Although 100% of chickens may be protected (against clinical signs and loss of ciliary activity in trachea), sometimes 10% of vaccinated chicks do not respond with a protective immune response. Protection is short lived, the start of the decline being apparent 9 weeks after vaccination with vaccines based on highly attenuated strains. IBV exists as scores of serotypes (defined by the neutralization test), cross-protection often being poor. Consequently, chickens may be re-vaccinated, with the same or another serotype, two or three weeks later. Single applications of inactivated virus has generally led to protection of <50% of chickens. Two applications have led to 90 to 100% protection in some reports, but remaining below 50% in others. In practice in the field, inactivated vaccines are used in laying birds that have previously been primed with two or three live attenuated virus vaccinations. This increases protection of the laying birds against egg production losses and induces a sustained level of serum antibody, which is passed to progeny. The large spike glycoprotein (S) comprises a carboxy-terminal S2 subunit (approximately 625 amino acid residues), which anchors S in the virus envelope, and an amino-terminal S1 subunit (approximately 520 residues), believed to largely form the distal bulbous part of S. The S1 subunit (purified from IBV virus, expressed using baculovirus or expressed in birds from a fowlpoxvirus vector) induced virus neutralizing antibody. Although protective immune responses were induced, multiple inoculations were required and the percentage of protected chickens was too low (<50%) for commercial application. Remarkably, expression of S1 in birds using a non-pathogenic fowl adenovirus vector induced protection in 90% and 100% of chickens in two experiments. Differences of as little as 5% between the S1 sequences can result in poor cross-protection. Differences in S1 of 2 to 3% (10 to 15 amino acids) can change serotype, suggesting that a small number of epitopes are immunodominant with respect to neutralizing antibody. Initial studies of the role of the IBV nucleocapsid protein (N) in immunity suggested that immunization with bacterially expressed N, while not inducing protection directly, improved the induction of protection by a subsequent inoculation with inactivated IBV. In another study, two intramuscular immunizations of a plasmid expressing N induced protective immunity. The basis of immunity to IBV is not well understood. Serum antibody levels do not correlate with protection, although local antibody is believed to play a role. Adoptive transfer of IBV-infection-induced alphabeta T cells bearing CD8 antigen protected chicks from challenge infection. In conclusion, live attenuated IBV vaccines induce good, although short-lived, protection against homologous challenge, although a minority of individuals may respond poorly. Inactivated IBV vaccines are insufficiently efficacious when applied only once and in the absence of priming by live vaccine. Two applications of inactivated IBV are much more efficacious, although this is not a commercially viable proposition in the poultry industry. However, the cost and logistics of multiple application of a SARS inactivated vaccine would be more acceptable for the protection of human populations, especially if limited to targeted groups (e.g. health care workers and high-risk contacts). Application of a SARS vaccine is perhaps best limited to a minimal number of targeted individuals who can be monitored, as some vaccinated persons might, if infected by SARS coronavirus, become asymptomatic excretors of virus, thereby posing a risk to non-vaccinated people. Looking further into the future, the high efficacy of the fowl adenovirus vector expressing the IBV S1 subunit provides optimism for a live SARS vaccine, if that were deemed to be necessary, with the possibility of including the N protein gene.
The angiotensin-converting enzyme 2 (ACE2) is an important regulator of the renin-angiotensin system and was very recently identified as a functional receptor for the SARS virus. The ACE2 sequence is similar (sequence identities 43% and 35%, and similarities 61% and 55%, respectively) to those of the testis-specific form of ACE (tACE) and the Drosophila homolog of ACE (AnCE). The high level of sequence similarity allowed us to build a robust homology model of the ACE2 structure with a root-mean-square deviation from the aligned crystal structures of tACE and AnCE less than 0.5A. A prominent feature of the model is a deep channel on the top of the molecule that contains the catalytic site. Negatively charged ridges surrounding the channel may provide a possible binding site for the positively charged receptor-binding domain (RBD) of the S-glycoprotein, which we recently identified [Biochem. Biophys. Res. Commun. 312 (2003) 1159]. Several distinct patches of hydrophobic residues at the ACE2 surface were noted at close proximity to the charged ridges that could contribute to binding. These results suggest a possible binding region for the SARS-CoV S-glycoprotein on ACE2 and could help in the design of experiments to further elucidate the structure and function of ACE2.
Early detection and identification of SARS-CoV-infected patients and actions to prevent transmission are absolutely critical to prevent another SARS outbreak. Antibodies that specifically recognize the SARS-CoV spike and nucleocapsid proteins may provide a rapid screening method to allow accurate identification and isolation of patients with the virus early in their infection. For this reason, we raised peptide-induced polyclonal antibodies against SARS-CoV spike protein and polyclonal antibodies against SARS-CoV nucleocapsid protein using 6x His nucleocapsid recombinant protein. Western blot analysis and immunofluorescent staining showed that these antibodies specifically recognized SARS-CoV.
Severe acute respiratory syndrome (SARS) is a novel infectious disease. No information is currently available on host-specific immunity against the SARS coronavirus (CoV), and detailed characteristics of the epidemiology of SARS CoV infection have not been identified.
ELISA was used to detect antibody to SARS CoV. Reverse-transcriptase polymerase chain reaction was used to detect SARS CoV RNA. T cells in peripheral blood of patients were quantified by flow cytometry.
Of 36 patients with probable SARS CoV infection, 30 (83.3%) were positive for IgG antibody to SARS CoV; in contrast, only 3 of 48 patients with suspected SARS CoV infection, 0 of 112 patients with fever but without SARS, and 0 of 96 healthy control individuals were positive for it. IgG antibody to SARS CoV was first detected between day 5 and day 47 after onset of illness (mean +/- SD, 18.7+/-10.4).
Detection of antibody to SARS CoV is useful in the diagnosis of SARS; however, at the incubation and initial phases of the illness, serological assay is of little value, because of late seroconversion in most patients.
Induction of effective cytotoxic T lymphocyte (CTL) and/or a specific antibody against conserved viral proteins may be essential to the development of a safe and effective severe acute respiratory syndrome coronavirus (SARS-Cov) vaccine. DNA vaccination represents a new strategy for induction of humoral and cellular immune response. To determine the ability of SARS-Cov nucleoprotein (N protein) to induce antiviral immunity, in this report, we established a stable C2C12 line expressing SARS-Cov N protein, which was used as a target for specific CTL assay. We also expressed recombinant N proteins in Escherichia coli and prepared N protein-specific polyclonal antibodies. C3H/He mice were immunized with N protein-expressible pcDN-fn vector by intramuscular injections. We found that the DNA vaccination induced both N protein-specific antibody and specific CTL activity to the target. When C3H/He mice were immunized by three separate injections, high antibody titre (1:3200-1:6400, average titre is 1:4580) and high CTL activity (67.4+/-8.4% (E:T = 25:1), 69.6+/-6.7% (E:T = 50:1) and 71.8+/-6.2% (E:T = 100:1)) were observed. In the case of two vaccine injections, CTL activity was also high (56.6+/-12.7% (E:T = 25:1), 57.4+/-11.7% (E:T = 50:1) and 63.0+/-6.3% (E:T = 100:1)) However, antibody titres were much lower (1:200-1:3200, average titre is 1:980). Our results suggest that SARS-Cov nucleocapsid gene might be a candidate gene for SARS DNA vaccination.
A novel coronavirus has been associated with a worldwide outbreak of atypical pneumonia referred to as Severe Acute Respiratory Syndrome (SARS-CoV). SARS-CoV nucleocapsid (N) protein has been cloned sequenced and expressed in Escherichia coli strain. Purified N protein was used to measure the SARS-CoV specific IgG antibodies from 16 SARS-CoV infected patients' sera and from 131 control subjects using ELISA assay. Specific antibody responses to the purified recombinant N protein after 10, 20, and 30 days of disease onset were observed in 13 of 16 (81.3%), 16 of 16 (100%) and 16 of 16 (100%) SARS patients sera, respectively. Comparison of detection results with a commercially available diagnostic kit coated with a mixture of SARS-CoV viral proteins showed 9 of 16 (56.3%), 13 of 16 (81.3%), and 15 of 16 (93.7%) positive responses, respectively. None of 131 control sera gave positive reaction in either assay. This data suggests that the N protein of SARS-CoV is immunodominant and this ELISA based test assay for detecting the SARS-CoV N antigen may hold a significant value for SARS diagnosis.