Determinants of species-specific utilization of ACE2 by human and animal coronaviruses (preprint)

Utilization of human ACE2 allowed several bat coronaviruses (CoVs), including the causative agent of COVID-19, to infect humans either directly or via intermediate hosts. Here, we analyzed the ability of Spike proteins from 24 human or animal CoVs to use ACE2 receptors across nine reservoir, potential intermediate and human hosts. We show that overall SARS-CoV-2 Omicron variants evolved more efficient ACE2 usage but mutation of R493Q in BA.5 Spike disrupts utilization of ACE2 from Greater horseshoe bats. Spikes from most CoVs showed species-specific differences in ACE2 usage, partly due to variations in ACE2 residues 31, 41 or 354. Mutation of T403R allowed the RaTG13 bat CoV Spike to use all ACE2 orthologs analysed for viral entry. Sera from COVID-19 vaccinated individuals neutralized the Spike proteins of a range of bat Sarbecoviruses. Our results define determinants of ACE2 receptor usage of diverse CoVs and suggest that COVID-19 vaccination may protect against future zoonoses of SARS-CoV-related bat viruses.

Heterologous ChAdOx1 nCoV-19 and BNT162b2 prime-boost vaccination elicits potent neutralizing antibody responses and T cell reactivity (preprint)

Background Heterologous prime-boost schedules with vector- and mRNA-based COVID-19 vaccines are already administered, but immunological responses and elicited protection have not been reported. Methods We here analyzed a cohort of 26 individuals aged 25-46 (median 30.5) years that received a ChAdOx1 nCoV-2019 prime followed by a BNT162b2 boost after an 8-week interval for reactogenicity, antibody responses and T cell reactivity. Results Self-reported solicited symptoms after ChAdOx1 nCoV-2019 prime were in line with previous reports and less severe after the BNT162b2 boost. Antibody titers increased significantly over time resulting in strong neutralization titers 2 weeks after the BNT162b2 boost. Neutralizing activity against the prevalent strain B.1.1.7 was 3.9-fold higher than in individuals receiving homologous BNT162b2 vaccination, only 2-fold reduced for variant of concern B.1.351, and similar for variant B.1.617. In addition, CD4+ and CD8+ T cells reacted to SARS-CoV-2 spike peptide stimulus 2 weeks after the full vaccination. Conclusions The heterologous ChAdOx1 nCoV-2019 / BNT162b2 prime-boost vaccination regimen is not associated with serious adverse events and results in a potent humoral immune response and elicits T cell reactivity. Variants of concern B.1.1.7, B.1.351 and B.1.617 are potently neutralized by sera of all participants. These results suggest that this heterologous vaccination regimen is at least as immunogenic and protective as homologous vaccinations.

COVID-19 risk haplogroups differ between populations, deviate from Neanderthal haplotypes and compromise risk assessment in non-Europeans (preprint)

Recent genome wide association studies (GWAS) have identified genetic risk factors for developing severe COVID-19 symptoms. The studies reported a 1bp insertion rs11385942 on chromosome 3 and furthermore two single nucleotide variants (SNVs) rs35044562 and rs67959919, all three correlated with each other. Zeberg and Paabo subsequently traced them back to Neanderthal origin. They found that a 49.4 kb genomic region including the risk allele of rs35044562 is inherited from Neanderthals of Vindija in Croatia. Here we add a differently focused evaluation of this major genetic risk factor to these recent analyses. We show that (i) COVID-19-related genetic factors of Neanderthals deviate from those of modern humans and that (ii) they differ among world-wide human populations, which compromises risk prediction in non-Europeans. Currently, caution is thus advised in the genetic risk assessment of non-Europeans during this world-wide COVID-19 pandemic.

Antiviral activity of plant juices and green tea against SARS-CoV-2 and influenza virus in vitro (preprint)

Respiratory viruses initially infect the naso- and oropharyngeal regions, where they amplify, cause symptoms and may also be transmitted to new hosts. Preventing initial infection or reducing viral loads upon infection might soothe symptoms, prevent dissemination into the lower airways, or transmission to the next individual. We here analyzed the potential of plant derived products to inactivate SARS-CoV-2 and influenza virus. We found that black chokeberry (Aronia melanocarpa) juice, pomegranate (Punica granatum) juice, and green tea (Camellia sinensis) have virucidal activity against both viruses, suggesting that oral rinsing may reduce viral loads in the oral cavity thereby lowering virus transmission.

Potential Opportunity Of Antisense Therapy Of COVID-19 on an in vitro model (preprint)

Data on potential effectiveness and prospects of treatment of new coronavirus infection of COVID-19 caused by virus SARS-CoV-2 with the help of antisense oligonucleotides acting against RNA of virus on an in vitro model are given. The ability of antisense oligonucleotides to suppress viral replication in diseases caused by coronaviruses using the example of SARS and MERS is shown. The identity of the initial regulatory section of RNA of various coronaviruses was found within 50 - 100 nucleotides from the 5'-end, which allows using antisense suppression of this RNA fragment. A new RNA fragment of the virus present in all samples of coronovirus SARS-CoV-2 has been identified, the suppression of which with the help of an antisense oligonucleotide can be effective in the treatment of COVID-19. The study of the synthesized antisense oligonucleotide 5`-AGCCGAGTGACAGCC ACACAG, complementary to the selected virus RNA sequence, was carried out. The low toxicity of the preparations of this group in the cell culture study and the ability to reduce viral load at high doses according to real time-PCR data are shown. The cytopathogenic dose exceeds 2 mg/ml. At a dosage of 1 mg/ml, viral replication is reduced by 5 - 13 times. Conclusions are made about the prospects of this direction and the feasibility of using the inhalation way of drug administration into the body.

IFITM proteins promote SARS-CoV-2 infection in human lung cells (preprint)

Interferon-induced transmembrane proteins (IFITMs 1, 2 and 3) restrict numerous viral pathogens and are thought to prevent infection by severe acute respiratory syndrome coronaviruses (SARS-CoVs). However, most evidence comes from single-round pseudoparticle infection of cells artificially overexpressing IFITMs. Here, we confirmed that overexpression of IFITMs blocks pseudoparticle infections mediated by the Spike proteins of {beta}-coronaviruses including pandemic SARS-CoV-2. In striking contrast, however, endogenous IFITM expression promoted genuine SARS-CoV-2 infection in human lung cells both in the presence and absence of interferon. IFITM2 was most critical for efficient entry of SARS-CoV-2 and enhanced virus production from Calu-3 cells by several orders of magnitude. IFITMs are expressed and further induced by interferons in the lung representing the primary site of SARS-CoV-2 infection as well as in other relevant tissues. Our finding that IFITMs enhance SARS-CoV-2 infection under conditions approximating the in vivo situation shows that they may promote viral invasion during COVID-19. HIGHLIGHTSO_LIOverexpression of IFITM1, 2 and 3 restricts SARS-CoV-2 infection C_LIO_LIEndogenous IFITM1, 2 and 3 boost SARS-CoV-2 infection of human lung cells C_LIO_LIIFITM2 is critical for efficient entry of SARS-CoV-2 in Calu-3 cells C_LI

Alpha-1 antitrypsin inhibits SARS-CoV-2 infection (preprint)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19). To identify factors of the respiratory tract that suppress SARS-CoV-2, we screened a peptide/protein library derived from bronchoalveolar lavage, and identified 1-antitrypsin (1-AT) as specific inhibitor of SARS-CoV-2. 1-AT targets the viral spike protein and blocks SARS-CoV-2 infection of human airway epithelium at physiological concentrations. Our findings show that endogenous 1-AT restricts SARS-CoV-2 and repurposes 1-AT-based drugs for COVID-19 therapy.

Holder Pasteurization Inactivates SARS-CoV-2 in Human Breast Milk (preprint)

SARS-CoV-2 RNA has been detected in the human breast milk of infected mothers, raising concerns regarding the safety of breastfeeding upon infection. We here show that holder pasteurization inactivates SARS-CoV-2 and provides an alternative and safe option for infected mothers to continue feeding breast milk to their infants.

An enzyme-based immunodetection assay to quantify SARS-CoV-2 infection (preprint)

SARS-CoV-2 is a novel pandemic coronavirus that caused a global health and economic crisis. The development of efficient drugs and vaccines against COVID-19 requires detailed knowledge about SARS-CoV-2 biology. Several techniques to detect SARS-CoV-2 infection have been established, mainly based on counting infected cells by staining plaques or foci, or by quantifying the viral genome by PCR. These methods are laborious, time-consuming and expensive and therefore not suitable for a high sample throughput or rapid diagnostics. We here report a novel enzyme-based immunodetection assay that directly quantifies the amount of de novo synthesized viral spike protein within fixed and permeabilized cells. This in-cell ELISA enables a rapid and quantitative detection of SARS-CoV-2 infection in microtiter format, regardless of the virus isolate or target cell culture. It follows the established method of performing ELISA assays and does not require expensive instrumentation. Utilization of the in-cell ELISA allows to e.g. determine TCID50 of virus stocks, antiviral efficiencies (IC50 values) of drugs or neutralizing activity of sera. Thus, the in-cell spike ELISA represents a promising alternative to study SARS-CoV-2 infection and inhibition and may facilitate future research. HighlightsO_LIDetermination of SARS-CoV-2 infection by enzymatically quantifying the expression of viral spike protein in bulk cell cultures C_LIO_LITargeting a highly conserved region in the S2 subunit of the S protein allows broad detection of several SARS-CoV-2 isolates in different cell lines C_LIO_LIScreening of antivirals in microtiter format and determining the antiviral activity as inhibitory concentrations 50 (IC50) C_LI

Remdesivir but not famotidine inhibits SARS-CoV-2 replication in human pluripotent stem cell-derived intestinal organoids (preprint)

Gastrointestinal symptoms in COVID-19 are associated with prolonged symptoms and increased severity. We employed human intestinal organoids derived from pluripotent stem cells (PSC-HIOs) to analyze SARS-CoV-2 pathogenesis and to validate efficacy of specific drugs in the gut. Certain, but not all cell types in PSC-HIOs express SARS-CoV-2 entry factors ACE2 and TMPRSS2, rendering them susceptible to SARS-CoV-2 infection. Remdesivir, a promising drug to treat COVID-19, effectively suppressed SARS-CoV-2 infection of PSC-HIOs. In contrast, the histamine-2-blocker famotidine showed no effect. Thus, PSC-HIOs provide an interesting platform to study SARS-CoV-2 infection and to identify or validate drugs.