Distinct evolution of SARS-CoV-2 Omicron XBB and BA.2.86 lineages combining increased fitness and antibody evasion (preprint)

The unceasing circulation of SARS-CoV-2 leads to the continuous emergence of novel viral sublineages. Here, we isolated and characterized XBB.1, XBB.1.5, XBB.1.9.1, XBB.1.16.1, EG.5.1.1, EG.5.1.3, XBF, BA.2.86.1 and JN.1 variants, representing >80% of circulating variants in November 2023. The XBB subvariants carry few but recurrent mutations in the spike, whereas BA.2.86.1 and JN.1 harbor >30 additional changes. These variants replicated in IGROV-1 but no longer in Vero E6 and were not markedly fusogenic. They potently infected nasal epithelial cells, with EG.5.1.3 exhibiting the highest fitness. Antivirals remained active. Neutralizing antibody (NAb) responses from vaccinees and BA.1/BA.2-infected individuals were markedly lower compared to BA.1, without major differences between variants. An XBB breakthrough infection enhanced NAb responses against both XBB and BA.2.86 variants. JN.1 displayed lower affinity to ACE2 and higher immune evasion properties compared to BA.2.86.1. Thus, while distinct, the evolutionary trajectory of these variants combines increased fitness and antibody evasion.

High fusion and cytopathy of SARS-CoV-2 variant B.1.640.1 (preprint)

SARS-CoV-2 variants with undetermined properties have emerged intermittently throughout the COVID-19 pandemic. Some variants possess unique phenotypes and mutations which allow further characterization of viral evolution and spike functions. Around 1100 cases of the B.1.640.1 variant were reported in Africa and Europe between 2021 and 2022, before the expansion of Omicron. Here, we analyzed the biological properties of a B.1.640.1 isolate and its spike. Compared to the ancestral spike, B.1.640.1 carried 14 amino acid substitutions and deletions. B.1.640.1 escaped binding by some anti-NTD and -RBD monoclonal antibodies, and neutralization by sera from convalescent and vaccinated individuals. In cell lines, infection generated large syncytia and a high cytopathic effect. In primary airway cells, B.1.640.1 replicated less than Omicron BA.1 and triggered more syncytia and cell death than other variants. The B.1.640.1 spike was highly fusogenic when expressed alone. This was mediated by two poorly characterized and infrequent mutations located in the spike S2 domain, T859N and D936H. Altogether, our results highlight the cytopathy of a hyper-fusogenic SARS-CoV-2 variant, supplanted upon the emergence of Omicron BA.1.

Resistance of Omicron subvariants BA.2.75.2, BA.4.6 and BQ.1.1 to neutralizing antibodies (preprint)

Convergent evolution of SARS-CoV-2 Omicron BA.2, BA.4 and BA.5 lineages has led to the emergence of several new subvariants, including BA.2.75.2, BA.4.6. and BQ.1.1. The subvariants BA.2.75.2 and BQ.1.1 are expected to become predominant in many countries in November 2022. They carry an additional and often redundant set of mutations in the spike, likely responsible for increased transmissibility and immune evasion. Here, we established a viral amplification procedure to easily isolate Omicron strains. We examined their sensitivity to 6 therapeutic monoclonal antibodies (mAbs) and to 72 sera from Pfizer BNT162b2-vaccinated individuals, with or without BA.1/BA.2 or BA.5 breakthrough infection. Ronapreve (Casirivimab and Imdevimab) and Evusheld (Cilgavimab and Tixagevimab) lost any antiviral efficacy against BA.2.75.2 and BQ.1.1, whereas Xevudy (Sotrovimab) remained weakly active. BQ.1.1 was also resistant to Bebtelovimab. Neutralizing titers in triply vaccinated individuals were low to undetectable against BQ.1.1 and BA.2.75.2, 4 months after boosting. A BA.1/BA.2 breakthrough infection increased these titers, which remained about 18-fold lower against BA.2.75.2 and BQ.1.1, than against BA.1. Reciprocally, a BA.5 breakthrough infection increased more efficiently neutralization against BA.5 and BQ.1.1 than against BA.2.75.2. Thus, the evolution trajectory of novel Omicron subvariants facilitated their spread in immunized populations and raises concerns about the efficacy of most currently available mAbs.

Retrospective and prospective studies evaluating the performance of the SARS-Cov-2 "AQ+ COVID-19 Ag Rapid Test" from InTec on symptomatic and non-symptomatic patients (preprint)

For the last two years, the SARS-CoV-2 virus spread all around the world and led to the COVID-19 pandemic. The need of methods to control the pandemic and to propose rapid and efficient diagnostic tools has emerged. In this perspective, SARS-CoV-2 rapid antigen detection tests (RADT) have been developed. We performed a retrospective study on 638 collected nasopharyngeal samples used for reference RT-qPCR diagnosis to compare the AQ+ COVID-19 Ag Rapid Test from InTec (AQ+ InTec test) performance with other commercially available RADT. We analysed the sensitivity and specificity of the different tests and showed a better overall performance of the AQ+ InTec test, which was confirmed on the SARS-Cov-2 Omicron variant. We then conducted a prospective study on 1428 patients, to evaluate the sensitivity and specificity of the AQ+ InTec test on nasal and nasopharyngeal samples in a point of care setting. We showed that sensitivity and specificity reach acceptable criteria regarding the official recommendations of the MDCG 2021-21 in both symptomatic and asymptomatic patients. Overall, the results of these two studies confirm that the AQ+ InTec test is a valuable tool for testing in a pandemic context with a high proportion of asymptomatic patients who are potential carriers for the SARS-CoV-2 virus and is performant on the most current circulating variant Omicron.

Longitudinal analysis of serum neutralization of SARS-CoV-2 Omicron BA.2, BA.4 and BA.5 in patients receiving monoclonal antibodies (preprint)

The emergence of novel Omicron lineages, such as BA.5, may impact the therapeutic efficacy of anti-SARS-CoV-2 neutralizing monoclonal antibodies (mAbs). Here, we evaluated the neutralization and ADCC activity of 6 therapeutic mAbs against Delta, BA.2, BA.4 and BA.5 isolates. The Omicron sub-variants escaped most of the antibodies but remained sensitive to Bebtelovimab and Cilgavimab. Consistent with their shared spike sequence, BA.4 and BA.5 displayed identical neutralization profiles. Sotrovimab was the most efficient at eliciting ADCC. We also analyzed 121 sera from 40 immunocompromised individuals up to 6 months after infusion of 1200 mg of Ronapreve (Imdevimab + Casirivimab), and 300 or 600 mg of Evusheld (Cilgavimab + Tixagevimab). Sera from Ronapreve-treated individuals did not neutralize Omicron subvariants. Evusheld-treated individuals neutralized BA.2 and BA.5, but titers were reduced by 41- and 130-fold, respectively, compared to Delta. A longitudinal evaluation of sera from Evusheld-treated patients revealed a slow decay of mAb levels and neutralization. The decline was more rapid against BA.5. Our data shed light on the antiviral activities of therapeutic mAbs and the duration of effectiveness of Evusheld pre-exposure prophylaxis.

Duration of BA.5 neutralization in sera and nasal swabs from SARS-CoV-2 vaccinated individuals, with or without Omicron breakthrough infection. (preprint)

Since early 2022, Omicron BA.1 has been eclipsed by BA.2, which was in turn outcompeted by BA.5, that displays enhanced antibody escape properties. Here, we evaluated the duration of the neutralizing antibody (Nab) response, up to 16 months after Pfizer BNT162b2 vaccination, in individuals with or without BA.1/BA.2 breakthrough infection. We measured neutralization of the ancestral D614G lineage, Delta and Omicron BA.1, BA.2, BA.5 variants in 291 sera and 35 nasal swabs from 27 individuals. Upon vaccination, serum Nab titers were reduced by 10, 15 and 25 fold for BA.1, BA.2 and BA.5, respectively, compared with D614G. The duration of neutralization was markedly shortened, from an estimated period of 11.5 months post-boost with D614G to 5.5 months with BA.5. After breakthrough, we observed a sharp increase of Nabs against Omicron subvariants, followed by a plateau and a slow decline after 4 or 5 months. In nasal swabs, infection, but not vaccination, triggered a strong IgA response and a detectable Omicron neutralizing activity. Thus, BA.5 spread is partly due to abbreviated vaccine efficacy, particularly in individuals who were not infected with previous Omicron variants.

Seroneutralization of Omicron BA.1 and BA.2 in patients receiving anti-SARS-CoV-2 monoclonal antibodies. (preprint)

The SARS-CoV-2 Omicron BA.1 variant has been supplanted in many countries by the BA.2 sub-lineage. BA.2 differs from BA.1 by about 21 mutations in its spike. Human anti-spike monoclonalantibodies(mAbs)areusedforpreventionortreatmentofCOVID-19. However, the capacity of therapeutic mAbs to neutralize BA.1 and BA.2 remains poorly characterized. Here, we first compared the sensitivity of BA.1 and BA.2 to neutralization by 9 therapeutic mAbs. In contrast to BA.1, BA.2 was sensitive to Cilgavimab, partly inhibited by Imdevimab and resistant to Adintrevimab and Sotrovimab. Two combinations of mAbs, Ronapreve (Casirivimab + Imdevimab) and Evusheld (Cilgavimab + Tixagevimab), are indicated as a pre-exposure prophylaxis in immunocompromised persons at risk of severe disease. We analyzed sera from 29 such individuals, up to one month after administration of Ronapreve and/or Evusheld. After treatment, all individuals displayed elevated antibody levels in their sera and neutralized Delta with high titers. Ronapreve recipients did not neutralize BA.1 and weakly impaired BA.2. With Evusheld, neutralization of BA.1 and BA.2 was detected in 19 and 29 out of 29 patients, respectively. As compared to Delta, titers were more severely decreased against BA.1 (344-fold) than BA.2 (9-fold). We further report 4 breakthrough Omicron infections among the 29 participants. Therefore, BA.1 and BA.2 exhibit noticeable differences in their sensitivity to therapeutic mAbs. Anti-Omicron activity of Ronapreve, and to a lesser extent that of Evusheld, is reduced in patients sera, a phenomenon associated with decreased clinical efficacy.

Fusogenicity and neutralization sensitivity of the SARS-CoV-2 Delta sublineage AY.4.2 (preprint)

SARS-CoV-2 lineages are continuously evolving. As of December 2021, the AY.4.2 Delta sub-lineage represented 20 % of sequenced strains in UK and has been detected in dozens of countries. It has since then been supplanted by the Omicron variant. AY.4.2 displays three additional mutations (T95I, Y145H and A222V) in the N-terminal domain (NTD) of the spike when compared to the original Delta variant (B.1.617.2) and remains poorly characterized. Here, we analyzed the fusogenicity of the AY.4.2 spike and the sensitivity of an authentic AY.4.2 isolate to neutralizing antibodies. The AY.4.2 spike exhibited similar fusogenicity and binding to ACE2 than Delta. The sensitivity of infectious AY.4.2 to a panel of monoclonal neutralizing antibodies was similar to Delta, except for the anti-RBD Imdevimab, which showed incomplete neutralization. Sensitivity of AY.4.2 to sera from individuals having received two or three doses of Pfizer or two doses of AstraZeneca vaccines was reduced by 1.7 to 2.1 fold, when compared to Delta. Our results suggest that mutations in the NTD remotely impair the efficacy of anti-RBD antibodies. The temporary spread of AY.4.2 was not associated with major changes in spike function but rather to a partially reduced neutralization sensitivity.

Considerable escape of SARS-CoV-2 variant Omicron to antibody neutralization (preprint)

The SARS-CoV-2 Omicron variant was first identified in November 2021 in Botswana and South Africa 1,2 . It has in the meantime spread to many countries and is expected to rapidly become dominant worldwide. The lineage is characterized by the presence of about 32 mutations in the Spike, located mostly in the N-terminal domain (NTD) and the receptor binding domain (RBD), which may enhance viral fitness and allow antibody evasion. Here, we isolated an infectious Omicron virus in Belgium, from a traveller returning from Egypt. We examined its sensitivity to 9 monoclonal antibodies (mAbs) clinically approved or in development 3 , and to antibodies present in 90 sera from COVID-19 vaccine recipients or convalescent individuals. Omicron was totally or partially resistant to neutralization by all mAbs tested. Sera from Pfizer or AstraZeneca vaccine recipients, sampled 5 months after complete vaccination, barely inhibited Omicron. Sera from COVID-19 convalescent patients collected 6 or 12 months post symptoms displayed low or no neutralizing activity against Omicron. Administration of a booster Pfizer dose as well as vaccination of previously infected individuals generated an anti-Omicron neutralizing response, with titers 5 to 31 fold lower against Omicron than against Delta. Thus, Omicron escapes most therapeutic monoclonal antibodies and to a large extent vaccine-elicited antibodies.

Evaluation Of A New All In One Sars-Cov-2 Antigen-Detecting Rapid Diagnostic Test And Self-Test: Diagnostic Performance And Usability On Child And Adult Population (preprint)

The control of the COVID-19 epidemics has been one of top global health priorities for the last eighteen months. To that end, more reliable and easy-to-use diagnostic tests are necessary. Young children are still not eligible to vaccination and it is important to find a way to easily test this key population regularly. With that in mind, we evaluated a new innovative easy two-step self-test named COVID-VIRO ALL IN(R) developed by AAZ that uses a sampling nasal sponge instead of a classic nasal swab. Mirroring the previous study conducted on the first generation of COVID-VIRO(R) antigenic self-test, we first performed a multicentre, prospective study on 124 adults and children, in a point-of-care setting. Sensitivity, specificity and overall acceptance of the COVID-VIRO ALL IN(R) antigen self-test compared to RT-PCR on nasopharyngeal samples were evaluated at 93.02%, 100% and 97,5%, respectively. We then performed a multicentre, prospective, usability study to evaluate the ease of use of COVID-VIRO ALL IN(R) in real life on 68 laypersons, all adults. Globally, 99% of them considered the instructions material good, 98% executed the procedure well, and all of them interpreted the results correctly. The usability was then specifically investigated on 40 children and teenagers participants, comparing both COVID-VIRO(R) first generation and the new COVID-VIRO ALL IN(R). All of them found COVID-VIRO ALL IN(R) much easier to use and much more comfortable. For young children, the COVID-VIRO ALL IN(R) self-test appears to be safer (less risk of trauma compare to nasal swabs and no liquid exposure) easier to use than classic COVID self-tests and giving immediate result which is not the case for RT-PCR done on saliva samples (currently done in routine for kids in French schools). It could be an adapted tool for future mass screening campaigns in schools or at home under adult supervision for kids from the age of 3.

Reduced sensitivity of infectious SARS-CoV-2 variant B.1.617.2 to monoclonal antibodies and sera from convalescent and vaccinated individuals (preprint)

The SARS-CoV-2 B.1.617 lineage emerged in October 2020 in India. It has since then become dominant in some indian regions and further spread to many countries. The lineage includes three main subtypes (B1.617.1, B.1617.2 and B.1.617.3), which harbour diverse Spike mutations in the N-terminal domain (NTD) and the receptor binding domain (RBD) which may increase their immune evasion potential. B.1.617.2 is believed to spread faster than the other versions. Here, we isolated infectious B.1.617.2 from a traveller returning from India. We examined its sensitivity to monoclonal antibodies (mAbs) and to antibodies present in sera from COVID-19 convalescent individuals or vaccine recipients, in comparison to other viral lineages. B.1.617.2 was resistant to neutralization by some anti-NTD and anti-RBD mAbs, including Bamlanivimab, which were impaired in binding to the B.1.617.2 Spike. Sera from convalescent patients collected up to 12 months post symptoms and from Pfizer Comirnaty vaccine recipients were 3 to 6 fold less potent against B.1.617.2, relative to B.1.1.7. Sera from individuals having received one dose of AstraZeneca Vaxzevria barely inhibited B.1.617.2. Thus, B.1.617.2 spread is associated with an escape to antibodies targeting non-RBD and RBD Spike epitopes.

SARS-CoV-2 in Ivory Coast: serosurveillance survey among mines workers (preprint)

BackgroundEight months after the detection of the first COVID-19 case in Africa, 1,262,476 cases have been reported in African countries compared to 72 million worldwide. The real burden of SARS-CoV-2 infection in West Africa is not clearly defined. The aim of the study was to evaluate the seroprevalence of SARS-CoV-2 in half of the 3,380 workers of several mining companies operating in two mines in the Ivory Coast and having its headquarters in the economic capital Abidjan. MethodsFrom 15th July to 13th October 2020, a voluntary serological test campaign was performed in the 3 sites where the companies operate: two mines, and the headquarters in Abidjan.We performed a COVID-PRESTO rapid test for the detection of IgG and IgM on capillary blood. A multivariate analysis was performed to identify independent sociodemographic characteristics associated with a higher SARS-CoV-2 seroprevalence rate. ResultsA total of 1,687 subjects were tested. 91% were male (n= 1,536) and mean age was 37 years old. The overall crude seroprevalence rate was 25.1% (n=422), but differing significantly between different sites, rising from 13.6% (11.2%-16.1%) in mine A to 34.4% (31.1%-37.7%) in mine B and 34.7% (26.2%-43.2%) in Abidjan. Non-resident workers in mines had a significantly lower prevalence rate than those living full-time in mines. Seroprevalence was 26.5% in natives of the Ivory Coast, while people coming from countries other than Africa were less likely to be SARS-CoV-2 seropositive. Among the 422 positive subjects, 74 reported mild symptoms in the three previous months and one was hospitalized for a severe COVID-19 infection. ConclusionThe prevalence of SARS-CoV-2 infection among mine workers in Ivory Coast is high. The low morbidity observed has probably led to an underestimation of the burden of this infection in West Africa. The high prevalence reported in subjects living in Abidjan, who have not any close contact with mine workers, may be indicative of the real seroprevalence in the Ivory Coast capital.

Real-life performances of a novel antigen detection test on nasopharyngeal specimens for SARS-CoV-2 infection diagnosis: a prospective study (preprint)

Introduction The SARS-CoV-2 pandemic has become a major public health issue worldwide. Developing and evaluating rapid and easy-to-perform diagnostic tests is an absolute priority. The current prospective study was designed to assess diagnostic performances of an antigen-based rapid detection test (COVID-VIRO) in a real-life setting. Methods Two nasopharyngeal specimens of symptomatic or asymptomatic adult patients hospitalized in the Infectious Diseases Department or voluntarily accessing the COVID-19 Screening Department of the Regional Hospital of Orleans, France, were concurrently collected. COVID VIRO diagnostic specificity and sensitivity were assessed in comparison to real-time reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) results. A subgroup of patients underwent an additional oropharyngeal and/or a saliva swab for rapid testing. Results 121 patients already having a confirmed infection and 127 patients having no evidence of recent or ongoing infection were enrolled, for a total of 248 couple of nasopharyngeal swab specimens. Overall COVID-VIRO sensitivity was 96.7% (IC: 93.5%-99.9%). In asymptomatic patients, patients having symptoms for more than 4 days and those having a RT-qPCR Cycle threshold value >32, sensitivity was of 100%, 95.8% and 96.9% respectively. The concordance between RT-qPCR and COVID VIRO rapid test was 100% for the 127 patients with no SARS-CoV-2 infection. Conclusion COVID-VIRO test had 100% specificity and above 95% sensitivity, better than WHO recommendations (specificity [≥]97-100%, sensitivity [≥]80%). These rapid tests are particularly interesting for large-scale screening in Emergency Department, low resource settings and airports.

Evaluation of the Practicability of a Finger-Stick Whole-Blood SARS-Cov-2 Self-Test Adapted for the General Population. (preprint)

BackgroundCOVID-19 (COronaVIrus Disease 2019) is an infectious respiratory disease caused by the novel SARS-CoV-2 virus. Rapid Diagnostic Tests (RDTs) have been developed to detect specific antibodies, IgG and IgM, to SARS-CoV-2 virus in human whole blood and easily usable by the general population are needed in order to alleviate the lockdown that many countries have initiated in response to the growing COVID-19 pandemic. A real-life study has been conducted in order to evaluate the performance of the COVID-PRESTO(R) RDT and the results have been submitted for publication and are currently under review. Even if this test showed very high sensitivity and specificity in a laboratory setting when used by trained professionals, it needs to be further evaluated for practicability when used by common folk in order to be approved by health authorities for in-home use Methods142 participants were recruited between March 2020 and April 2020 among non-medical populations in central France (nuclear plants workers, individuals attending the Orleans University Hospital vaccination clinic and Orleans University Hospital non-medical staff). Instructions for use with or without a tutorial video was made available to the volunteers. Two separate objectives were pursued: evaluation of the capability of participants to obtain an interpretable result, and evaluation of the users ability to read the results. Results88.4 % of the test users judged the instruction for use leaflet to be clear and understandable. 99.3 % of the users obtained a valid results and according to the supervisors 92.7% of the tests were properly performed by the user. Overall, 95% of the users gave positive feedback toward the COVID PRESTO(R) as a potential self-test. No influence of age and education was observed. ConclusionCOVID-PRESTO(R) was successfully used by an overwhelming majority of participants and its utilization was judged very satisfactory, therefore showing a promising potential as a self-test to be used by the general population. This RDT can become an easy-to-use tool to help know whether individuals are protected or not, particularly in the perspective of a second wave or a mass vaccination program.

Evaluation of performance of two SARS-CoV-2 Rapid whole-blood finger-stick IgM-IgGCombined Antibody Tests (preprint)

Background The SARS-CoV-2 virus is responsible for the infectious respiratory disease called COVID-19 (COronaVIrus Disease). In response to the growing COVID-19 pandemic, Rapid Diagnostic Tests (RDTs) have been developed to detect specific antibodies, IgG and IgM, to SARS-CoV-2 virus in human whole blood. We conducted a real-life study to evaluate the performance of two RDTs, COVID-PRESTO and COVID-DUO, compared to the gold standard, RT-PCR. Methods RT-PCR testing of SARS-Cov-2 was performed from nasopharyngeal swab specimens collected in adult patients visiting the infectious disease department at the hospital (Orleans, France). Fingertip whole blood samples taken at different time points after onset of the disease were tested with RDTs. The specificity and sensitivity of the rapid test kits compared to test of reference (RT-PCR) were calculated. Results Among 381 patients with symptoms of COVID-19 who went to the hospital for a diagnostic, 143 patients were RT-PCR negative. Results of test with RDTs were all negative for these patients, indicating a specificity of 100% for both RDTs. In the RT-PCR positive subgroup (n=238), 133 patients were tested with COVID-PRESTO and 129 patients were tested with COVID-DUO (24 patients tested with both). The further the onset of symptoms was from the date of collection, the greater the sensitivity. The sensitivity of COVID-PRESTO test ranged from 10.00% for patients having experienced their 1st symptoms from 0 to 5 days ago to 100% in patients where symptoms had occurred more than 15 days before the date of tests. For COVID-DUO test, the sensitivity ranged from 35.71% [0-5 days] to 100% (> 15 days). Conclusion COVID-PRESTO and DUO RDTs turned out to be very specific (none false positive) and to be sensitive enough after 15 days from onset of symptom. These easy to use IgG/IgM combined test kits are the first ones allowing a screening with capillary blood sample, by typing from a finger prick. These rapid tests are particularly interesting for screening in low resource settings.