TMPRSS2 is a functional receptor for human coronavirus HKU1.

Four endemic seasonal human coronaviruses causing common colds circulate worldwide: HKU1, 229E, NL63 and OC43 (ref. 1). After binding to cellular receptors, coronavirus spike proteins are primed for fusion by transmembrane serine protease 2 (TMPRSS2) or endosomal cathepsins2-9. NL63 uses angiotensin-converting enzyme 2 as a receptor10, whereas 229E uses human aminopeptidase-N11. HKU1 and OC43 spikes bind cells through 9-O-acetylated sialic acid, but their protein receptors remain unknown12. Here we show that TMPRSS2 is a functional receptor for HKU1. TMPRSS2 triggers HKU1 spike-mediated cell-cell fusion and pseudovirus infection. Catalytically inactive TMPRSS2 mutants do not cleave HKU1 spike but allow pseudovirus infection. Furthermore, TMPRSS2 binds with high affinity to the HKU1 receptor binding domain (Kd 334 and 137 nM for HKU1A and HKU1B genotypes) but not to SARS-CoV-2. Conserved amino acids in the HKU1 receptor binding domain are essential for binding to TMPRSS2 and pseudovirus infection. Newly designed anti-TMPRSS2 nanobodies potently inhibit HKU1 spike attachment to TMPRSS2, fusion and pseudovirus infection. The nanobodies also reduce infection of primary human bronchial cells by an authentic HKU1 virus. Our findings illustrate the various evolution strategies of coronaviruses, which use TMPRSS2 to either directly bind to target cells or prime their spike for membrane fusion and entry.

Reduced sensitivity of SARS-CoV-2 variant Delta to antibody neutralization.

The SARS-CoV-2 B.1.617 lineage was identified in October 2020 in India1-5. Since then, it has become dominant in some regions of India and in the UK, and has spread to many other countries6. The lineage includes three main subtypes (B1.617.1, B.1.617.2 and B.1.617.3), which contain diverse mutations in the N-terminal domain (NTD) and the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein that may increase the immune evasion potential of these variants. B.1.617.2-also termed the Delta variant-is believed to spread faster than other variants. Here we isolated an infectious strain of the Delta variant from an individual with COVID-19 who had returned to France from India. We examined the sensitivity of this strain to monoclonal antibodies and to antibodies present in sera from individuals who had recovered from COVID-19 (hereafter referred to as convalescent individuals) or who had received a COVID-19 vaccine, and then compared this strain with other strains of SARS-CoV-2. The Delta variant was resistant to neutralization by some anti-NTD and anti-RBD monoclonal antibodies, including bamlanivimab, and these antibodies showed impaired binding to the spike protein. Sera collected from convalescent individuals up to 12 months after the onset of symptoms were fourfold less potent against the Delta variant relative to the Alpha variant (B.1.1.7). Sera from individuals who had received one dose of the Pfizer or the AstraZeneca vaccine had a barely discernible inhibitory effect on the Delta variant. Administration of two doses of the vaccine generated a neutralizing response in 95% of individuals, with titres three- to fivefold lower against the Delta variant than against the Alpha variant. Thus, the spread of the Delta variant is associated with an escape from antibodies that target non-RBD and RBD epitopes of the spike protein.

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

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 1,100 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-N-terminal domain and anti-receptor-binding domain 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. (This study has been registered at ClinicalTrials.gov under registration no. NCT04750720.)IMPORTANCEOur results highlight the plasticity of SARS-CoV-2 Spike to generate highly fusogenic and cytopathic strains with the causative mutations being uncharacterized in previous variants. We describe mechanisms regulating the formation of syncytia and the subsequent consequences in a primary culture model, which are poorly understood.

Development of a highly specific and sensitive VHH-based sandwich immunoassay for the detection of the SARS-CoV-2 nucleoprotein.

The current COVID-19 pandemic illustrates the importance of obtaining reliable methods for the rapid detection of SARS-CoV-2. A highly specific and sensitive diagnostic test able to differentiate the SARS-CoV-2 virus from common human coronaviruses is therefore needed. Coronavirus nucleoprotein (N) localizes to the cytoplasm and the nucleolus and is required for viral RNA synthesis. N is the most abundant coronavirus protein, so it is of utmost importance to develop specific antibodies for its detection. In this study, we developed a sandwich immunoassay to recognize the SARS-CoV-2 N protein. We immunized one alpaca with recombinant SARS-CoV-2 N and constructed a large single variable domain on heavy chain (VHH) antibody library. After phage display selection, seven VHHs recognizing the full N protein were identified by ELISA. These VHHs did not recognize the nucleoproteins of the four common human coronaviruses. Hydrogen Deuterium eXchange-Mass Spectrometry (HDX-MS) analysis also showed that these VHHs mainly targeted conformational epitopes in either the C-terminal or the N-terminal domains. All VHHs were able to recognize SARS-CoV-2 in infected cells or on infected hamster tissues. Moreover, the VHHs could detect the SARS variants B.1.17/alpha, B.1.351/beta, and P1/gamma. We propose that this sandwich immunoassay could be applied to specifically detect the SARS-CoV-2 N in human nasal swabs.

Gaseous nitric oxide failed to inhibit the replication cycle of SARS-CoV-2 in vitro.

Nitric oxide (NO) has been shown to have antimicrobial activity in vitro and in some in vivo models, while the virucidal activity of NO remains elusive. Some studies using NO donors have suggested that NO could be a potential candidate to treat SARS-CoV infection. The Covid-19 pandemic raised the hypothesis that NO gas might have an impact on Sars-CoV-2 replication cycle and might be considered as a candidate therapy to treat COVID-19. To our knowledge, there are no in vitro preclinical studies demonstrating a virucidal effect of gaseous NO on SARS-CoV-2. This study aims to determine whether gaseous NO has an impact on the replication cycle of SARS-CoV-2 in vitro. To that end, SARS-CoV-2 infected epithelial (VeroE6) and pulmonary (A549-hACE2) cells were treated with repeated doses of gaseous NO at different concentrations known to be efficient against bacteria. Our results show that exposing SARS-CoV-2 infected-cells to NO gas even at high doses (160 ppm, 6 h) does not influence the replication cycle of the virus in vitro. We report here that NO gas has no antiviral properties in vitro on SARS-COV-2. Therefore, there is no rationale for its usage in clinical settings to treat COVID-19 patients for direct antiviral purposes, which does not exclude other potential physiological benefits of this gas.

Untargeted metagenomic sequencing identifies Toscana virus in patients with idiopathic meningitis, southern Spain, 2015 to 2019.

BackgroundVarious pathogens, including bacteria, fungi, parasites, and viruses can lead to meningitis. Among viruses causing meningitis, Toscana virus (TOSV), a phlebovirus, is transmitted through sandfly bites. TOSV infection may be suspected if patients with enterovirus- and herpesvirus-negative aseptic (non-bacterial) meningitis recall recent insect bites. Other epidemiological factors (season, rural area) may be considered. The broad range of possible meningitis aetiologies poses considerable diagnosis challenges. Untargeted metagenomic next-generation sequencing (mNGS) can potentially identify pathogens, which are not considered or detected in routine diagnostic panels.AimIn this retrospective, single-centre observational study, we investigated mNGS usefulness to understand the cause of meningitis when conventional approaches fail.MethodsCerebrospinal fluid (CSF) samples from patients hospitalised in southern Spain in 2015-2019 with aseptic meningitis and no aetiology found by conventional testing, were subjected to mNGS. Patients' demographic characteristics had been recorded and physicians had asked them about recent insect bites. Obtained viral genome sequences were phylogenetically analysed.ResultsAmong 23 idiopathic cases, TOSV was identified in eight (all male; median age: 39 years, range: 15-78 years). Five cases lived in an urban setting, three occurred in autumn and only one recalled insect bites. Phylogenetic analysis of TOSV segment sequences supported one intra-genotype reassortment event.ConclusionsOur study highlights the usefulness of mNGS for identifying viral pathogens directly in CSF. In southern Spain, TOSV should be considered regardless of recalling of insect bites or other epidemiological criteria. Detection of a disease-associated reassortant TOSV emphasises the importance of monitoring the spread and evolution of phleboviruses in Mediterranean countries.

Genetic Diversity of Collaborative Cross Mice Controls Viral Replication, Clinical Severity, and Brain Pathology Induced by Zika Virus Infection, Independently of Oas1b.

The explosive spread of Zika virus (ZIKV) has been associated with major variations in severe disease and congenital afflictions among infected populations, suggesting an influence of host genes. We investigated how genome-wide variants could impact susceptibility to ZIKV infection in mice. We first describe that the susceptibility of Ifnar1-knockout mice is largely influenced by their genetic background. We then show that Collaborative Cross (CC) mice, which exhibit a broad genetic diversity, in which the type I interferon receptor (IFNAR) was blocked by an anti-IFNAR antibody expressed phenotypes ranging from complete resistance to severe symptoms and death, with large variations in the peak and the rate of decrease in the plasma viral load, in the brain viral load, in brain histopathology, and in the viral replication rate in infected cells. The differences in susceptibility to ZIKV between CC strains correlated with the differences in susceptibility to dengue and West Nile viruses between the strains. We identified highly susceptible and resistant mouse strains as new models to investigate the mechanisms of human ZIKV disease and other flavivirus infections. Genetic analyses revealed that phenotypic variations are driven by multiple genes with small effects, reflecting the complexity of ZIKV disease susceptibility in the human population. Notably, our results rule out the possibility of a role of the Oas1b gene in the susceptibility to ZIKV. Altogether, the findings of this study emphasize the role of host genes in the pathogeny of ZIKV infection and lay the foundation for further genetic and mechanistic studies.IMPORTANCE In recent outbreaks, ZIKV has infected millions of people and induced rare but potentially severe complications, including Guillain-Barré syndrome and encephalitis in adults. While several viral sequence variants were proposed to enhance the pathogenicity of ZIKV, the influence of host genetic variants in mediating the clinical heterogeneity remains mostly unexplored. We addressed this question using a mouse panel which models the genetic diversity of the human population and a ZIKV strain from a recent clinical isolate. Through a combination of in vitro and in vivo approaches, we demonstrate that multiple host genetic variants determine viral replication in infected cells and the clinical severity, the kinetics of blood viral load, and brain pathology in mice. We describe new mouse models expressing high degrees of susceptibility or resistance to ZIKV and to other flaviviruses. These models will facilitate the identification and mechanistic characterization of host genes that influence ZIKV pathogenesis.

A Single Dose of NILV-Based Vaccine Provides Rapid and Durable Protection against Zika Virus.

Zika virus, a member of the Flaviviridae family, is primarily transmitted by infected Aedes species mosquitoes. In 2016, Zika infection emerged as a global health emergency for its explosive spread and the remarkable neurological defects in the developing fetus. Development of a safe and effective Zika vaccine remains a high priority owing to the risk of re-emergence and limited understanding of Zika virus epidemiology. We engineered a non-integrating lentiviralvector(NILV)-based Zika vaccine encoding the consensus pre-membrane and envelope glycoprotein of circulating Zika virus strains. We further evaluated the immunogenicity and protective efficacy of this vaccine in both immunocompromised and immunocompetent mouse models. A single immunization in both mouse models elicited a robust neutralizing antibody titer and afforded full protection against Zika challenge as early as 7 days post-immunization. This NILV-based vaccine also induced a long-lasting immunity when immunized mice were challenged 6 months after immunization. Altogether, our NILV Zika vaccine provides a rapid yet durable protection through a single dose of immunization without extra adjuvant formulation. Our data suggest a promising Zika vaccine candidate for an emergency situation, and demonstrate the capacity of lentiviral vector as an efficient vaccine delivery platform.

Genomic Epidemiology of 2015-2016 Zika Virus Outbreak in Cape Verde.

During 2015-2016, Cape Verde, an island nation off the coast of West Africa, experienced a Zika virus (ZIKV) outbreak involving 7,580 suspected Zika cases and 18 microcephaly cases. Analysis of the complete genomes of 3 ZIKV isolates from the outbreak indicated the strain was of the Asian (not African) lineage. The Cape Verde ZIKV sequences formed a distinct monophylogenetic group and possessed 1-2 (T659A, I756V) unique amino acid changes in the envelope protein. Phylogeographic and serologic evidence support earlier introduction of this lineage into Cape Verde, possibly from northeast Brazil, between June 2014 and August 2015, suggesting cryptic circulation of the virus before the initial wave of cases were detected in October 2015. These findings underscore the utility of genomic-scale epidemiology for outbreak investigations.

Visualization of self-delivering hydrophobically modified siRNA cellular internalization.

siRNAs are a new class of therapeutic modalities with promising clinical efficacy that requires modification or formulation for delivery to the tissue and cell of interest. Conjugation of siRNAs to lipophilic groups supports efficient cellular uptake by a mechanism that is not well characterized. Here we study the mechanism of internalization of asymmetric, chemically stabilized, cholesterol-modified siRNAs (sd-rxRNAs®) that efficiently enter cells and tissues without the need for formulation. We demonstrate that uptake is rapid with significant membrane association within minutes of exposure followed by the formation of vesicular structures and internalization. Furthermore, sd-rxRNAs are internalized by a specific class of early endosomes and show preferential association with epidermal growth factor (EGF) but not transferrin (Tf) trafficking pathways as shown by live cell TIRF and structured illumination microscopy (SIM). In fixed cells, we observe ∼25% of sd-rxRNA co-localizing with EGF and <5% with Tf, which is indicative of selective endosomal sorting. Likewise, preferential sd-rxRNA co-localization was demonstrated with EEA1 but not RBSN-containing endosomes, consistent with preferential EGF-like trafficking through EEA1-containing endosomes. sd-rxRNA cellular uptake is a two-step process, with rapid membrane association followed by internalization through a selective, saturable subset of the endocytic process. However, the mechanistic role of EEA1 is not yet known. This method of visualization can be used to better understand the kinetics and mechanisms of hydrophobic siRNA cellular uptake and will assist in further optimization of these types of compounds for therapeutic intervention.

A Blood RNA Signature Detecting Severe Disease in Young Dengue Patients at Hospital Arrival.

Background: Early detection of severe dengue can improve patient care and survival. To date, no reliable single-gene biomarker exists. We hypothesized that robust multigene signatures exist. Methods: We performed a prospective study on Cambodian dengue patients aged 4 to 22 years. Peripheral blood mononuclear cells (PBMCs) were obtained at hospital admission. We analyzed 42 transcriptomic profiles of patients with secondary dengue infected with dengue serotype 1. Our novel signature discovery approach controls the number of included genes and captures nonlinear relationships between transcript concentrations and severity. We evaluated the signature on secondary cases infected with different serotypes using 2 datasets: 22 PBMC samples from additional patients in our cohort and 32 whole blood samples from an independent cohort. Results: We identified an 18-gene signature for detecting severe dengue in patients with secondary infection upon hospital admission with a sensitivity of 0.93 (95% confidence interval [CI], .82-.98), specificity of 0.67 (95% CI, .53-.80), and area under the receiver operating characteristic curve (AUC) of 0.86 (95% CI, .75-.97). At validation, the signature had empirical AUCs of 0.85 (95% CI, .69-1.00) and 0.83 (95% CI, .68-.98) for the PBMCs and whole blood datasets, respectively. Conclusions: The signature could detect severe dengue in secondary-infected patients upon hospital admission. Its genes offer new insights into the pathogenesis of severe dengue.

Inhibition of Polyamine Biosynthesis Is a Broad-Spectrum Strategy against RNA Viruses.

RNA viruses present an extraordinary threat to human health, given their sudden and unpredictable appearance, the potential for rapid spread among the human population, and their ability to evolve resistance to antiviral therapies. The recent emergence of chikungunya virus, Zika virus, and Ebola virus highlights the struggles to contain outbreaks. A significant hurdle is the availability of antivirals to treat the infected or protect at-risk populations. While several compounds show promise in vitro and in vivo, these outbreaks underscore the need to accelerate drug discovery. The replication of several viruses has been described to rely on host polyamines, small and abundant positively charged molecules found in the cell. Here, we describe the antiviral effects of two molecules that alter polyamine levels: difluoromethylornithine (DFMO; also called eflornithine), which is a suicide inhibitor of ornithine decarboxylase 1 (ODC1), and diethylnorspermine (DENSpm), an activator of spermidine/spermine N1-acetyltransferase (SAT1). We show that reducing polyamine levels has a negative effect on diverse RNA viruses, including several viruses involved in recent outbreaks, in vitro and in vivo These findings highlight the importance of the polyamine biosynthetic pathway to viral replication, as well as its potential as a target in the development of further antivirals or currently available molecules, such as DFMO. IMPORTANCE: RNA viruses present a significant hazard to human health, and combatting these viruses requires the exploration of new avenues for targeting viral replication. Polyamines, small positively charged molecules within the cell, have been demonstrated to facilitate infection for a few different viruses. Our study demonstrates that diverse RNA viruses rely on the polyamine pathway for replication and highlights polyamine biosynthesis as a promising drug target.

Zanamivir and baloxavir combination to cure persistent influenza and coronavirus infections after hematopoietic stem cell transplant.

OBJECTIVES: Immunocompromised patients may experience prolonged shedding of influenza virus potentially leading to severe infections. Alternatives to monotherapy with neuraminidase inhibitors should be evaluated to entirely suppress viral replication and prevent drug-resistant mutations. METHODS: We investigated the clinical and virological evolution in a case of persistent influenza A and human coronavirus OC43 (HCoV-OC43) coinfection in a hematopoietic stem cell transplant recipient after different therapeutic strategies. RESULTS: Successive oseltamivir and zanamivir monotherapies failed to control both infections, with positive results persisting for over 110 days each. This led to the emergence of highly resistant oseltamivir strains due to neuraminidase mutations (E119V and R292K) followed by a deletion (del245-248), while maintaining sensitivity to zanamivir. The intra-host viral diversity data showed that the treatments impacted viral diversity of influenza virus, but not of HCoV-OC43. Considering the patient's underlying condition and the impact of prolonged viral shedding on pulmonary function, eradicating the influenza virus was necessary. A 10-day regimen combining zanamivir and baloxavir-marboxil effectively controlled influenza virus replication and was associated with the clearance of HCoV-OC43, finally resulting in comprehensive respiratory recovery. CONCLUSION: These observations underscore the importance of further investigating combination treatments as the primary approach to achieve influenza eradication in immunocompromised patients.

Escape of SARS-CoV-2 Variants KP.1.1, LB.1, and KP3.3 From Approved Monoclonal Antibodies.

Background: First-generation anti-SARS-CoV-2 monoclonal antibodies (mAbs) used for prophylaxis or therapeutic purposes in immunocompromised patients have been withdrawn because of the emergence of resistant Omicron variants. In 2024, 2 novel mAbs, VYD222/Pemivibart and AZD3152/Sipavibart, were approved by health authorities, but their activity against contemporary JN.1 sublineages is poorly characterized. Methods: We isolated authentic JN.1.1, KP.1.1, LB.1, and KP.3.3 viruses and evaluated their sensitivity to neutralization by these mAbs in 2 target cell lines. Results: Compared to ancestral strains, VYD222/Pemivibart remained moderately active against JN.1 subvariants, with a strong increase of 50% Inhibitory Concentration (IC50), reaching up to 3 to 15 µg/mL for KP3.3. AZD3152/Sipavibart neutralized JN.1.1 but lost antiviral efficacy against KP.1.1, LB.1, and KP3.3. Conclusions: Our results highlight the need for a close clinical monitoring of VYD222/Pemivibart and raise concerns about the clinical efficacy of AZD3152/Sipavibart.

Distinct evolution of SARS-CoV-2 Omicron XBB and BA.2.86/JN.1 lineages combining increased fitness and antibody evasion.

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 January 2024. 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.

Distinct evolution of SARS-CoV-2 Omicron XBB and BA.2.86/JN.1 lineages combining increased fitness and antibody evasion.

The unceasing circulation of SARS-CoV-2 leads to the continuous emergence of novel viral sublineages. Here, we isolate and characterize 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 January 2024. 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 replicate in IGROV-1 but no longer in Vero E6 and are not markedly fusogenic. They potently infect nasal epithelial cells, with EG.5.1.3 exhibiting the highest fitness. Antivirals remain active. Neutralizing antibody (NAb) responses from vaccinees and BA.1/BA.2-infected individuals are markedly lower compared to BA.1, without major differences between variants. An XBB breakthrough infection enhances NAb responses against both XBB and BA.2.86 variants. JN.1 displays 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.

Broad sarbecovirus neutralization by combined memory B cell antibodies to ancestral SARS-CoV-2.

Antibodies play a pivotal role in protecting from SARS-CoV-2 infection, but their efficacy is challenged by the continuous emergence of viral variants. In this study, we describe two broadly neutralizing antibodies cloned from the memory B cells of a single convalescent individual after infection with ancestral SARS-CoV-2. Cv2.3194, a resilient class 1 anti-RBD antibody, remains active against Omicron sub-variants up to BA.2.86. Cv2.3132, a near pan-Sarbecovirus neutralizer, targets the heptad repeat 2 membrane proximal region. When combined, Cv2.3194 and Cv2.3132 form a complementary SARS-CoV-2 neutralizing antibody cocktail exhibiting a local dose-dependent synergy. Thus, remarkably robust neutralizing memory B cell antibodies elicited in response to ancestral SARS-CoV-2 infection can withstand viral evolution and immune escape. The cooperative effect of such antibody combination may confer a certain level of protection against the latest SARS-CoV-2 variants.

Sotrovimab therapy elicits antiviral activities against Omicron BQ.1.1 and XBB.1.5 in sera of immunocompromised patients.

Antibodies effective against the recent Omicron sublineages are missing. By taking advantage of a multi-centric prospective cohort of immunocompromised individuals treated for mild-to-moderate COVID-19, Bruel et al. show that administration of 500 mg of sotrovimab induces serum neutralization and antibody-dependent cellular cytotoxicity of BQ.1.1 and XBB.1.5. Therefore, sotrovimab may remain a therapeutic option against these variants.

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

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 subvariant BQ.1.1 became predominant in many countries in December 2022. The subvariants 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) lose antiviral efficacy against BA.2.75.2 and BQ.1.1, whereas Xevudy (Sotrovimab) remaine weakly active. BQ.1.1 is also resistant to Bebtelovimab. Neutralizing titers in triply vaccinated individuals are low to undetectable against BQ.1.1 and BA.2.75.2, 4 months after boosting. A BA.1/BA.2 breakthrough infection increases these titers, which remains about 18-fold lower against BA.2.75.2 and BQ.1.1, than against BA.1. Reciprocally, a BA.5 breakthrough infection increases more efficiently neutralization against BA.5 and BQ.1.1 than against BA.2.75.2. Thus, the evolution trajectory of novel Omicron subvariants facilitates their spread in immunized populations and raises concerns about the efficacy of most available mAbs.