Coinfections and antimicrobial treatment in a cohort of falciparum malaria in a non-endemic country: a 10-year experience.

INTRODUCTION: Falciparum malaria remains one of the deadliest infectious diseases worldwide. In Germany, it is mainly an imported infection among travellers. Rates of coinfection are often unknown, and a clinical rationale for the beneficial use of calculated antibiotic therapy in patients with malaria and suspected coinfection is lacking. METHODS: We conducted an analysis of all in-patients treated with falciparum malaria at a German infectious diseases centre in vicinity to one of Europe's major airports for 2010-2019. Logistic regression and time-to-event analysis were used to evaluate predictors for bacterial coinfection, the use of antibacterial substances, as well as their influence on clinical course. RESULTS: In total, 264 patients were included. Of those, 64% received an additional antibacterial therapy (n = 169). Twenty-nine patients (11.0%) were found to have suffered from a relevant bacterial coinfection, while only a small fraction had relevant bacteremia (n = 3, 1.4%). However, patients with severe malaria did not suffer from coinfections more frequently (p = 0.283). CRP levels were not a reliable predictor for a bacterial coinfection (OR 0.99, 95% CI 0.94-1.06, p = 0.850), while another clinical focus of infection was positively associated (OR 3.86, 95% CI 1.45-11.55, p = 0.010). CONCLUSION: Although bacterial coinfections were rare in patients with malaria at our centre, the risk does not seem negligible. These data point rather towards individual risk assessment in respective patients than to general empiric antibiotic use.

Riding the Omicron BA.5 Wave: Improved Humoral Response after Vaccination with Bivalent Omicron BA.4-5-Adapted mRNA SARS-CoV-2 Vaccine in Chronic Hemodialysis Patients.

Hemodialysis patients faced an excess morbidity and mortality during the COVID-19 pandemic. We evaluated the effect of second-generation mRNA vaccines against Omicron BA.4 and BA.5 variants of SARS-CoV-2 on humoral immunity. The study population comprised 66 adult hemodialysis patients who have encountered four SARS-CoV-2 antigen contacts through vaccination or infection. We assessed their humoral response using an anti-SARS-CoV-2 spike receptor binding domain IgG antibody assay (S-RBD-ab), measuring neutralizing antibodies against ancestral strain of SARS-CoV-2, Delta, and Omicron in a surrogate virus neutralization test (SVNT), and specifically against BA.5 in a plaque reduction neutralization test (PRNT) before and four weeks after vaccination with Comirnaty Original/Omicron BA.4-5. During the following six months, SARS-CoV-2 infections and symptom severity were documented. The bivalent mRNA vaccine led to a 7.6-fold increase in S-RBD-ab levels and an augmented inhibition of the Omicron variant in SVNT by 35% (median). Seroconversion in the Omicron BA.5-specific PRNT was attained by in 78.4% of previously negative patients (29/37). Levels of S-RBD-ab correlated with inhibition in the Omicron-specific SVNT and neutralization titers in the BA.5-PRNT. Eleven SARS-CoV-2 infections occurred in the six-month follow-up, none of which took a life-threatening course. The bivalent mRNA vaccine improved the SARS-CoV-2 virus variant-specific humoral immunity in chronic hemodialysis patients. Measurement of S-RBD-ab can be used in hemodialysis patients to estimate their humoral immunity status against Omicron BA.5.

Progressive loss of conserved spike protein neutralizing antibody sites in Omicron sublineages is balanced by preserved T cell immunity.

Evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant has led to the emergence of sublineages with different patterns of neutralizing antibody evasion. We report that Omicron BA.4/BA.5 breakthrough infection of individuals immunized with SARS-CoV-2 wild-type-strain-based mRNA vaccines results in a boost of Omicron BA.4.6, BF.7, BQ.1.1, and BA.2.75 neutralization but does not efficiently boost BA.2.75.2, XBB, or XBB.1.5 neutralization. In silico analyses showed that the Omicron spike glycoprotein lost most neutralizing B cell epitopes, especially in sublineages BA.2.75.2, XBB, and XBB.1.5. In contrast, T cell epitopes are conserved across variants including XBB.1.5. T cell responses of mRNA-vaccinated, SARS-CoV-2-naive individuals against the wild-type strain, Omicron BA.1, and BA.4/BA.5 were comparable, suggesting that T cell immunity against recent sublineages including XBB.1.5 may remain largely unaffected. While some Omicron sublineages effectively evade B cell immunity, spike-protein-specific T cell immunity, due to the nature of polymorphic cell-mediated immune responses, may continue to contribute to prevention/limitation of severe COVID-19 manifestation.

Impact of different classes of immune-modulating treatments on B cell-related and T cell-related immune response before and after COVID-19 booster vaccination in patients with immune-mediated diseases and primary immunodeficiency: a cohort study.

OBJECTIVES: To evaluate the potential of immunosuppressed patients to mount B-cell and T-cell responses to COVID-19 booster vaccination (third vaccination). METHODS: Patients with primary immunodeficiency (PID), immune-mediated inflammatory diseases (IMIDs) on CD20-depleting treatment with rituximab (RTX), or IMIDs treated with conventional synthetic disease-modifying antirheumatic drugs (csDMARDs) or biological disease-modifying antirheumatic drug (bDMARDs) were included and assessed before (baseline visit (BL)) and 2, 4 and 8 weeks after COVID-19 booster vaccination. Serum B-cell responses were assessed by antibody levels against SARS-CoV-2 spike protein (anti-spike IgG antibody (S-AB)) and a surrogate virus neutralisation test (sVNT). T-cell responses were assessed by an interferon gamma release assay (IGRA). RESULTS: Fifty patients with PID (n=6), treated with RTX therapy (n=13), or treated with csDMARDs/bDMARDs (n=31) were included. At BL, anti-S-AB titres in PID and csDMARD/bDMARD-treated patients were low (although significantly higher than RTX patients); measures of B-cell-mediated response increased significantly after booster vaccination. In the RTX cohort, low BL anti-S-AB and sVNT values did not improve after booster vaccination, but patients had significantly elevated IGRA responses post booster vaccination compared with the other groups. csDMARD/bDMARD-treated patients showed the highest BL values in all three assays with greater increases in all parameters after booster vaccination compared with patients with PID. CONCLUSION: Patients with IMID on therapeutic B-cell depletion have low anti-S-AB and sVNT values before and after booster vaccination but show significantly higher levels of IGRA compared with other immunosuppressed patients, suggesting an underlying mechanism attempting to compensate compromised humoral immunity by upregulating T-cell responsiveness. PID appears to have a stronger impact on antiviral immune response than csDMARD/bDMARD treatment.

Comparison of the Ct-values for genomic and subgenomic SARS-CoV-2 RNA reveals limited predictive value for the presence of replication competent virus.

SARS-CoV-2 is the causative agent of the acute respiratory disease COVID-19. In addition to the full length positive-sensed, single-stranded genomic RNA (gRNA), viral subgenomic RNAs (sgRNAs) that are required for expression of the 3' region of the genome are synthesized in virus-infected cells. However, whether these sgRNA-species might be used as a measure of active virus replication and to predict infectivity is still under debate. The commonly used methods to monitor and quantitate SARS-CoV-2 infections are based on RT-qPCR analysis and the detection of gRNA. The infectivity of a sample obtained from nasopharyngeal or throat swabs is associated with the viral load and inversely correlates with Ct-values, however, a cut-off value predicting the infectivity highly depends on the performance of the assay. Furthermore, gRNA derived Ct-values result from nucleic acid detection and do not necessarily correspond to active replicating virus. We established a multiplex RT-qPCR assay on the cobas 6800 omni utility channel concomitantly detecting SARS-CoV-2 gRNAOrf1a/b, sgRNAE,7a,N, and human RNaseP-mRNA used as human input control. We compared the target specific Ct-values with the viral culture frequency and performed ROC curve analysis to determine the assay sensitivity and specificity. We found no advantage in the prediction of viral culture when using sgRNA detection compared to gRNA only, since Ct-values for gRNA and sgRNA were highly correlated and gRNA offered a slightly more reliable predictive value. Single Ct-values alone only provide a very limited prediction for the presence of replication competent virus. Hence, careful consideration of the medical history including symptom onset has to be considered for risk stratification.

Polio type 2 and 3 eradication: Relevance to the immunity status of individuals living in Germany, 2005-2020.

Since October 2019, poliovirus type 3 (PV3) has been certified as globally eradicated, and further laboratory use of PV3 will be restricted according to the WHO Polio Eradication Initiative and containment measures. To examine a possible gap in PV3 immunity and a lack of immunity against poliovirus type 2 (PV2), which was already declared as eradicated in 2015, neutralising antibodies against polioviruses (PV) of individuals living in Germany (n = 91,530 samples; mainly outpatients (≈90%) who received immune status testing) were investigated from 2005 to 2020 (age distribution: <18 years 15.8%, 18-64 years 71.2% and ≥65 years 9.5% for 2005-2015; <18 years 19.6%, 18-64 years 67% and ≥65 years 11.5% for 2016-2020). The results showed that the proportion of sera exclusively lacking antibodies against PV3 was 10.6% in 2005-2015 and 9.6% in 2016-2020 and against PV2 2.8% in 2005-2015. As there is decreased protection against PV3 and to detect potential antigenically (immune escape) variant PVs not covered by used vaccines, we recommend continued testing of PV1 and PV3.

Heterologous prime-boost immunization with ChAdOx1-S and BNT162b2: reactogenicity and immunogenicity in a prospective cohort study.

OBJECTIVES: Regarding reactogenicity and immunogenicity, heterologous COVID-19 vaccination regimens are considered as an alternative to conventional immunization schemes. METHODS: Individuals receiving either heterologous (ChAdOx1-S [AstraZeneca, Cambridge, UK]/BNT162b2 [Pfizer-BioNTech, Mainz, Germany]; n = 306) or homologous (messenger RNA [mRNA]-1273 [Moderna, Cambridge, Massachusetts, USA]; n = 139) vaccination were asked to participate when receiving their second dose. Reactogenicity was assessed after 1 month, immunogenicity after 1, 3, and/or 6 months, including a third dose, through SARS-CoV-2 antispike immunoglobulin G, surrogate virus neutralization test, and a plaque reduction neutralization test against the Delta (B.1.167.2) and Omicron (B.1.1.529; BA.1) variants of concern. RESULTS: The overall reactogenicity was lower after heterologous vaccination. In both cohorts, SARS-CoV-2 antispike immunoglobulin G concentrations waned over time with the heterologous vaccination demonstrating higher neutralizing activity than homologous mRNA vaccination after 3 months to low neutralizing levels in the Delta plaque reduction neutralization test after 6 months. At this point, 3.2% of the heterologous and 11.4% of the homologous cohort yielded low neutralizing activity against Omicron. After a third dose of an mRNA vaccine, ≥99% of vaccinees demonstrated positive neutralizing activity against Delta. Depending on the vaccination scheme and against Omicron, 60% to 87.5% of vaccinees demonstrated positive neutralizing activity. CONCLUSION: ChAdOx1-S/BNT162b2 vaccination demonstrated an acceptable reactogenicity and immunogenicity profile. A third dose of an mRNA vaccine is necessary to maintain neutralizing activity against SARS-CoV-2. However, variants of concern-adapted versions of the vaccines would be desirable.

Exposure to BA.4/5 S protein drives neutralization of Omicron BA.1, BA.2, BA.2.12.1, and BA.4/5 in vaccine-experienced humans and mice.

The SARS-CoV-2 Omicron variant and its sublineages show pronounced viral escape from neutralizing antibodies elicited by vaccination or prior SARS-CoV-2 variant infection owing to over 30-amino acid alterations within the spike (S) glycoprotein. Breakthrough infection of vaccinated individuals with Omicron sublineages BA.1 and BA.2 is associated with distinct patterns of cross-neutralizing activity against SARS-CoV-2 variants of concern (VOCs). In continuation of our previous work, we characterized the effect of Omicron BA.4/BA.5 S glycoprotein exposure on the neutralizing antibody response upon breakthrough infection in vaccinated individuals and upon variant-adapted booster vaccination in mice. We found that immune sera from triple mRNA-vaccinated individuals with subsequent breakthrough infection during the Omicron BA.4/BA.5 wave showed cross-neutralizing activity against previous Omicron variants BA.1, BA.2, BA.2.12.1, and BA.4/BA.5 itself. Administration of a prototypic BA.4/BA.5-adapted mRNA booster vaccine to mice after SARS-CoV-2 wild-type strain-based primary immunization is associated with broader cross-neutralizing activity than a BA.1-adapted booster. Whereas the Omicron BA.1-adapted mRNA vaccine in a bivalent format (wild-type + BA.1) broadens cross-neutralizing activity relative to the BA.1 monovalent booster, cross-neutralization of BA.2 and descendants is more effective in mice boosted with a bivalent wild-type + BA.4/BA.5 vaccine. In naïve mice, primary immunization with the bivalent wild-type + Omicron BA.4/BA.5 vaccine induces strong cross-neutralizing activity against Omicron VOCs and previous variants. These findings suggest that, when administered as boosters, mono- and bivalent Omicron BA.4/BA.5-adapted vaccines enhance neutralization breadth and that the bivalent version also has the potential to confer protection to individuals with no preexisting immunity against SARS-CoV-2.

SARS-CoV-2 Vaccination in Kidney Transplant Recipients-Stratified Analysis of the Humoral Immune Response.

Kidney transplant recipients are at increased risk of SARS-CoV-2 infection and a more severe course of COVID-19. Methods: We conducted a quantitative serologic testing of antibodies specific for the wild type of SARS-CoV-2 and the Omicron variant of concern before and after a third-dose vaccination, either mRNA-1273 (Moderna) or BNT162b2 (Pfizer-BioNTech) in a cohort of 103 stable kidney transplant recipients (median [range] age, 58 [22-84] y, 57 men [55.3%]). Results: Third-dose vaccination increased the seroconversion rate from 57.3% to 71.8%. However, despite a marked rise of the antibody concentrations after the booster, 55.4% and 11.6% only formed neutralizing antibodies against the SARS-CoV-2 wild type and Omicron, respectively. Treatment with mycophenolic acid/mycophenolate mofetil (in strata of the dose quartiles), advanced age, and' above all' impaired renal function (eGFR <60 mL/min) adversely influenced the humoral immunity regarding seroconversion and inhibition of the wild type of SARS-CoV-2. Conclusions: Apart from immunosuppressive therapy, the humoral vaccination response is largely affected by nonmodifiable factors in kidney transplant recipients. With the currently leading and clinically easier Omicron variant, this puts into perspective the strategy to significantly enhance the protective efficacy of the available vaccines by reducing or temporarily stopping proliferation inhibitors, not least considering the inherent rejection risk with a possible deterioration of graft function.

Reply to Fabbris et al. A Viable Alternative. Comment on "Kohmer et al. Self-Collected Samples to Detect SARS-CoV-2: Direct Comparison of Saliva, Tongue Swab, Nasal Swab, Chewed Cotton Pads and Gargle Lavage. J. Clin. Med. 2021, 10, 5751".

We thank Fabbris et al. for their remarks [...].

Limited neutralisation of the SARS-CoV-2 Omicron subvariants BA.1 and BA.2 by convalescent and vaccine serum and monoclonal antibodies.

BACKGROUND: In recent months, Omicron variants of SARS-CoV-2 have become dominant in many regions of the world, and case numbers with Omicron subvariants BA.1 and BA.2 continue to increase. Due to numerous mutations in the spike protein, the efficacy of currently available vaccines, which are based on Wuhan-Hu 1 isolate of SARS-CoV-2, is reduced, leading to breakthrough infections. Efficacy of monoclonal antibody therapy is also likely impaired. METHODS: In our in vitro study using A549-AT cells constitutively expressing ACE2 and TMPRSS2, we determined and compared the neutralizing capacity of vaccine-elicited sera, convalescent sera and monoclonal antibodies against authentic SARS-CoV-2 Omicron BA.1 and BA.2 compared with Delta. FINDINGS: Almost no neutralisation of Omicron BA.1 and BA.2 was observed using sera from individuals vaccinated with two doses 6 months earlier, regardless of the type of vaccine taken. Shortly after the booster dose, most sera from triple BNT162b2-vaccinated individuals were able to neutralise both Omicron variants. In line with waning antibody levels three months after the booster, only weak residual neutralisation was observed for BA.1 (26%, n = 34, 0 median NT50) and BA.2 (44%, n = 34, 0 median NT50). In addition, BA.1 but not BA.2 was resistant to the neutralising monoclonal antibodies casirivimab/imdevimab, while BA.2 exhibited almost a complete evasion from the neutralisation induced by sotrovimab. INTERPRETATION: Both SARS-CoV-2 Omicron subvariants BA.1 and BA.2 escape antibody-mediated neutralisation elicited by vaccination, previous infection with SARS-CoV-2, and monoclonal antibodies. Waning immunity renders the majority of tested sera obtained three months after booster vaccination negative in BA.1 and BA.2 neutralisation. Omicron subvariant specific resistance to the monoclonal antibodies casirivimab/imdevimab and sotrovimab emphasizes the importance of genotype-surveillance and guided application. FUNDING: This study was supported in part by the Goethe-Corona-Fund of the Goethe University Frankfurt (M.W.) and the Federal Ministry of Education and Research (COVIDready; grant 02WRS1621C (M.W.).

Impact of Moderna mRNA-1273 Booster Vaccine on Fully Vaccinated High-Risk Chronic Dialysis Patients after Loss of Humoral Response.

The long-term effect of protection by two doses of SARS-CoV-2 vaccination in patients receiving chronic intermittent hemodialysis (CIHD) is an urging question. We investigated the humoral and cellular immune response of 42 CIHD patients who had received two doses of SARS-CoV-2 vaccine, and again after a booster vaccine with mRNA-1273 six months later. We measured antibody levels and SARS-CoV-2-specific surrogate neutralizing antibodies (SNA). Functional T cell immune response to vaccination was assessed by quantifying interferon-γ (IFN-γ) and IL-2 secreting T cells specific for SARS-CoV-2 using an ELISpot assay. Our data reveal a moderate immune response after the second dose of vaccination, with significantly decreasing SARS-CoV-2-specific antibody levels and less than half of the study group showed neutralizing antibodies six months afterwards. Booster vaccines increased the humoral response dramatically and led to a response rate of 89.2% for antibody levels and a response rate of 94.6% for SNA. Measurement in a no response/low response (NR/LR) subgroup of our cohort, which differed from the whole group in age and rate of immunosuppressive drugs, indicated failure of a corresponding T cell response after the booster vaccine. We strongly argue in favor of a regular testing of surrogate neutralizing antibodies and consecutive booster vaccinations for CIHD patients to provide a stronger and persistent immunity.

Heterologous immunization with BNT162b2 followed by mRNA-1273 in dialysis patients: seroconversion and presence of neutralizing antibodies.

INTRODUCTION: The vital renal replacement therapy makes it impossible for dialysis patients to distance themselves socially. This results in a high risk of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and developing coronavuris disease 2019, with excess mortality due to disease burden and immunosuppression. We determined the efficacy of a 100-µg booster of mRNA-1273 (Moderna, Cambridge, MA, USA) 6 months after two doses of BNT162b2 (BioNTech/Pfizer, Mainz, Germany/New York, USA) in 194 SARS-CoV-2-naïve dialysis patients. METHODS: Anti-SARS-CoV-2 spike antibodies were measured with the Elecsys Anti-SARS-CoV-2 S assay (Roche Diagnostics, Mannheim, Germany) 4 and 10-12 weeks after two doses of BNT162b2 as well as 4 weeks after the mRNA-1273 booster. The presence of neutralizing antibodies was measured by the SARS-CoV-2 Surrogate Virus Neutralization Test (GenScript Biotech, Piscataway, NJ, USA). Two different cut-offs for positivity were used, one according to the manufacturer's specifications and one correlating with positivity in a plaque reduction neutralization test (PRNT). Receiver operating characteristics analyses were performed to match the anti-SARS-CoV-2 spike antibody cut-offs with the cut-offs in the surrogate neutralization assay accordingly. RESULTS: Any level of immunoreactivity determined by the anti-SARS-CoV-2 spike antibody assay was found in 87.3% (n = 144/165) and 90.6% (n = 164/181) of patients 4 and 10-12 weeks, respectively, after two doses of BNT162b2. This was reduced to 68.5% or 60.6% 4 weeks and 51.7% or 35.4% 10-12 weeks, respectively, when using the ROC cut-offs for neutralizing antibodies in the surrogate neutralization test (manufacturer's cut-off ≥103 U/mL and cut-off correlating with PRNT ≥196 U/mL). Four weeks after the mRNA-1273 booster, the concentration of anti-SARS-CoV-2 spike antibodies increased to 23 119.9 U/mL and to 97.3% for both cut-offs of neutralizing antibodies. CONCLUSION: Two doses of BNT162b2 followed by one dose of mRNA-1273 within 6 months in patients receiving maintenance dialysis resulted in significant titres of SARS-CoV-2 spike antibodies. While two doses of mRNA vaccine achieved adequate humoral immunity in a minority, the third vaccination boosts the development of virus-neutralizing quantities of SARS-CoV-2 spike antibodies (against wild-type SARS-CoV-2) in almost all patients.

Suitability of Different Diagnostic Platforms for Virological Testing of Blood Samples from Cornea Donors.

Background: To minimize the risk of disease transmission in cornea transplantation, donor screening for blood-derived viral infections is mandatory. Ideally, pre-mortem blood samples are used, but based on availability, cadaveric blood samples of cornea donors may also be used. However, serological and nucleic acid amplification tests (NATs) need to be validated for the use of cadaveric specimens. Methods: Hepatitis B virus (HBV), hepatitis C virus (HCV), human immunodeficiency virus (HIV), human T-lymphotropic virus (HTLV) 1/2, and Treponema pallidum (syphilis)-specific serological and/or NAT assays were validated on different platforms (Abbott Alinity i, Alinity m, Roche Cobas 6800, and Roche Cobas AmpliPrep/Cobas TaqMan (CAP/CTM)) using (un)spiked paired pre- and post-mortem cornea donor blood samples from the same individual (up to 23.83 h after death) of 28 individuals in accordance with the specifications of the German Federal Institute for Vaccines and Biomedicines (Paul-Ehrlich-Institut [PEI]). In addition, routinely HBV-, HCV- and HIV-PCR-negative tested post-mortem blood samples of 24 individuals were used to assess NAT specificity. Results: For the majority of serological parameters on the Abbott Alinity i (HBsAg, anti-HBc, anti-HBs, anti-HCV, anti-HIV, anti-HTLV 1/2, and anti-Treponema pallidum), ratios of generated test results of (un)spiked paired pre- and post-mortem blood samples differed ≤25%, with an agreement of qualitative pre- and post-mortem test results ranging from 91.2 to 100%. For NAT parameters (HBV, HCV, and HIV) on the Cobas 6800, Alinity m, and CAP/CTM, no significant deviation in virus concentrations (factor >5) of spiked pre- and post-mortem blood samples could be observed. Ct-values of corresponding internal controls did also not differ significantly (>1.5 Ct-values). In addition, no false-positive test results were generated when specificity was assessed. Conclusion: Overall, fluctuations of test results for serological and NAT parameters in pre- and post-mortem blood samples examined in this study, were only limited and within the range of what is also observed when routinely testing fresh patient specimens. We conclude that all examined assays are eligible for the screening of blood samples taken up to about 24 h after the occurrence of death.

Self-Collected Samples to Detect SARS-CoV-2: Direct Comparison of Saliva, Tongue Swab, Nasal Swab, Chewed Cotton Pads and Gargle Lavage.

Testing for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) by RT-PCR is a vital public health tool in the pandemic. Self-collected samples are increasingly used as an alternative to nasopharyngeal swabs. Several studies suggested that they are sufficiently sensitive to be a useful alternative. However, there are limited data directly comparing several different types of self-collected materials to determine which material is preferable. A total of 102 predominantly symptomatic adults with a confirmed SARS-CoV-2 infection self-collected native saliva, a tongue swab, a mid-turbinate nasal swab, saliva obtained by chewing a cotton pad and gargle lavage, within 48 h of initial diagnosis. Sample collection was unsupervised. Both native saliva and gargling with tap water had high diagnostic sensitivity of 92.8% and 89.1%, respectively. Nasal swabs had a sensitivity of 85.1%, which was not significantly inferior to saliva (p = 0.092), but 16.6% of participants reported they had difficult in self-collection of this sample. A tongue swab and saliva obtained by chewing a cotton pad had a significantly lower sensitivity of 74.2% and 70.2%, respectively. Diagnostic sensitivity was not related to the presence of clinical symptoms or to age. When comparing self-collected specimens from different material, saliva, gargle lavage or mid-turbinate nasal swabs may be considered for most symptomatic patients. However, complementary experiments are required to verify that differences in performance observed among the five sampling modes were not attributed to collection impairment.

Limited Neutralization of Authentic Severe Acute Respiratory Syndrome Coronavirus 2 Variants Carrying E484K In Vitro.

Whether monoclonal antibodies are able to neutralize severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern has been investigated using pseudoviruses. In this study we show that bamlanivimab, casirivimab, and imdevimab efficiently neutralize authentic SARS-CoV-2, including variant B.1.1.7 (alpha), but variants B.1.351 (beta) and P.2 (zeta) were resistant against bamlanivimab and partially resistant to casirivimab. Whether antibodies are able to neutralize severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variantshas been investigated using pseudoviruses. We show that authentic SARS-CoV-2 carrying E484K were resistant against bamlanivimab and less susceptible to casirivimab, convalescent and vaccine-elicited sera.

Utility of Different Surrogate Enzyme-Linked Immunosorbent Assays (sELISAs) for Detection of SARS-CoV-2 Neutralizing Antibodies.

The plaque reduction neutralization test (PRNT) is a preferred method for the detection of functional, SARS-CoV-2 specific neutralizing antibodies from serum samples. Alternatively, surrogate enzyme-linked immunosorbent assays (ELISAs) using ACE2 as the target structure for the detection of neutralization-competent antibodies have been developed. They are capable of high throughput, have a short turnaround time, and can be performed under standard laboratory safety conditions. However, there are very limited data on their clinical performance and how they compare to the PRNT. We evaluated three surrogate immunoassays (GenScript SARS-CoV-2 Surrogate Virus Neutralization Test Kit (GenScript Biotech, Piscataway Township, NJ, USA), the TECO® SARS-CoV-2 Neutralization Antibody Assay (TECOmedical AG, Sissach, Switzerland), and the Leinco COVID-19 ImmunoRank™ Neutralization MICRO-ELISA (Leinco Technologies, Fenton, MO, USA)) and one automated quantitative SARS-CoV-2 Spike protein-based IgG antibody assay (Abbott GmbH, Wiesbaden, Germany) by testing 78 clinical samples, including several follow-up samples of six BNT162b2 (BioNTech/Pfizer, Mainz, Germany/New York, NY, USA) vaccinated individuals. Using the PRNT as a reference method, the overall sensitivity of the examined assays ranged from 93.8 to 100% and specificity ranged from 73.9 to 91.3%. Weighted kappa demonstrated a substantial to almost perfect agreement. The findings of our study allow these assays to be considered when a PRNT is not available. However, the latter still should be the preferred choice. For optimal clinical performance, the cut-off value of the TECO assay should be individually adapted.

Surveillance of SARS-CoV-2 in Frankfurt am Main from October to December 2020 Reveals High Viral Diversity Including Spike Mutation N501Y in B.1.1.70 and B.1.1.7.

BACKGROUND: International travel is a major driver of the introduction and spread of SARS-CoV-2. AIM: To investigate SARS-CoV-2 genetic diversity in the region of a major transport hub in Germany, we characterized the viral sequence diversity of the SARS-CoV-2 variants circulating in Frankfurt am Main, the city with the largest airport in Germany, from the end of October to the end of December 2020. METHODS: In total, we recovered 136 SARS-CoV-2 genomes from nasopharyngeal swab samples. We isolated 104 isolates that were grown in cell culture and RNA from the recovered viruses and subjected them to full-genome sequence analysis. In addition, 32 nasopharyngeal swab samples were directly sequenced. RESULTS AND CONCLUSION: We found 28 different lineages of SARS-CoV-2 circulating during the study period, including the variant of concern B.1.1.7 (Δ69/70, N501Y). Six of the lineages had not previously been observed in Germany. We detected the spike protein (S) deletion Δ69/Δ70 in 15% of all sequences, a four base pair (bp) deletion (in 2.9% of sequences) and a single bp deletion (in 0.7% of sequences) in ORF3a, leading to ORF3a truncations. In four sequences (2.9%), an amino acid deletion at position 210 in S was identified. In a single sample (0.7%), both a 9 bp deletion in ORF1ab and a 7 bp deletion in ORF7a were identified. One sequence in lineage B.1.1.70 had an N501Y substitution while lacking the Δ69/70 in S. The high diversity of sequences observed over two months in Frankfurt am Main highlights the persisting need for continuous SARS-CoV-2 surveillance using full-genome sequencing, particularly in cities with international airport connections.