Omicron BA.2 breakthrough infection enhances crossneutralization of BA.2.12.1 and BA.4/BA.5

BNT162b2-vaccinated individuals after Omicron BA.1 breakthrough infection have strong serum-neutralizing activity against Omicron BA.1, BA.2, and previous SARS-CoV-2 variants of concern (VOCs) yet less against the highly contagious Omicron sublineages BA.4 and BA.5 that have displaced previous variants. Because the latter sublineages are derived from Omicron BA.2, we characterized serum-neutralizing activity of COVID-19 mRNA vaccine triple-immunized individuals who experienced BA. 2 breakthrough infection. We demonstrate that sera of these individuals have broadly neutralizing activity against previous VOCs and all tested Omicron sublineages, including BA.2-derived variants BA.2.12.1 and BA.4/BA.5. Furthermore, applying antibody depletion, we showed that neutralization of BA.2 and BA.4/BA.5 sublineages by BA.2 convalescent sera is driven to a considerable extent by antibodies targeting the N-terminal domain (NTD) of the spike glycoprotein. However, neutralization by Omicron BA.1 convalescent sera depends exclusively on antibodies targeting the receptor binding domain (RBD). These findings suggest that exposure to Omicron BA.2, in contrast to BA.1 spike glycoprotein, triggers substantial NTD-specific recall responses in vaccinated individuals and thereby enhances the neutralization of BA.4/BA.5 sublineages. Given the current epidemiology with a predominance of BA.2-derived sublineages such as BA.4/BA.5 and rapidly ongoing evolution, these findings helped to inform development of our Omicron BA.4/BA.5-adapted vaccine.

Respiratory viruses in medicolegal autopsies during the winter season 2021/2022: observations after reduction of coronavirus disease-19 (COVID-19) pandemic restrictions.

In the context of the coronavirus disease (COVID-19) pandemic, measures were taken to protect the population from infection. These were almost completely lifted in several countries in the spring of 2022. To obtain an overview of the spectrum of respiratory viruses encountered in autoptical routine case work, and their infectivity, all autopsy cases at the Institute of Legal Medicine in Frankfurt/M. with flu-like symptoms (among others) were examined for at least 16 different viruses via multiplex PCR and cell culture. Out of 24 cases, 10 were virus-positive in PCR: specifically, 8 cases with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), 1 with respiratory syncytial virus (RSV), and 1 with SARS-CoV-2 and the human coronavirus OC43 (HCoV-OC43), as a double infection. The RSV infection and one of the SARS-CoV-2 infections were only detected due to the autopsy. Two SARS-CoV-2 cases (postmortem interval of 8 and 10 days, respectively) showed infectious virus in cell culture; the 6 other cases did not show infectious virus. In the RSV case, virus isolation by cell culture was unsuccessful (Ct value of 23.15 for PCR on cryoconserved lung tissue). HCoV-OC43 was measured as non-infectious in cell culture, with a Ct value of 29.57. The detection of RSV and HCoV-OC43 infections may shed light on the relevance of respiratory viruses other than SARS-CoV-2 in postmortem settings; however, further, more extensive studies are needed for a robust assessment of the hazard potential due to infectious postmortem fluids and tissues in medicolegal autopsy settings.

Humoral and Cellular Immunity against Different Sars-Cov-2 Variants Is Detectable but Reduced in Vaccinated Kidney Transplant Recipients

Solid organ transplant recipients have an increased morbidity and mortality after SARS-CoV-2 infection. Immune responses after the third/fourth vaccination against SARS-CoV-2 are insufficiently studied in patients after kidney transplantation (KTX). We analyzed immune responses at a median of 4 months after the third/fourth vaccination in KTX patients and compared them to healthy controls. Cellular immunity was assessed using interferon-gamma (IFN-gamma) and interleukin-2 (IL-2) ELISpot assays. Neutralizing antibody titers were assessed against SARS-CoV-2 D614G (wild type) and the variants alpha, delta and omicron by a cell culture-based neutralization assay. Humoral immunity was also determined by a competitive fluorescence assay, using 11 different variants of SARS-CoV-2. Antibody ratios were measured by ELISA. KTX patients showed significantly lower SARSCoV- 2-specific IFN-gamma responses after booster vaccination than healthy controls. However, the SARS-CoV-2 specific IL-2 responses were comparable to the T cell responses of healthy controls. Cell culture-based neutralizing antibody titers were 1.3-fold higher in healthy controls for D614G, alpha and delta and 7.8-fold higher for omicron (p < 0.01). Healthy controls had approximately 2-fold higher concentrations of potential neutralizing antibodies against all 11 variants than KTX patients. However, more than 60% of the KTX patients exhibited antibodies to variants of SARS-CoV-2. In conclusion, KTX patients should be partly protected against SARSCoV- 2, either by cross-reactive T cells, especially those producing IL-2, or by neutralizing antibodies to SARSCoV- 2 variants.

Novel SARS-CoV-2 variants induce higher toxicity in cardiovascular cells

Objective: SARS-CoV-2 causes the coronavirus disease 2019 (COVID-19) and has spawned a global health crisis. Virus infection can lead to elevated markers of cardiac injury and inflammation associated with a higher risk of mortality. However, it is so far unclear whether cardiovascular damage is caused by direct virus infection or is mainly secondary due to inflammation. Recently, additional novel SARS-CoV-2 variants have emerged accounting for more than 70% of all cases in Germany. To what extend these variants differ from the original strain in their pathology remains to be elucidated. Here, we investigated the effect of the novel SARS-CoV-2 variants on cardiovascular cells. Results: To study whether cardiovascular cells are permissive for SARSCoV-2, we inoculated human iPS-derived cardiomyocytes and endothelial cells from five different origins, including umbilical vein endothelial cells, coronary artery endothelial cells (HCAEC), cardiac and lung microvascular endothelial cells, or pulmonary arterial cells, in vitro with SARS-CoV-2 isolates (G614 (original strain), B.1.1.7 (British variant), B.1.351 (South African variant) and P.1 (Brazilian variant)). While the original virus strain infected iPS-cardiomyocytes and induced cell toxicity 96h post infection (290±10 cells vs. 130±10 cells;p=0.00045), preliminary data suggest a more severe infection by the novel variants. To what extend the response to the novel variants differ from the original strain is currently investigated by phosphoproteom analysis. Of the five endothelial cells studied, only human coronary artery EC took up the original virus strain, without showing viral replication and cell toxicity. Spike protein was only detected in the perinuclear region and was co-localized with calnexin-positive endosomes, which was accompanied by elevated ER-stress marker genes, such as EDEM1 (1.5±0.2-fold change;p=0.04). Infection with the novel SARS-CoV-2 variants resulted in significant higher levels of viral spike compared to the current strain. Surprisingly, viral up-take was also seen in other endothelial cell types (e.g. HUVEC). Although no viral replication was observed (850±158 viral RNA copies at day 0 vs. 197±43 viral RNA copies at day 3;p=0.01), the British SARS-CoV-2 variant B.1.1.7 reduced endothelial cell numbers (0.63±0.03-fold change;p=0.0001). Conclusion: Endothelial cells and cardiomyocytes showed a distinct response to SARS-CoV-2. Whereas cardiomyocytes were permissively infected, endothelial cells took up the virus, but were resistant to viral replication. However, both cell types showed signs of increased toxicity induced by the British SARS-CoV-2 variant. These data suggest that cardiac complications observed in COVID-19 patients might at least in part be based on direct infection of cardiovascular cells. The more severe cytotoxic effects of the novel variants implicate that patients infected with the new variants should be even more closely monitored.

Detection and infectivity of SARS-CoV-2 in exhumated corpses.

Postmortem detection of severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) after the exhumation of a corpse can become important, e.g. in the case of subsequent medical malpractice allegations. To date, data on possible detection periods [e.g. by reverse transcription polymerase chain reaction (RT-PCR)] or on the potential infectivity of the virus after an exhumation are rare. In the present study, these parameters were examined in two cases with a time span of approximately 4 months between day of death and exhumation. Using SARS-CoV-2 RT-PCR on swabs of both lungs and the oropharynx detection was possible with cycle threshold (Ct) values of about 30 despite signs of beginning decay. RT-PCR testing of perioral and perinasal swabs and swabs collected from the inside of the body bag, taken to estimate the risk of infection of those involved in the exhumation, was negative. Cell culture-based infectivity testing was negative for both, lung and oropharyngeal swabs. In one case, RT-PCR testing at the day of death of an oropharyngeal swab showed almost identical Ct values as postmortem testing of an oropharyngeal swab, impressively demonstrating the stability of viral RNA in the intact corpse. However, favorable climatic conditions in the grave have to be taken into account, as it was wintertime with constant low temperatures. Nevertheless, it was possible to demonstrate successful postmortem detection of SARS-CoV-2 infection following exhumation even after months in an earth grave.

Infectivity of deceased COVID-19 patients.

The duration of infectivity of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) in living patients has been demarcated. In contrast, a possible SARS-CoV-2 infectivity of corpses and subsequently its duration under post mortem circumstances remain to be elucidated. The aim of this study was to investigate the infectivity and its duration of deceased COVID-19 (coronavirus disease) patients. Four SARS-CoV-2 infected deceased patients were subjected to medicolegal autopsy. Post mortem intervals (PMI) of 1, 4, 9 and 17 days, respectively, were documented. During autopsy, swabs and organ samples were taken and examined by RT-qPCR (real-time reverse transcription-polymerase chain reaction) for the detection of SARS-CoV-2 ribonucleic acid (RNA). Determination of infectivity was performed by means of virus isolation in cell culture. In two cases, virus isolation was successful for swabs and tissue samples of the respiratory tract (PMI 4 and 17 days). The two infectious cases showed a shorter duration of COVID-19 until death than the two non-infectious cases (2 and 11 days, respectively, compared to > 19 days), which correlates with studies of living patients, in which infectivity could be narrowed to about 6 days before to 12 days after symptom onset. Most notably, infectivity was still present in one of the COVID-19 corpses after a post-mortem interval of 17 days and despite already visible signs of decomposition. To prevent SARS-CoV-2 infections in all professional groups involved in the handling and examination of COVID-19 corpses, adequate personal safety standards (reducing or avoiding aerosol formation and wearing FFP3 [filtering face piece class 3] masks) have to be enforced for routine procedures.

[The virology of SARS-CoV-2]. / Die Virologie von SARS-CoV-2.

The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has far-reaching effects on society, the economy and medical treatment. Therefore, it is all the more important to understand the characteristics of the virus and to utilize them in the diagnostics, treatment and epidemiology. This article firstly elucidates the medical significance of coronaviruses in general. Then angiotensin-converting enzyme 2 (ACE2) as the binding site of SARS-CoV­2 and the possible influence on disease susceptibility are explained. The gold standard for detection of an active SARS-CoV­2 infection is the direct detection of the pathogen with nucleic acid amplification techniques. At the onset of symptoms a swab of the upper airway is especially suitable due to the high viral load. At a later stage direct detection can be achieved in samples from the lower airway or in stool and anal swabs. Antibody tests cannot replace the direct detection of the pathogen; however, the detection of immunoglobulin G antibodies is of special interest for epidemiological questions (seroconversion time of sometimes several weeks). The plaque reduction neutralization test exclusively detects antibodies which neutralize viruses but the procedure is complicated and can only be carried out in secure laboratories (L3). In addition, the importance of these antibodies with respect to immunity against a second infection is uncertain. Thanks to modern techniques thousands of SARS-CoV­2 sequences are already available, which show a genomic variability. The D614G mutation in the S spikes seems to cause a higher infectiousness. Mutations can impair the diagnostics and treatment, which makes monitoring necessary.

[The virology of SARS-CoV-2]. / Die Virologie von SARS-CoV-2.

The new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has far reaching effects on society, the economy and medical treatment. It is all the more important to understand the characteristics of the virus and to utilize them diagnostically, therapeutically and epidemiologically. This article firstly elucidates the medical importance of coronaviruses in general. Then angiotensin-converting enzyme 2 (ACE2) as the binding site of SARS-CoV­2 and the possible influence on the disease susceptibility are explained. The gold standard for detection of an active SARS-CoV­2 infection is the direct detection of the pathogen with nucleic acid amplification techniques. At the onset of symptoms, a swab of the upper airway is especially suitable due to the high viral burden. At a later stage direct detection can be achieved in samples from the lower airway or a stool or anal swab. Antigen or antibody tests cannot replace the direct detection of the pathogen; however, the detection of immunoglobulin G antibodies are of special interest for epidemiological questions (seroconversion time of sometimes several weeks). The plaque reduction neutralization test exclusively detects antibodies which neutralize viruses but the procedure is complicated. In addition, the importance of these antibodies with respect to immunity against a second infection is uncertain. Thanks to modern techniques thousands of SARS-CoV­2 sequences are already available, which show a genomic variability. The D614G mutation in the S spikes seems to cause a higher infectiosity. Mutations can impair the diagnostics and treatment, which makes monitoring necessary.