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1.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-335222

ABSTRACT

Variant of concern (VOC) Omicron-BA1 has achieved global predominance in early 2022. Therefore, surveillance and comprehensive characterization of Omicron-BA.1 in advanced primary cell culture systems and multiple animal models is urgently needed. Here, we characterized Omicron-BA.1 and recombinant Omicron-BA.1 spike gene mutants in comparison with VOC Delta in well-differentiated primary human nasal and bronchial epithelial cells in vitro, followed by in vivo fitness characterization in naïve hamsters, ferrets and hACE2-expressing mice, and in immunized hACE2-mice. We demonstrate a spike-mediated enhancement of early replication of Omicron-BA.1 in nasal epithelial cultures, but limited replication in bronchial epithelial cultures. In Syrian hamsters, Delta showed dominance over Omicron-BA.1 and in ferrets, Omicron-BA.1 infection was abortive. In mice expressing the authentic hACE2-receptor, Delta and a Delta spike clone also showed dominance over Omicron-BA.1 and an Omicron-BA.1 spike clone, respectively. Interestingly, in naïve K18-hACE2 mice, we observed Delta spike-mediated increased replication and pathogenicity and Omicron-BA.1 spike-mediated reduced replication and pathogenicity, suggesting that the spike gene is a major determinant of both Delta and Omicron-BA.1 replication and pathogenicity. Finally, the Omicron-BA.1 spike clone was less well controlled by mRNA-vaccination in K18-hACE2-mice and became more competitive compared to the progenitor and Delta spike clones, suggesting that spike gene-mediated immune evasion is another important factor that led to Omicron-BA.1 dominance.

2.
Mod Pathol ; 2022 Apr 01.
Article in English | MEDLINE | ID: covidwho-1773954

ABSTRACT

The rate of SARS-CoV-2 infections in vaccinees has become a relevant serious issue. This study aimed to determine the causes of death, histological organ alteration, and viral spread in relation to demographic, clinical-pathological, viral variants, and vaccine types for deceased individuals with proven SARS-CoV-2 infection after vaccination who died between January and November 2021. Twenty-nine consecutively collected cases were analyzed and compared to 141 nonvaccinated control cases. Autopsies were performed on 16 partially and 13 fully vaccinated individuals. Most patients were elderly and suffered from several relevant comorbidities. Real-time RT-PCR (RT-qPCR) identified a significantly increased rate of generalized viral dissemination within organ systems in vaccinated cases versus nonvaccinated cases (45% vs. 16%, respectively; P = 0.008) mainly with Ct-values of higher than 25 in non-respiratory samples. However, vaccinated cases also showed high viral loads, reaching Ct-values below 10, especially in the upper airways and lungs. This was accompanied by high rates of pulmonal bacterial or mycotic superinfections and the occurrence of immunocompromising factors, such as malignancies, immunosuppressive drug intake, or decreased immunoglobulin levels. All these findings were particularly accentuated in partially vaccinated patients compared to fully vaccinated individuals. The virus dissemination observed in our case study may indicate that patients with an impaired immune system have a decreased ability to eliminate the virus. However, the potential role of antibody-dependent enhancement must also be ruled out in future studies. Fatal cases of COVID-19 in vaccinees were rare and often associated with severe comorbidities or other immunosuppressive conditions.

3.
Zoonoses Public Health ; 2022 Mar 02.
Article in English | MEDLINE | ID: covidwho-1723416

ABSTRACT

During the first months of the coronavirus disease (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), cases of human-to-cat transmission were reported. Seroconversion was shown in cats infected under experimental and natural conditions. This large-scale survey of 1,005 serum samples was conducted to investigate anti-SARS-CoV-2 antibody prevalence in domestic cats during the first 7 months of the pandemic in Germany and other European countries. In addition, we compared the sensitivity and specificity of two multispecies SARS-CoV-2 antibody enzyme-linked immunosorbent assays (ELISA). Results were confirmed by using an indirect immunofluorescence test (iIFT) and a surrogate virus neutralization test (sVNT). Sera that were highly positive for feline coronavirus (FCoV) antibodies (n = 103) were included to correct for cross-reactivity of the tests used. Our results showed an overall SARS-CoV-2 seropositivity of 1.9% (n = 19) in a receptor-binding domain (RBD)-based ELISA, additional 0.8% (n = 8) were giving inconclusive results. In contrast, a nucleocapsid-based ELISA revealed 0.5% (n = 5) positive and 0.2% (n = 2) inconclusive results. While the iIFT and sVNT confirmed 100% of positive and 50%-57.1% of the doubtful results as determined in the RBD ELISA, the nucleocapsid-based assay showed a high discrepancy and only one of the five positive results could be confirmed. The results indicate significant deficits of the nucleocapsid-based ELISA with respect to sensitivity and specificity. Due to a significantly higher rate (5.8%) of positive results in the group of highly FCoV antibody-positive samples, cross-reactivity of the FCoV-ELISA with SARS-CoV-2 antibodies cannot be excluded. Furthermore, we investigated the impact of direct contact of domestic cats (n = 23) to SARS-CoV-2 positive owners. Considering one inconclusive result, which got confirmed by iIFT, this exposure did not lead to a significantly higher prevalence (4.4%; p = .358) among tested subjects. Overall, we conclude that cats are a negligible entity with respect to virus transmission in Europe.

4.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-329545

ABSTRACT

Wildlife animals may be susceptible for multiple infectious agents of public health or veterinary relevance, thereby potentially forming a reservoir that bears the constant risk of re-introduction into the human or livestock population. Here, we serologically investigated 493 wild ruminant samples collected in the 2021/22 hunting season in Germany for the presence of antibodies against the severe acute respiratory coronavirus 2 (SARS-CoV-2) and four viruses pathogenic for domestic ruminants, namely the orthobunyavirus Schmallenberg virus (SBV), the reovirus bluetongue virus (BTV) and ruminant pestiviruses like bovine viral diarrhoea virus or border disease virus. The animal species comprised fallow deer, red deer, roe deer, mouflon and wisent. For coronavirus serology, additional 307 fallow, roe and red deer samples collected between 2017 and 2020 at three military training areas were included. While antibodies against SBV could be detected in about 13.6% of the samples collected in 2021/22, only one fallow deer of unknown age tested positive for anti-BTV antibodies and all samples reacted negative for antibodies against ruminant pestiviruses. In an ELISA based on the receptor-binding domain (RBD) of SARS-CoV-2, 25 out of 493 (5.1%) samples collected in autumn and winter 2021/22 scored positive. This sero-reactivity could not be confirmed by the highly specific virus neutralization test, occurred also in 2017, 2018 and 2019, i.e. prior to the human SARS-CoV-2 pandemic, and was likewise observed against the RBD of the related SARS-CoV-1. Therefore, the SARS-CoV-2-seroreactivity was most likely induced by another, hitherto unknown deer virus belonging to the subgenus Sarbecovirus of betacoronaviruses.

5.
PLoS Pathog ; 18(1): e1010161, 2022 01.
Article in English | MEDLINE | ID: covidwho-1703195

ABSTRACT

The global response to Coronavirus Disease 2019 (COVID-19) is now facing new challenges such as vaccine inequity and the emergence of SARS-CoV-2 variants of concern (VOCs). Preclinical models of disease, in particular animal models, are essential to investigate VOC pathogenesis, vaccine correlates of protection and postexposure therapies. Here, we provide an update from the World Health Organization (WHO) COVID-19 modeling expert group (WHO-COM) assembled by WHO, regarding advances in preclinical models. In particular, we discuss how animal model research is playing a key role to evaluate VOC virulence, transmission and immune escape, and how animal models are being refined to recapitulate COVID-19 demographic variables such as comorbidities and age.


Subject(s)
COVID-19/etiology , Disease Models, Animal , SARS-CoV-2 , Age Factors , Animals , COVID-19/prevention & control , COVID-19/therapy , COVID-19 Vaccines/adverse effects , COVID-19 Vaccines/immunology , Comorbidity , Humans , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity
6.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-317008

ABSTRACT

Vector-based SARS-CoV-2 vaccines have been associated with vaccine-induced thrombosis with thrombocytopenia syndrome (VITT/TTS), but the causative factors are still unresolved. We comprehensively analyzed ChAdOx1 nCov-19 (AstraZeneca) and Ad26.COV2.S (Johnson & Johnson). ChAdOx1 nCoV-19 contains significant amounts of host cell protein impurities, including functionally active proteasomes, and adenoviral proteins. In Ad26.COV2.S much less impurities were found. Platelet-factor 4 (PF4) formed complexes with ChAdOx1 nCoV-19 constituents, but not with purified virions from ChAdOx1 nCoV-19 or with Ad26.COV2.S. Vascular hyperpermeability was induced by ChAdOx nCoV-19 but not by Ad26.COV2.S.These differences in impurities together with EDTA-induced capillary leakage might contribute to the higher incidence rate of VITT associated with ChAdOx1 nCoV-19 compared to Ad26.COV2.S.

7.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-311513

ABSTRACT

Background: SARS-CoV-2 vaccine ChAdOx1 nCov-19 rarely causes vaccine-induced immune thrombotic thrombocytopenia (VITT) that—like autoimmune heparin-induced thrombocytopenia—is mediated by platelet-activating anti-platelet factor 4 (PF4) antibodies. Methods: We investigated vaccine, PF4, and VITT patient-derived anti-PF4 antibody interactions using dynamic light scattering, 3D-super-resolution microscopy, and electron microscopy. Mass spectrometry was used to analyze vaccine composition. We investigated the mechanism for early post-vaccine inflammatory reactions as potential co-stimulant for anti-PF4 immune response. Finally, we evaluated VITT antibodies for inducing release of procoagulant DNA-containing neutrophil extracellular traps (NETs), and measured DNase activity in VITT patient serum. Results: Biophysical analyses showed formation of complexes between PF4 and vaccine constituents, including virus proteins that were recognized by VITT antibodies. EDTA, a vaccine constituent, increased microvascular leakage in mice allowing for circulation of virus- and virus-producing cell culture-derived proteins. Antibodies in normal sera cross-reacted with human proteins in the vaccine and likely contribute to commonly observed acute ChAdOx1 nCov-19 post-vaccination inflammatory reactions. Polyphosphates and DNA enhanced PF4-dependent platelet activation by VITT antibodies. In the presence of platelets, PF4 enhanced VITT antibody-driven procoagulant NETs formation, while DNase activity was reduced in VITT sera, with granulocyte-rich cerebral vein thrombosis observed in a VITT patient. Conclusions: ChAdOx1 nCoV-19 vaccine constituents (i) form antigenic complexes with PF4, (ii) EDTA increases microvascular permeability, and (iii) vaccine components cause acute inflammatory reactions. Antigen formation in a proinflammatory milieu offers an explanation for anti-PF4 antibody production. High-titer anti-PF4 antibodies activate platelets and induce neutrophil activation and NETs formation, fueling the VITT prothrombotic response.

8.
Haematologica ; 107(4): 947-957, 2022 Apr 01.
Article in English | MEDLINE | ID: covidwho-1635447

ABSTRACT

Vector-based SARS-CoV-2 vaccines have been associated with vaccine- induced thrombosis with thrombocytopenia syndrome (VITT/TTS), but the causative factors are still unresolved. We comprehensively analyzed the ChAdOx1 nCoV-19 (AstraZeneca) and Ad26.COV2.S (Johnson and Johnson) vaccines. ChAdOx1 nCoV-19 contains significant amounts of host cell protein impurities, including functionally active proteasomes, and adenoviral proteins. A much smaller amount of impurities was found in Ad26.COV2.S. Platelet factor 4 formed complexes with ChAdOx1 nCoV-19 constituents, but not with purified virions from ChAdOx1 nCoV-19 or with Ad26.COV2.S. Vascular hyperpermeability was induced by ChAdOx nCoV-19 but not by Ad26.COV2.S. These differences in impurities together with EDTAinduced capillary leakage might contribute to the higher incidence rate of VITT associated with ChAdOx1 nCoV-19 compared to Ad26.COV2.S.


Subject(s)
COVID-19 , Vaccines , COVID-19 Vaccines/adverse effects , Humans , SARS-CoV-2
9.
Nature ; 602(7896): 307-313, 2022 02.
Article in English | MEDLINE | ID: covidwho-1585832

ABSTRACT

Emerging variants of concern (VOCs) are driving the COVID-19 pandemic1,2. Experimental assessments of replication and transmission of major VOCs and progenitors are needed to understand the mechanisms of replication and transmission of VOCs3. Here we show that the spike protein (S) from Alpha (also known as B.1.1.7) and Beta (B.1.351) VOCs had a greater affinity towards the human angiotensin-converting enzyme 2 (ACE2) receptor than that of the progenitor variant S(D614G) in vitro. Progenitor variant virus expressing S(D614G) (wt-S614G) and the Alpha variant showed similar replication kinetics in human nasal airway epithelial cultures, whereas the Beta variant was outcompeted by both. In vivo, competition experiments showed a clear fitness advantage of Alpha over wt-S614G in ferrets and two mouse models-the substitutions in S were major drivers of the fitness advantage. In hamsters, which support high viral replication levels, Alpha and wt-S614G showed similar fitness. By contrast, Beta was outcompeted by Alpha and wt-S614G in hamsters and in mice expressing human ACE2. Our study highlights the importance of using multiple models to characterize fitness of VOCs and demonstrates that Alpha is adapted for replication in the upper respiratory tract and shows enhanced transmission in vivo in restrictive models, whereas Beta does not overcome Alpha or wt-S614G in naive animals.


Subject(s)
COVID-19/transmission , COVID-19/virology , Mutation , SARS-CoV-2/classification , SARS-CoV-2/physiology , Virus Replication , Amino Acid Substitution , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Animals , Animals, Laboratory/virology , COVID-19/veterinary , Cricetinae , Disease Models, Animal , Epithelial Cells/virology , Female , Ferrets/virology , Humans , Male , Mesocricetus/virology , Mice , Mice, Transgenic , SARS-CoV-2/genetics , SARS-CoV-2/growth & development , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , Virulence/genetics
10.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-294937

ABSTRACT

Background: The rate of SARS-CoV-2 breakthrough infections in vaccinees is becoming an increasingly serious issue. Objective: To determine the causes of death, histological organ alteration, and viral spread in relation to demographic, clinical-pathological, viral variants, and vaccine types. Design: Comprehensive retrospective observational cohort study. Setting: Consecutive cases from four German academic medical centers. Patients: Deceased with proven SARS-CoV-2 infection after vaccination who died between January and November 2021. Collections of 29 vaccinees which were analyzed and compared to 141 nonvaccinated control cases. Results: Autopsies were performed on 16 partially and 13 fully vaccinated individuals. Most patients were elderly and suffered from several relevant comorbidities. Real-time RT-PCR (RT-qPCR) identified a significantly increased rate of generalized viral dissemination within the organism in vaccinated cases versus nonvaccinated cases (45% vs. 16%, respectively;P = 0.008). Vaccinated cases also showed high viral loads, reaching Ct values below 10, especially in the upper airways and lungs. This was accompanied by high rates of pulmonal bacterial or mycotic superinfections and the occurrence of immunocompromising factors such as malignancies, immunosuppressive drug intake, or decreased immunoglobulin levels. All these findings were particularly accentuated in partially vaccinated patients compared to fully vaccinated individuals. A fatal course after vaccination occurred in only 14% of all COVID-19 deceased in Augsburg. Limitations: Restricted number of cases Conclusions: Fatal cases of COVID-19 in vaccinees were rare and often associated with severe comorbidities or other immunosuppressive conditions. Interestingly, we observed striking virus dissemination in our case study, which may indicate a decreased ability to eliminate the virus in patients with an impaired immune system. However, the potential role of antibody-dependent enhancement must also be ruled out in future studies.

11.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-293969

ABSTRACT

Background: A novel zoonotic SARS-related coronavirus emerged in China at the end of 2019. The novel SARS-CoV-2 became pandemic within weeks and the number of human infections and severe cases is increasing. The role of potential animal hosts is still understudied.Methods: We intranasally inoculated fruit bats (Rousettus aegyptiacus;n=9), ferrets (n=9), pigs (n=9) and chickens (n=17) with 105 TCID50 of a SARS-CoV-2 isolate per animal. Animals were monitored clinically and for virus shedding. Direct contact animals (n=3) were included. Animals were humanely sacrificed for virological and immune-pathohistological analysis at different time points.Findings: Under these settings, pigs and chickens were not susceptible to SARS-CoV-2. All swabs as well as organ samples and contact animals remained negative for viral RNA, and none of the animals seroconverted. Rousettus aegyptiacus fruit bats experienced a transient infection, with virus detectable by RT-qPCR, immunohistochemistry (IHC) and in situ hybridization (ISH) in the nasal cavity, associated with rhinitis. Viral RNA was also identified in the trachea, lung and lung associated lymphatic tissue. One of three contact bats became infected. More efficient virus replication but no clinical signs were observed in ferrets with transmission to all direct contact animals. Prominent viral RNA loads of up to 104 viral genome copies/ml were detected in the upper respiratory tract. Mild rhinitis was associated with viral antigen detection in the respiratory and olfactory epithelium. Both fruit bats and ferrets developed SARS-CoV-2 reactive antibodies reaching neutralizing titers of up to 1:1024.Interpretation: Pigs and chickens could not be infected intranasally by SARS-CoV-2, whereas fruit bats showed characteristics of a reservoir host. Virus replication in ferrets resembled a subclinical human infection with efficient spread. These animals might serve as a useful model for further studies e.g. testing vaccines or antivirals.Funding Statement: Intramural funding of the German Federal Ministry of Food and Agriculture provided to the Friedrich-Loeffler-Institut.Declaration of Interests: All authors declare no competing interest.Ethics Approval Statement: The animal experiments were evaluated and approved by the ethics committee of the State Office of Agriculture, Food safety, and Fishery in Mecklenburg – Western Pomerania (LALLF M-V: LVL MV/TSD/7221.3-2-010/18-12). All procedures were carried out in approved biosafety level 3 (BSL3) facilities.

12.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-293536

ABSTRACT

Background: The rate of SARS-CoV-2 breakthrough infections in vaccinees is becoming an increasingly serious issue. Objective: To determine the causes of death, histological organ alteration, and viral spread in relation to demographic, clinical-pathological, viral variants, and vaccine types. Design: Comprehensive retrospective observational cohort study. Setting: Consecutive cases from four German academic medical centers. Patients: Deceased with proven SARS-CoV-2 infection after vaccination who died between January and November 2021. Collections of 29 vaccinees which were analyzed and compared to 141 nonvaccinated control cases. Results: Autopsies were performed on 16 partially and 13 fully vaccinated individuals. Most patients were elderly and suffered from several relevant comorbidities. Real-time RT-PCR (RT-qPCR) identified a significantly increased rate of generalized viral dissemination within the organism in vaccinated cases versus nonvaccinated cases (45% vs. 16%, respectively;P = 0.008). Vaccinated cases also showed high viral loads, reaching Ct values below 10, especially in the upper airways and lungs. This was accompanied by high rates of pulmonal bacterial or mycotic superinfections and the occurrence of immunocompromising factors such as malignancies, immunosuppressive drug intake, or decreased immunoglobulin levels. All these findings were particularly accentuated in partially vaccinated patients compared to fully vaccinated individuals. A fatal course after vaccination occurred in only 14% of all COVID-19 deceased in Augsburg. Limitations: Restricted number of cases Conclusions: Fatal cases of COVID-19 in vaccinees were rare and often associated with severe comorbidities or other immunosuppressive conditions. Interestingly, we observed striking virus dissemination in our case study, which may indicate a decreased ability to eliminate the virus in patients with an impaired immune system. However, the potential role of antibody-dependent enhancement must also be ruled out in future studies.

13.
Diseases ; 9(4)2021 Nov 15.
Article in English | MEDLINE | ID: covidwho-1523903

ABSTRACT

Since the beginning of 2020, the betacoronavirus SARS-CoV-2 is causing a global pandemic of an acute respiratory disease termed COVID-19. The diagnostics of the novel disease is primarily based on direct virus detection by RT-PCR; however, the availability of test kits may become a major bottleneck, when millions of tests are performed per week. To increase the flexibility of SARS-CoV-2 diagnostics, three real-time RT-PCR assays listed on the homepage of the World Health Organization were selected and investigated regarding their compatibility with three different RT-PCR kits. Furthermore, the reaction volume of the PCR chemistry was reduced up to half of the original protocol to make the individual reactions more cost- and resource-effective. When testing dilution series of culture-grown virus, nearly identical quantification cycle values (Cq) were obtained for all RT-PCR assay/chemistry combinations. Regarding the SARS-CoV-2 detection in clinical samples, agreeing results were obtained for all combinations for virus negative specimens and swabs containing high to medium viral genome loads. In cases of very low SARS-CoV-2 genome loads (Cq > 36), inconsistent results were observed, with some test runs scoring negative and some positive. However, no preference of a specific target within the viral genome (E, RdRp, or N) or of a certain chemistry was seen. In summary, a reduction of the reaction volume and the type of PCR chemistry did not influence the PCR sensitivity.

14.
Blood ; 138(22): 2256-2268, 2021 12 02.
Article in English | MEDLINE | ID: covidwho-1443788

ABSTRACT

SARS-CoV-2 vaccine ChAdOx1 nCoV-19 (AstraZeneca) causes a thromboembolic complication termed vaccine-induced immune thrombotic thrombocytopenia (VITT). Using biophysical techniques, mouse models, and analysis of VITT patient samples, we identified determinants of this vaccine-induced adverse reaction. Super-resolution microscopy visualized vaccine components forming antigenic complexes with platelet factor 4 (PF4) on platelet surfaces to which anti-PF4 antibodies obtained from VITT patients bound. PF4/vaccine complex formation was charge-driven and increased by addition of DNA. Proteomics identified substantial amounts of virus production-derived T-REx HEK293 proteins in the ethylenediaminetetraacetic acid (EDTA)-containing vaccine. Injected vaccine increased vascular leakage in mice, leading to systemic dissemination of vaccine components known to stimulate immune responses. Together, PF4/vaccine complex formation and the vaccine-stimulated proinflammatory milieu trigger a pronounced B-cell response that results in the formation of high-avidity anti-PF4 antibodies in VITT patients. The resulting high-titer anti-PF4 antibodies potently activated platelets in the presence of PF4 or DNA and polyphosphate polyanions. Anti-PF4 VITT patient antibodies also stimulated neutrophils to release neutrophil extracellular traps (NETs) in a platelet PF4-dependent manner. Biomarkers of procoagulant NETs were elevated in VITT patient serum, and NETs were visualized in abundance by immunohistochemistry in cerebral vein thrombi obtained from VITT patients. Together, vaccine-induced PF4/adenovirus aggregates and proinflammatory reactions stimulate pathologic anti-PF4 antibody production that drives thrombosis in VITT. The data support a 2-step mechanism underlying VITT that resembles the pathogenesis of (autoimmune) heparin-induced thrombocytopenia.


Subject(s)
Antigen-Antibody Complex/immunology , Autoantibodies/immunology , COVID-19/prevention & control , Capsid Proteins/adverse effects , Drug Contamination , Genetic Vectors/adverse effects , HEK293 Cells/immunology , Immunoglobulin G/immunology , Platelet Factor 4/immunology , Purpura, Thrombocytopenic, Idiopathic/etiology , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/adverse effects , Adenoviridae/immunology , Animals , Antigen-Antibody Complex/ultrastructure , Autoantibodies/biosynthesis , Capillary Leak Syndrome/etiology , Capsid Proteins/immunology , Cell Line, Transformed , /immunology , Dynamic Light Scattering , Epitopes/chemistry , Epitopes/immunology , Extracellular Traps/immunology , Extravasation of Diagnostic and Therapeutic Materials/etiology , Genetic Vectors/immunology , HEK293 Cells/chemistry , Humans , Imaging, Three-Dimensional , Immunoglobulin G/biosynthesis , Inflammation , Mice , Microscopy/methods , Platelet Activation , Proteomics , Purpura, Thrombocytopenic, Idiopathic/blood , Purpura, Thrombocytopenic, Idiopathic/immunology , Sinus Thrombosis, Intracranial/diagnostic imaging , Sinus Thrombosis, Intracranial/immunology , Spike Glycoprotein, Coronavirus/immunology , Virus Cultivation
15.
Res Vet Sci ; 140: 229-232, 2021 Nov.
Article in English | MEDLINE | ID: covidwho-1401839

ABSTRACT

Several non-variant of concern SARS-CoV-2 infections in pets have been reported as documented in the OIE and GISAID databases and there is only one fully documented case of an alpha variant of concern (VOC)(B.1.1.7) in the United States so far. Here, we describe the first case in a cat infected with the alpha SARS-CoV-2 variant in Germany. A cat suffering from pneumonia was presented to a veterinary practice. The pneumonia was treated symptomatically, but 16 days later the cat was presented again. Since the owner had been tested positive for a SARS-CoV-2 infection in the meantime, swab samples were taken from the cat and analyzed for SARS-CoV-2 specific nucleic acids. The various RT-qPCR analyses and whole-genome sequencing revealed the presence of the SARS-CoV-2 B.1.1.7 variant in this cat. This study shows that pets living in close contact with SARS-CoV-2 B.1.1.7 infected owners can contract this virus and also suffer from a respiratory disease. It is not clear yet whether onward transmissions to other cats and humans can occur. To minimize transmission risks, pet owners and veterinarians should comply to the hygienic rules published by OIE and others. It must be stated, that infections of cats with SARS-CoV-2 is still a rare event. Cats with clinical signs of a respiratory disease should be presented to a veterinarian, who will decide on further steps.


Subject(s)
COVID-19 , Cat Diseases , Animals , COVID-19/veterinary , Cat Diseases/diagnosis , Cats , Germany , Humans , Real-Time Polymerase Chain Reaction/veterinary , SARS-CoV-2
16.
Viruses ; 13(8)2021 08 19.
Article in English | MEDLINE | ID: covidwho-1367921

ABSTRACT

The recent emergence of SARS-CoV-2 in humans from a yet unidentified animal reservoir and the capacity of the virus to naturally infect pets, farmed animals and potentially wild animals has highlighted the need for serological surveillance tools. In this study, the luciferase immunoprecipitation systems (LIPS), employing the spike (S) and nucleocapsid proteins (N) of SARS-CoV-2, was used to examine the suitability of the assay for antibody detection in different animal species. Sera from SARS-CoV-2 naturally-infected mink (n = 77), SARS-CoV-2 experimentally-infected ferrets, fruit bats and hamsters and a rabbit vaccinated with a purified spike protein were examined for antibodies using the SARS-CoV-2 N and/or S proteins. From comparison with the known neutralization status of the serum samples, statistical analyses including calculation of the Spearman rank-order-correlation coefficient and Cohen's kappa agreement were used to interpret the antibody results and diagnostic performance. The LIPS immunoassay robustly detected the presence of viral antibodies in naturally infected SARS-CoV-2 mink, experimentally infected ferrets, fruit bats and hamsters as well as in an immunized rabbit. For the SARS-CoV-2-LIPS-S assay, there was a good level of discrimination between the positive and negative samples for each of the five species tested with 100% agreement with the virus neutralization results. In contrast, the SARS-CoV-2-LIPS-N assay did not consistently differentiate between SARS-CoV-2 positive and negative sera. This study demonstrates the suitability of the SARS-CoV-2-LIPS-S assay for the sero-surveillance of SARS-CoV-2 infection in a range of animal species.


Subject(s)
Antibodies, Viral/blood , COVID-19/veterinary , Mink/immunology , SARS-CoV-2/immunology , Animals , COVID-19/diagnosis , COVID-19/epidemiology , COVID-19/immunology , COVID-19 Serological Testing , Chiroptera/immunology , Coronavirus Nucleocapsid Proteins/immunology , Epidemiological Monitoring , Ferrets/immunology , Immunoprecipitation , Mesocricetus/immunology , Phosphoproteins/immunology , Rabbits/immunology , Seroepidemiologic Studies , Spike Glycoprotein, Coronavirus/immunology
17.
Diagn Microbiol Infect Dis ; 101(4): 115520, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1363963

ABSTRACT

Sample panels of SARS-CoV-2 cases were retrospectively whole-genome sequenced. In three individuals, samples of upper and lower respiratory tract resulted in identical sequences suggesting virus stability including the spike protein cleavage site. In a fourth case, low-level intra-host genomic evolution and a unique 5-nucleotide deletion was observed.


Subject(s)
Adaptation, Physiological/genetics , COVID-19/virology , Respiratory System/virology , SARS-CoV-2/isolation & purification , Whole Genome Sequencing , Genome, Viral , Humans , Retrospective Studies , Tissue Distribution
18.
PLoS One ; 16(7): e0254872, 2021.
Article in English | MEDLINE | ID: covidwho-1317145

ABSTRACT

BACKGROUND: COVID-19 is only partly understood, and the level of evidence available in terms of pathophysiology, epidemiology, therapy, and long-term outcome remains limited. During the early phase of the pandemic, it was necessary to effectively investigate all aspects of this new disease. Autopsy can be a valuable procedure to investigate the internal organs with special techniques to obtain information on the disease, especially the distribution and type of organ involvement. METHODS: During the first wave of COVID-19 in Germany, autopsies of 19 deceased patients were performed. Besides gross examination, the organs were analyzed with standard histology and polymerase-chain-reaction for SARS-CoV-2. Polymerase chain reaction positive localizations were further analyzed with immunohistochemistry and RNA-in situ hybridization for SARS-CoV-2. RESULTS: Eighteen of 19 patients were found to have died due to COVID-19. Clinically relevant histological changes were only observed in the lungs. Diffuse alveolar damage in considerably different degrees was noted in 18 cases. Other organs, including the central nervous system, did not show specific micromorphological alterations. In terms of SARS-CoV-2 detection, the focus remains on the upper airways and lungs. This is true for both the number of positive samples and the viral load. A highly significant inverse correlation between the stage of diffuse alveolar damage and viral load was found on a case and a sample basis. Mediastinal lymph nodes and fat were also affected by the virus at high frequencies. By contrast, other organs rarely exhibited a viral infection. Moderate to strong correlations between the methods for detecting SARS-CoV-2 were observed for the lungs and for other organs. CONCLUSIONS: The lung is the most affected organ in gross examination, histology and polymerase chain reaction. SARS-CoV-2 detection in other organs did not reveal relevant or specific histological changes. Moreover, we did not find CNS involvement.


Subject(s)
COVID-19/virology , Central Nervous System/virology , Lung/virology , Lymph Nodes/virology , Viral Load , Aged , Aged, 80 and over , Autopsy/statistics & numerical data , COVID-19/epidemiology , COVID-19/pathology , Central Nervous System/pathology , Female , Humans , Lung/pathology , Lymph Nodes/pathology , Male , Middle Aged
19.
Nat Commun ; 12(1): 4048, 2021 06 30.
Article in English | MEDLINE | ID: covidwho-1290662

ABSTRACT

The ongoing SARS-CoV-2 pandemic necessitates the fast development of vaccines. Recently, viral mutants termed variants of concern (VOC) which may escape host immunity have emerged. The efficacy of spike encoding mRNA vaccines (CVnCoV and CV2CoV) against the ancestral strain and the VOC B.1.351 was tested in a K18-hACE2 transgenic mouse model. Naive mice and mice immunized with a formalin-inactivated SARS-CoV-2 preparation were used as controls. mRNA-immunized mice develop elevated SARS-CoV-2 RBD-specific antibody and neutralization titers which are readily detectable, but significantly reduced against VOC B.1.351. The mRNA vaccines fully protect from disease and mortality caused by either viral strain. SARS-CoV-2 remains undetected in swabs, lung, or brain in these groups. Despite lower neutralizing antibody titers compared to the ancestral strain BavPat1, CVnCoV and CV2CoV show complete disease protection against the novel VOC B.1.351 in our studies.


Subject(s)
Antibodies, Neutralizing/blood , Antibodies, Viral/blood , COVID-19 Vaccines/immunology , COVID-19/prevention & control , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/immunology , Animals , Cell Line , Chlorocebus aethiops , Genome, Viral/genetics , Humans , Mice , Mice, Transgenic , SARS-CoV-2/genetics , Vero Cells
20.
Vaccines (Basel) ; 9(6)2021 Jun 15.
Article in English | MEDLINE | ID: covidwho-1282658

ABSTRACT

Emerging infectious diseases represent an increasing threat to human and animal health. Therefore, safe and effective vaccines that could be available within a short time frame after an outbreak are required for adequate prevention and control. Here, we developed a robust and versatile self-assembling multimeric protein scaffold particle (MPSP) vaccine platform using lumazine synthase (LS) from Aquifex aeolicus. This scaffold allowed the presentation of peptide epitopes by genetic fusion as well as the presentation of large antigens by bacterial superglue-based conjugation to the pre-assembled particle. Using the orthobunyavirus model Schmallenberg virus (SBV) we designed MPSPs presenting major immunogens of SBV and assessed their efficacy in a mouse model as well as in cattle, a target species of SBV. All prototype vaccines conferred protection from viral challenge infection and the multivalent presentation of the selected antigens on the MPSP markedly improved their immunogenicity compared to the monomeric subunits. Even a single shot vaccination protected about 80% of mice from an otherwise lethal dose of SBV. Most importantly, the MPSPs induced a virtually sterile immunity in cattle. Altogether, LS represents a promising platform for modular and rapid vaccine design.

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