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1.
Nat Commun ; 12(1): 3587, 2021 06 11.
Article in English | MEDLINE | ID: covidwho-1387350

ABSTRACT

There is a great need for the development of vaccines that induce potent and long-lasting protective immunity against SARS-CoV-2. Multimeric display of the antigen combined with potent adjuvant can enhance the potency and longevity of the antibody response. The receptor binding domain (RBD) of the spike protein is a primary target of neutralizing antibodies. Here, we developed a trimeric form of the RBD and show that it induces a potent neutralizing antibody response against live virus with diverse effector functions and provides protection against SARS-CoV-2 challenge in mice and rhesus macaques. The trimeric form induces higher neutralizing antibody titer compared to monomer with as low as 1µg antigen dose. In mice, adjuvanting the protein with a TLR7/8 agonist formulation alum-3M-052 induces 100-fold higher neutralizing antibody titer and superior protection from infection compared to alum. SARS-CoV-2 infection causes significant loss of innate cells and pathology in the lung, and vaccination protects from changes in innate cells and lung pathology. These results demonstrate RBD trimer protein as a suitable candidate for vaccine against SARS-CoV-2.


Subject(s)
Adjuvants, Immunologic/administration & dosage , COVID-19 Vaccines/immunology , COVID-19/immunology , COVID-19/prevention & control , Heterocyclic Compounds, 3-Ring/administration & dosage , Stearic Acids/administration & dosage , Alum Compounds/administration & dosage , Angiotensin-Converting Enzyme 2/immunology , Angiotensin-Converting Enzyme 2/metabolism , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Antibody Formation/immunology , COVID-19 Vaccines/administration & dosage , Disease Models, Animal , Heterocyclic Compounds, 3-Ring/immunology , Humans , Macaca mulatta , Mice , Protein Binding , SARS-CoV-2/isolation & purification , Spike Glycoprotein, Coronavirus/immunology , Stearic Acids/immunology
2.
Sci Immunol ; 6(60)2021 06 15.
Article in English | MEDLINE | ID: covidwho-1270873

ABSTRACT

The inclusion of infants in the SARS-CoV-2 vaccine roll-out is important to prevent severe complications of pediatric SARS-CoV-2 infections and to limit transmission and could possibly be implemented via the global pediatric vaccine schedule. However, age-dependent differences in immune function require careful evaluation of novel vaccines in the pediatric population. Toward this goal, we assessed the safety and immunogenicity of two SARS-CoV-2 vaccines. Two groups of 8 infant rhesus macaques (RMs) were immunized intramuscularly at weeks 0 and 4 with stabilized prefusion SARS-CoV-2 S-2P spike (S) protein encoded by mRNA encapsulated in lipid nanoparticles (mRNA-LNP) or the purified S protein mixed with 3M-052, a synthetic TLR7/8 agonist in a squalene emulsion (Protein+3M-052-SE). Neither vaccine induced adverse effects. Both vaccines elicited high magnitude IgG binding to RBD, N terminus domain, S1, and S2, ACE2 blocking activity, and high neutralizing antibody titers, all peaking at week 6. S-specific memory B cells were detected by week 4 and S-specific T cell responses were dominated by the production of IL-17, IFN-γ, or TNF-α. Antibody and cellular responses were stable through week 22. The immune responses for the mRNA-LNP vaccine were of a similar magnitude to those elicited by the Moderna mRNA-1273 vaccine in adults. The S-2P mRNA-LNP and Protein-3M-052-SE vaccines were well-tolerated and highly immunogenic in infant RMs, providing proof-of concept for a pediatric SARS-CoV-2 vaccine with the potential for durable immunity that might decrease the transmission of SARS-CoV-2 and mitigate the ongoing health and socioeconomic impacts of COVID-19.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19 Vaccines/immunology , COVID-19/immunology , Animals , Animals, Newborn , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , COVID-19/prevention & control , Macaca mulatta , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/administration & dosage , Spike Glycoprotein, Coronavirus/immunology
3.
Emerg Microbes Infect ; 10(1): 1320-1330, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1266083

ABSTRACT

Ebola virus (EBOV) is a negative single-stranded RNA virus within the Filoviridae family and the causative agent of Ebola virus disease (EVD). Nonhuman primates (NHPs), including cynomolgus and rhesus macaques, are considered the gold standard animal model to interrogate mechanisms of EBOV pathogenesis. However, despite significant genetic similarity (>90%), NHP species display different clinical presentation following EBOV infection, notably a ∼1-2 days delay in disease progression. Consequently, evaluation of therapeutics is generally conducted in rhesus macaques, whereas cynomolgus macaques are utilized to determine efficacy of preventative treatments, notably vaccines. This observation is in line with reported differences in disease severity and host responses between these two NHP following infection with simian varicella virus, influenza A and SARS-CoV-2. However, the molecular underpinnings of these differential outcomes following viral infections remain poorly defined. In this study, we compared published transcriptional profiles obtained from cynomolgus and rhesus macaques infected with the EBOV-Makona Guinea C07 using bivariate and regression analyses to elucidate differences in host responses. We report the presence of a shared core of differentially expressed genes (DEGs) reflecting EVD pathology, including aberrant inflammation, lymphopenia, and coagulopathy. However, the magnitudes of change differed between the two macaque species. These findings suggest that the differential clinical presentation of EVD in these two species is mediated by altered transcriptional responses.


Subject(s)
Gene Expression Regulation/immunology , Hemorrhagic Fever, Ebola/veterinary , Macaca fascicularis , Macaca mulatta , Monkey Diseases/immunology , Transcription, Genetic/immunology , Animals , COVID-19 , Ebolavirus , Hemorrhagic Fever, Ebola/genetics , Hemorrhagic Fever, Ebola/immunology , Hemorrhagic Fever, Ebola/mortality , Humans , Immunity , Monkey Diseases/genetics , Monkey Diseases/mortality , RNA, Viral/metabolism , SARS-CoV-2 , Species Specificity
4.
MAbs ; 13(1): 1930636, 2021.
Article in English | MEDLINE | ID: covidwho-1258715

ABSTRACT

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), which causes coronavirus disease-2019 (COVID-19), interacts with the host cell receptor angiotensin-converting enzyme 2 (hACE2) via its spike 1 protein during infection. After the virus sequence was published, we identified two potent antibodies against the SARS-CoV-2 receptor binding domain (RBD) from antibody libraries using a phage-to-yeast (PtY) display platform in only 10 days. Our lead antibody JMB2002, now in a Phase 1 clinical trial (ChiCTR2100042150), showed broad-spectrum in vitro blocking activity against hACE2 binding to the RBD of multiple SARS-CoV-2 variants, including B.1.351 that was reportedly much more resistant to neutralization by convalescent plasma, vaccine sera and some clinical-stage neutralizing antibodies. Furthermore, JMB2002 has demonstrated complete prophylactic and potent therapeutic efficacy in a rhesus macaque disease model. Prophylactic and therapeutic countermeasure intervention of SARS-CoV-2 using JMB2002 would likely slow down the transmission of currently emerged SARS-CoV-2 variants and result in more efficient control of the COVID-19 pandemic.


Subject(s)
Angiotensin-Converting Enzyme 2/antagonists & inhibitors , Antibodies, Neutralizing/pharmacology , Antiviral Agents/pharmacology , COVID-19/drug therapy , COVID-19/prevention & control , SARS-CoV-2/drug effects , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/immunology , Animals , Antibody Specificity , Binding Sites, Antibody , CHO Cells , COVID-19/immunology , COVID-19/metabolism , COVID-19/virology , Chlorocebus aethiops , Cricetulus , Disease Models, Animal , Epitopes , Macaca mulatta , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Vero Cells
5.
R Soc Open Sci ; 8(4): 201891, 2021 Apr 28.
Article in English | MEDLINE | ID: covidwho-1234204

ABSTRACT

Measles virus (MV) is a highly contagious respiratory morbillivirus that results in many disabilities and deaths. A crucial challenge in studying MV infection is to understand the so-called 'measles paradox'-the progression of the infection to severe immunosuppression before clearance of acute viremia, which is also observed in canine distemper virus (CDV) infection. However, a lack of models that match in vivo data has restricted our understanding of this complex and counter-intuitive phenomenon. Recently, progress was made in the development of a model that fits data from acute measles infection in rhesus macaques. This progress motivates our investigations to gain additional insights from this model into the control mechanisms underlying the paradox. In this paper, we investigated analytical conditions determining the control and robustness of viral clearance for MV and CDV, to untangle complex feedback mechanisms underlying the dynamics of acute infections in their natural hosts. We applied control theory to this model to help resolve the measles paradox. We showed that immunosuppression is important to control and clear the virus. We also showed under which conditions T-cell killing becomes the primary mechanism for immunosuppression and viral clearance. Furthermore, we characterized robustness properties of T-cell immunity to explain similarities and differences in the control of MV and CDV. Together, our results are consistent with experimental data, advance understanding of control mechanisms of viral clearance across morbilliviruses, and will help inform the development of effective treatments. Further the analysis methods and results have the potential to advance understanding of immune system responses to a range of viral infections such as COVID-19.

6.
J Immunol ; 206(11): 2527-2535, 2021 06 01.
Article in English | MEDLINE | ID: covidwho-1227097

ABSTRACT

The T cell response is an important detection index in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine development. The present study was undertaken to determine the T cell epitopes in the spike (S) protein of SARS-CoV-2 that dominate the T cell responses in SARS-CoV-2-infected patients. PBMCs from rhesus macaques vaccinated with a DNA vaccine encoding the full-length S protein were isolated, and an ELISPOT assay was used to identify the recognized T cell epitopes among a total of 158 18-mer and 10-aa-overlapping peptides spanning the full-length S protein. Six multipeptide-based epitopes located in the S1 region, with four of the six located in the receptor-binding domain, were defined as the most frequently recognized epitopes in macaques. The conservation of the epitopes across species was also verified, and peptide mixtures for T cell response detection were established. Six newly defined T cell epitopes were found in the current study, which may provide a novel potential target for T cell response detection and the diagnosis and vaccine design of SARS-CoV-2 based on multipeptide subunit-based epitopes.


Subject(s)
Epitopes, T-Lymphocyte/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Macaca mulatta
7.
Bioconjug Chem ; 32(5): 1034-1046, 2021 05 19.
Article in English | MEDLINE | ID: covidwho-1217668

ABSTRACT

SARS-CoV-2 caused the COVID-19 pandemic that lasted for more than a year. Globally, there is an urgent need to use safe and effective vaccines for immunization to achieve comprehensive protection against SARS-CoV-2 infection. Focusing on developing a rapid vaccine platform with significant immunogenicity as well as broad and high protection efficiency, we designed a SARS-CoV-2 spike protein receptor-binding domain (RBD) displayed on self-assembled ferritin nanoparticles. In a 293i cells eukaryotic expression system, this candidate vaccine was prepared and purified. After rhesus monkeys are immunized with 20 µg of RBD-ferritin nanoparticles three times, the vaccine can elicit specific humoral immunity and T cell immune response, and the neutralizing antibodies can cross-neutralize four SARS-CoV-2 strains from different sources. In the challenge protection test, after nasal infection with 2 × 105 CCID50 SARS-CoV-2 virus, compared with unimmunized control animals, virus replication in the vaccine-immunized rhesus monkeys was significantly inhibited, and respiratory pathology observations also showed only slight pathological damage. These analyses will benefit the immunization program of the RBD-ferritin nanoparticle vaccine in the clinical trial design and the platform construction to present a specific antigen domain in the self-assembling nanoparticle in a short time to harvest stable, safe, and effective vaccine candidates for new SARS-CoV-2 isolates.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19/prevention & control , Nanoparticles/chemistry , Spike Glycoprotein, Coronavirus/metabolism , T-Lymphocytes/immunology , Animals , Antibodies, Neutralizing/immunology , Binding Sites , CD8-Positive T-Lymphocytes/immunology , COVID-19/immunology , Ferritins/chemistry , Ferritins/metabolism , Immunity, Humoral , Macaca mulatta , Male , Nanoparticles/metabolism , SARS-CoV-2/isolation & purification , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , T-Lymphocytes/metabolism , Ultracentrifugation
8.
JCI Insight ; 6(10)2021 05 24.
Article in English | MEDLINE | ID: covidwho-1197299

ABSTRACT

Emerging coronaviruses from zoonotic reservoirs, including severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), have been associated with human-to-human transmission and significant morbidity and mortality. Here, we study both intradermal and intramuscular 2-dose delivery regimens of an advanced synthetic DNA vaccine candidate encoding a full-length MERS-CoV spike (S) protein, which induced potent binding and neutralizing antibodies as well as cellular immune responses in rhesus macaques. In a MERS-CoV challenge, all immunized rhesus macaques exhibited reduced clinical symptoms, lowered viral lung load, and decreased severity of pathological signs of disease compared with controls. Intradermal vaccination was dose sparing and more effective in this model at protecting animals from disease. The data support the further study of this vaccine for preventing MERS-CoV infection and transmission, including investigation of such vaccines and simplified delivery routes against emerging coronaviruses.


Subject(s)
Coronavirus Infections/veterinary , Macaca mulatta/immunology , Middle East Respiratory Syndrome Coronavirus/immunology , Vaccines, DNA/therapeutic use , Viral Vaccines/therapeutic use , Animals , Coronavirus Infections/immunology , Coronavirus Infections/prevention & control , Immunogenicity, Vaccine , Injections, Intradermal , Middle East Respiratory Syndrome Coronavirus/genetics , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , Vaccines, DNA/administration & dosage , Vaccines, DNA/genetics , Viral Vaccines/administration & dosage , Viral Vaccines/genetics
9.
J Virol ; 2021 Jan 20.
Article in English | MEDLINE | ID: covidwho-1195822

ABSTRACT

Respiratory virus challenge studies involve administration of the challenge virus and sampling to assess for protection from the same anatomical locations. It can therefore be difficult to differentiate actively replicating virus from input challenge virus. For SARS-CoV-2, specific monitoring of actively replicating virus is critical to investigate the protective and therapeutic efficacy of vaccines, monoclonal antibodies, and antiviral drugs. We developed a SARS-CoV-2 subgenomic RNA (sgRNA) RT-PCR assay to differentiate productive infection from inactivated or neutralized virus. Subgenomic RNAs are generated after cell entry and are poorly incorporate into mature virions, and thus may provide a marker for actively replicating virus. We show envelope (E) sgRNA was degraded by RNase in infected cell lysates, while genomic RNA (gRNA) was protected, presumably due to packaging into virions. To investigate the capacity of the sgRNA assay to distinguish input challenge virus from actively replicating virus in vivo, we compared the E sgRNA assay to a standard nucleoprotein (N) or E total RNA assay in convalescent rhesus macaques and in antibody-treated rhesus macaques after experimental SARS-CoV-2 challenge. In both studies, the E sgRNA assay was negative, suggesting protective efficacy, whereas the N and E total RNA assays remained positive. These data suggest the potential utility of sgRNA to monitor actively replicating virus in prophylactic and therapeutic SARS-CoV-2 studies.ImportanceDeveloping therapeutic and prophylactic countermeasures for the SARS-CoV-2 virus is a public health priority. During challenge studies, respiratory viruses are delivered and sampled from the same anatomical location. It is therefore important to distinguish actively replicating virus from input challenge virus. The most common assay for detecting SARS-CoV-2 virus, reverse transcription polymerase chain reaction (RT-PCR) targeting nucleocapsid total RNA, cannot distinguish neutralized input virus from replicating virus. In this study, we assess SARS-CoV-2 subgenomic RNA as a potential measure of replicating virus in rhesus macaques.

10.
Cell Rep Med ; 2(4): 100252, 2021 04 20.
Article in English | MEDLINE | ID: covidwho-1164615

ABSTRACT

The outbreak and spread of SARS-CoV-2 (severe acute respiratory syndrome-coronavirus-2) is a current global health emergency, and effective prophylactic vaccines are needed urgently. The spike glycoprotein of SARS-CoV-2 mediates entry into host cells, and thus is the target of neutralizing antibodies. Here, we show that adjuvanted protein immunization with soluble SARS-CoV-2 spike trimers, stabilized in prefusion conformation, results in potent antibody responses in mice and rhesus macaques, with neutralizing antibody titers exceeding those typically measured in SARS-CoV-2 seropositive humans by more than one order of magnitude. Neutralizing antibody responses were observed after a single dose, with exceptionally high titers achieved after boosting. A follow-up to monitor the waning of the neutralizing antibody responses in rhesus macaques demonstrated durable responses that were maintained at high and stable levels at least 4 months after boosting. These data support the development of adjuvanted SARS-CoV-2 prefusion-stabilized spike protein subunit vaccines.


Subject(s)
Antibodies, Neutralizing/blood , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Viral/blood , COVID-19/prevention & control , COVID-19/veterinary , COVID-19/virology , COVID-19 Vaccines/immunology , Female , Macaca mulatta , Male , /metabolism , Mice , Mice, Inbred C57BL , Protein Domains/immunology , Protein Subunits/immunology , SARS-CoV-2/isolation & purification , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism , Time Factors , Vaccination
11.
Cell Rep Med ; 2(4): 100230, 2021 04 20.
Article in English | MEDLINE | ID: covidwho-1147272

ABSTRACT

The deployment of a vaccine that limits transmission and disease likely will be required to end the coronavirus disease 2019 (COVID-19) pandemic. We recently described the protective activity of an intranasally administered chimpanzee adenovirus-vectored vaccine encoding a pre-fusion stabilized spike (S) protein (ChAd-SARS-CoV-2-S [chimpanzee adenovirus-severe acute respiratory syndrome-coronavirus-2-S]) in the upper and lower respiratory tracts of mice expressing the human angiotensin-converting enzyme 2 (ACE2) receptor. Here, we show the immunogenicity and protective efficacy of this vaccine in non-human primates. Rhesus macaques were immunized with ChAd-Control or ChAd-SARS-CoV-2-S and challenged 1 month later by combined intranasal and intrabronchial routes with SARS-CoV-2. A single intranasal dose of ChAd-SARS-CoV-2-S induces neutralizing antibodies and T cell responses and limits or prevents infection in the upper and lower respiratory tracts after SARS-CoV-2 challenge. As ChAd-SARS-CoV-2-S confers protection in non-human primates, it is a promising candidate for limiting SARS-CoV-2 infection and transmission in humans.

12.
J Virol ; 2021 Mar 10.
Article in English | MEDLINE | ID: covidwho-1127540

ABSTRACT

Vaccines are being rapidly developed with the goal of ending the SARS-CoV-2 pandemic. However, the extent to which SARS-CoV-2 vaccination induces serum responses that cross-react with other coronaviruses remains poorly studied. Here we define serum profiles in rhesus macaques after vaccination with DNA or Ad26 based vaccines expressing SARS-CoV-2 Spike protein followed by SARS-CoV-2 challenge, or SARS-CoV-2 infection alone. Analysis of serum responses showed robust reactivity to the SARS-CoV-2 full-length Spike protein and receptor binding domain (RBD), both included in the vaccine. However, serum cross-reactivity to the closely related sarbecovirus SARS-CoV-1 Spike and RBD, was reduced. Reactivity was also measured to the distantly related common cold alpha-coronavirus, 229E and NL63, and beta-coronavirus, OC43 and HKU1, Spike proteins. Using SARS-COV-2 and SARS-CoV-1 lentivirus based pseudoviruses, we show that neutralizing antibody responses were predominantly SARS-CoV-2 specific. These data define patterns of cross-reactive binding and neutralizing serum responses induced by SARS-CoV-2 infection and vaccination in rhesus macaques. Our observations have important implications for understanding polyclonal responses to SARS-CoV-2 Spike, which will facilitate future CoV vaccine assessment and development.ImportanceThe rapid development and deployment of SARS-CoV-2 vaccines has been unprecedented. In this study, we explore the cross-reactivity of SARS-CoV-2 specific antibody responses to other coronaviruses. By analyzing responses from NHPs both before and after immunization with DNA or Ad26 vectored vaccines, we find patterns of cross reactivity that mirror those induced by SARS-CoV-2 infection. These data highlight the similarities between infection and vaccine induced humoral immunity for SARS-CoV-2 and cross-reactivity of these responses to other CoVs.

13.
J Virol Methods ; 293: 114120, 2021 07.
Article in English | MEDLINE | ID: covidwho-1117217

ABSTRACT

BACKGROUND: Primary rhesus monkey kidney cells (RhMK) can be used for the detection of respiratory viruses, including influenza and parainfluenza. The human colon adeno-carcinoma cell line, CACO-2, has been previously used for the growth of multiple influenza viruses, including seasonal, novel and avian lineages. OBJECTIVE: We compared CACO-2, Madin-Darby Canine Kidney (MDCK), and RhMK cells for the isolation of viruses from patients presenting with influenza like-illness (ILI). STUDY DESIGN: Nasopharyngeal specimens from patients with ILI in primary care settings were processed for conventional viral culture in MDCK, RhMK, and CACO-2. Cells were examined microscopically for cytopathic effect (CPE) and confirmatory testing included immunofluorescent antigen (IFA) detection and real-time RT-PCR. Additionally, 16 specimens positive for respiratory syncytial virus (RSV) by PCR were inoculated on CACO-2 cells. Statistical analysis was done using Chi-square test with IBM Statistical Program. RESULTS: Of 1031 respiratory specimens inoculated, viruses were isolated and confirmed from 331 (32.1 %) in MDCK cells, 304 (29.5 %) in RhMk cells, and 433 (42.0 %) in CACO-2 cells. These included influenza A/(H1N1)pdm09, influenza A(H3N2), influenza B, parainfluenza virus (PIV) types 1, 2, and 3, human coronavirus 229E (CoV-229E), human adenovirus (HAdV), herpes simplex virus 1 (HSV 1), and enterovirus (EV). Influenza A viruses grew best in the CACO-2 cell line. Time to observation of CPE was similar for all three cell types but unlike RhMK and MDCK cells, virus-specific morphological changes were indistinguishable in CACO-2 cells. None of the 16 specimens positive for RSV by PCR grew on CACO-2 cells. CONCLUSIONS: The overall respiratory virus culture isolation rate in CACO-2 cells was significantly higher than that in RhMK or MDCK cells (p < 0.05). CACO-2 cells also supported the growth of some viruses that did not grow in either RhMK or MDCK cells. Except for RSV, CACO-2 cells provide a worthwhile addition to culture algorithms for respiratory specimens.


Subject(s)
Influenza, Human/virology , Nasopharynx/virology , Adenoviruses, Human/growth & development , Adenoviruses, Human/isolation & purification , Adolescent , Adult , Aged , Aged, 80 and over , Animals , Caco-2 Cells , Child , Child, Preschool , Dogs , Female , Humans , Infant , Madin Darby Canine Kidney Cells , Male , Middle Aged , Orthomyxoviridae/growth & development , Orthomyxoviridae/isolation & purification , Respiratory Syncytial Viruses/growth & development , Respiratory Syncytial Viruses/isolation & purification , Young Adult
14.
Nat Commun ; 12(1): 1260, 2021 02 24.
Article in English | MEDLINE | ID: covidwho-1101645

ABSTRACT

A novel coronavirus, SARS-CoV-2, has been identified as the causative agent of the current COVID-19 pandemic. Animal models, and in particular non-human primates, are essential to understand the pathogenesis of emerging diseases and to assess the safety and efficacy of novel vaccines and therapeutics. Here, we show that SARS-CoV-2 replicates in the upper and lower respiratory tract and causes pulmonary lesions in both rhesus and cynomolgus macaques. Immune responses against SARS-CoV-2 are also similar in both species and equivalent to those reported in milder infections and convalescent human patients. This finding is reiterated by our transcriptional analysis of respiratory samples revealing the global response to infection. We describe a new method for lung histopathology scoring that will provide a metric to enable clearer decision making for this key endpoint. In contrast to prior publications, in which rhesus are accepted to be the preferred study species, we provide convincing evidence that both macaque species authentically represent mild to moderate forms of COVID-19 observed in the majority of the human population and both species should be used to evaluate the safety and efficacy of interventions against SARS-CoV-2. Importantly, accessing cynomolgus macaques will greatly alleviate the pressures on current rhesus stocks.


Subject(s)
COVID-19/immunology , COVID-19/virology , Lung/pathology , Lung/virology , Animals , Disease Models, Animal , Female , Immunity, Cellular/physiology , Interferon-gamma/metabolism , Macaca fascicularis , Macaca mulatta , Male , Pandemics , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity
15.
J Biol Chem ; 296: 100435, 2021.
Article in English | MEDLINE | ID: covidwho-1087033

ABSTRACT

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic represents a global threat, and the interaction between the virus and angiotensin-converting enzyme 2 (ACE2), the primary entry receptor for SARS-CoV-2, is a key determinant of the range of hosts that can be infected by the virus. However, the mechanisms underpinning ACE2-mediated viral entry across species remains unclear. Using infection assay, we evaluated SARS-CoV-2 entry mediated by ACE2 of 11 different animal species. We discovered that ACE2 of Rhinolophus sinicus (Chinese rufous horseshoe bat), Felis catus (domestic cat), Canis lupus familiaris (dog), Sus scrofa (wild pig), Capra hircus (goat), and Manis javanica (Malayan pangolin) facilitated SARS-CoV-2 entry into nonsusceptible cells. Moreover, ACE2 of the pangolin also mediated SARS-CoV-2 entry, adding credence to the hypothesis that SARS-CoV-2 may have originated from pangolins. However, the ACE2 proteins of Rhinolophus ferrumequinum (greater horseshoe bat), Gallus gallus (red junglefowl), Notechis scutatus (mainland tiger snake), or Mus musculus (house mouse) did not facilitate SARS-CoV-2 entry. In addition, a natural isoform of the ACE2 protein of Macaca mulatta (rhesus monkey) with the Y217N mutation was resistant to SARS-CoV-2 infection, highlighting the possible impact of this ACE2 mutation on SARS-CoV-2 studies in rhesus monkeys. We further demonstrated that the Y217 residue of ACE2 is a critical determinant for the ability of ACE2 to mediate SARS-CoV-2 entry. Overall, these results clarify that SARS-CoV-2 can use the ACE2 receptors of multiple animal species and show that tracking the natural reservoirs and intermediate hosts of SARS-CoV-2 is complex.


Subject(s)
Angiotensin-Converting Enzyme 2/genetics , COVID-19/epidemiology , COVID-19/transmission , Pandemics , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/genetics , Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/immunology , Animals , COVID-19/diagnosis , COVID-19/immunology , Cats , Chickens/virology , Chiroptera/virology , Dogs , Elapidae/virology , Eutheria/virology , Gene Expression , Goats/virology , HEK293 Cells , Host-Pathogen Interactions/genetics , Host-Pathogen Interactions/immunology , Humans , Immunity, Innate , Macaca mulatta/virology , Mice , Models, Molecular , Mutation , Protein Binding , Protein Structure, Secondary , Recombinant Proteins/genetics , Recombinant Proteins/immunology , SARS-CoV-2/genetics , SARS-CoV-2/immunology , Species Specificity , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Swine/virology , Virus Internalization
16.
Emerg Microbes Infect ; 10(1): 342-355, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1069193

ABSTRACT

The current study aims to develop a safe and highly immunogenic COVID-19 vaccine. The novel combination of a DNA vaccine encoding the full-length Spike (S) protein of SARS-CoV-2 and a recombinant S1 protein vaccine induced high level neutralizing antibody and T cell immune responses in both small and large animal models. More significantly, the co-delivery of DNA and protein components at the same time elicited full protection against intratracheal challenge of SARS-CoV-2 viruses in immunized rhesus macaques. As both DNA and protein vaccines have been proven safe in previous human studies, and DNA vaccines are capable of eliciting germinal center B cell development, which is critical for high-affinity memory B cell responses, the DNA and protein co-delivery vaccine approach has great potential to serve as a safe and effective approach to develop COVID-19 vaccines that provide long-term protection.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19/prevention & control , Spike Glycoprotein, Coronavirus/immunology , Vaccines, DNA/immunology , Vaccines, Subunit/immunology , Animals , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/immunology , Antibodies, Viral/blood , Antibodies, Viral/immunology , Cell Line , DNA/immunology , HEK293 Cells , Humans , Lymphocyte Count , Macaca mulatta , Mice , Mice, Inbred C57BL , Plasmids/genetics , Rabbits , Recombinant Proteins/immunology , SARS-CoV-2/genetics , SARS-CoV-2/immunology , T-Lymphocytes/immunology
17.
PLoS One ; 16(2): e0246366, 2021.
Article in English | MEDLINE | ID: covidwho-1059447

ABSTRACT

Airborne transmission is predicted to be a prevalent route of human exposure with SARS-CoV-2. Aside from African green monkeys, nonhuman primate models that replicate airborne transmission of SARS-CoV-2 have not been investigated. A comparative evaluation of COVID-19 in African green monkeys, rhesus macaques, and cynomolgus macaques following airborne exposure to SARS-CoV-2 was performed to determine critical disease parameters associated with disease progression, and establish correlations between primate and human COVID-19. Respiratory abnormalities and viral shedding were noted for all animals, indicating successful infection. Cynomolgus macaques developed fever, and thrombocytopenia was measured for African green monkeys and rhesus macaques. Type II pneumocyte hyperplasia and alveolar fibrosis were more frequently observed in lung tissue from cynomolgus macaques and African green monkeys. The data indicate that, in addition to African green monkeys, macaques can be successfully infected by airborne SARS-CoV-2, providing viable macaque natural transmission models for medical countermeasure evaluation.


Subject(s)
COVID-19/physiopathology , Disease Models, Animal , Macaca mulatta , SARS-CoV-2/physiology , Animals , COVID-19/pathology , COVID-19/transmission , Chlorocebus aethiops , Disease Transmission, Infectious , Female , Lung/pathology , Macaca fascicularis , Male , Virus Shedding
20.
Nat Commun ; 12(1): 541, 2021 01 22.
Article in English | MEDLINE | ID: covidwho-1044084

ABSTRACT

CD4 T follicular helper (Tfh) cells are important for the generation of durable and specific humoral protection against viral infections. The degree to which SARS-CoV-2 infection generates Tfh cells and stimulates the germinal center (GC) response is an important question as we investigate vaccine induced immunity against COVID-19. Here, we report that SARS-CoV-2 infection in rhesus macaques, either infused with convalescent plasma, normal plasma, or receiving no infusion, resulted in transient accumulation of pro-inflammatory monocytes and proliferating Tfh cells with a Th1 profile in peripheral blood. CD4 helper cell responses skewed predominantly toward a Th1 response in blood, lung, and lymph nodes. SARS-CoV-2 Infection induced GC Tfh cells specific for the SARS-CoV-2 spike and nucleocapsid proteins, and a corresponding early appearance of antiviral serum IgG antibodies. Collectively, the data show induction of GC responses in a rhesus model of mild COVID-19.


Subject(s)
COVID-19/immunology , Germinal Center/immunology , SARS-CoV-2/immunology , T Follicular Helper Cells/immunology , Th1 Cells/immunology , Animals , Antibodies, Viral/blood , COVID-19/therapy , Cell Line , Chlorocebus aethiops , Coronavirus Nucleocapsid Proteins/immunology , Disease Models, Animal , Female , Humans , Immunity, Humoral/immunology , Immunization, Passive , Immunogenicity, Vaccine/immunology , Immunoglobulin G/blood , Macaca mulatta , Male , Phosphoproteins/immunology , SARS-CoV-2/isolation & purification , Spike Glycoprotein, Coronavirus/immunology , Vero Cells
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