Broadly neutralizing antibodies reactive against most and even all variants of the same viral species have been described for influenza and HIV-1 (ref. 1). However, whether a neutralizing antibody could have the breadth of range to target different viral species was unknown. Human respiratory syncytial virus (HRSV) and human metapneumovirus (HMPV) are common pathogens that cause severe disease in premature newborns, hospitalized children and immune-compromised patients, and play a role in asthma exacerbations. Although antisera generated against either HRSV or HMPV are not cross-neutralizing, we speculated that, because of the repeated exposure to these viruses, cross-neutralizing antibodies may be selected in some individuals. Here we describe a human monoclonal antibody (MPE8) that potently cross-neutralizes HRSV and HMPV as well as two animal paramyxoviruses: bovine RSV (BRSV) and pneumonia virus of mice (PVM). In its germline configuration, MPE8 is HRSV-specific and its breadth is achieved by somatic mutations in the light chain variable region. MPE8 did not result in the selection of viral escape mutants that evaded antibody targeting and showed potent prophylactic efficacy in animal models of HRSV and HMPV infection, as well as prophylactic and therapeutic efficacy in the more relevant model of lethal PVM infection. The core epitope of MPE8 was mapped on two highly conserved anti-parallel β-strands on the pre-fusion viral F protein, which are rearranged in the post-fusion F protein conformation. Twenty-six out of the thirty HRSV-specific neutralizing antibodies isolated were also found to be specific for the pre-fusion F protein. Taken together, these results indicate that MPE8 might be used for the prophylaxis and therapy of severe HRSV and HMPV infections and identify the pre-fusion F protein as a candidate HRSV vaccine.

Background Monitoring the adaptive immune responses during the natural course of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection provides useful information for the development of vaccination strategies against this virus and its emerging variants. We thus profiled the serum anti-SARS-CoV-2 antibody levels and specific memory B- and T-cell responses in convalescent coronavirus disease-2019 (COVID-19) patients. Methods Altogether 119 samples from 88 convalescent donors who experienced mild to critical disease were tested for the presence of elevated anti-spike and anti-receptor binding domain antibody levels over a period of eight months. In addition, level of SARS-CoV-2 neutralizing antibodies, specific memory B- and T-cell responses were tested in a subset of samples. Findings Anti-SARS-CoV-2 antibodies were present in 85% of the samples collected within 4 weeks after onset of symptoms in COVID-19 patients. Levels of specific IgM/IgA antibodies declined after 1 month while levels of specific IgG antibodies and plasma neutralizing activities remained relatively stable up to 6 months after diagnosis. Anti-SARS-CoV-2 IgG antibodies were still present, though at a significantly lower level, in 80% of the samples collected at 6-8 months after symptom onset. SARS-CoV-2-specific memory B- and T-cell responses developed with time and were persistent in all patients followed up till 6-8 months. Conclusions Our data suggest that protective adaptive immunity following natural infection of SARS-CoV-2 might persist for at least 6-8 months, regardless of disease severity. Development of medium or long-term protective immunity through vaccination might thus be possible. Funding EU-ATAC consortium, the Italian Ministry of Health and SciLife/KAW.

Enteroviruses (EV) can cause severe neurological and respiratory infections, and occasionally lead to devastating outbreaks as previously demonstrated with EV-A71 and EV-D68 in Europe. However, these infections are still often underdiagnosed and EV typing data is not currently collected at European level. In order to improve EV diagnostics, collate data on severe EV infections and monitor the circulation of EV types, we have established European non-polio enterovirus network (ENPEN). First task of this cross-border network has been to ensure prompt and adequate diagnosis of these infections in Europe, and hence we present recommendations for non-polio EV detection and typing based on the consensus view of this multidisciplinary team including experts from over 20 European countries. We recommend that respiratory and stool samples in addition to cerebrospinal fluid (CSF) and blood samples are submitted for EV testing from patients with suspected neurological infections. This is vital since viruses like EV-D68 are rarely detectable in CSF or stool samples. Furthermore, reverse transcriptase PCR (RT-PCR) targeting the 5'noncoding regions (5'NCR) should be used for diagnosis of EVs due to their sensitivity, specificity and short turnaround time. Sequencing of the VP1 capsid protein gene is recommended for EV typing; EV typing cannot be based on the 5'NCR sequences due to frequent recombination events and should not rely on virus isolation. Effective and standardized laboratory diagnostics and characterisation of circulating virus strains are the first step towards effective and continuous surveillance activities, which in turn will be used to provide better estimation on EV disease burden.

Neutralizing antibodies that target the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein are among the most promising approaches against COVID-19 1,2 . A bispecific IgG1-like molecule (CoV-X2) has been developed on the basis of C121 and C135, two antibodies derived from donors who had recovered from COVID-19 3 . Here we show that CoV-X2 simultaneously binds two independent sites on the RBD and, unlike its parental antibodies, prevents detectable spike binding to the cellular receptor of the virus, angiotensin-converting enzyme 2 (ACE2). Furthermore, CoV-X2 neutralizes wild-type SARS-CoV-2 and its variants of concern, as well as escape mutants generated by the parental monoclonal antibodies. We also found that in a mouse model of SARS-CoV-2 infection with lung inflammation, CoV-X2 protects mice from disease and suppresses viral escape. Thus, the simultaneous targeting of non-overlapping RBD epitopes by IgG-like bispecific antibodies is feasible and effective, and combines the advantages of antibody cocktails with those of single-molecule approaches.The COVID-19 pandemic has prompted substantial efforts to develop effective countermeasures against SARS-CoV-2. Preclinical data and phase-III clinical studies indicate that monoclonal antibodies could be effectively deployed for prevention or treatment during the viral symptoms phase of the disease 1,2 . Cocktails of two or more monoclonal antibodies are preferred over a single antibody as these cocktails result in increased efficacy and the prevention of viral escape. However, this approach requires increased manufacturing costs and volumes, which are problematic at a time when the supply chain is under pressure to meet the high demand for COVID-19 therapeutic agents, vaccines and biologics in general 4 . Cocktails also complicate formulation 5,6 and hinder strategies such as antibody delivery by viral vectors or by nonvectored nucleic acids 7,8 . One alternative is to use multispecific antibodies, which have the advantages of cocktails and single-molecule strategies.To this end, we used structural information 9 and computational simulations to design bispecific antibodies that would simultaneously bind to (i) independent sites on the same RBD and (ii) distinct RBDs on a spike (S) trimer. We evaluated several designs using atomistic molecular dynamics simulations, and produced four constructs: of these, CoV-X2 was the most potent neutralizer of SARS-CoV-2 pseudovirus, and had a half-maximal inhibitory concentration (IC 50 ) of 0.04 nM (5.8 ng ml −1 ) (Extended Data Fig. 1). CoV-X2 is a human-derived IgG1-like bispecific antibody in the CrossMAb format 10 that is the result of the combination of the Fragment antigen binding (Fab) of the monoclonal antibodies C121 and C135, which are two potent neutralizers of SARS-CoV-2 3 . Structural predictions showed that CoV-X2-but not its parental monoclonal antibodies-can bind bivalently to all RBD conformations on the S trimer, which prevents the binding of ACE2 receptor 11 (Fig. 1a, Extended Data Fig. 2).CoV-X2 bou...

Objectives: To detect possible severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) RNA contamination of inanimate surfaces in areas at high risk of aerosol formation by patients with coronavirus disease 2019 (COVID-19). Methods: Sampling was performed in the emergency unit and the sub-intensive care ward. SARS-CoV-2 RNA was extracted from swabbed surfaces and objects and subjected to real-time RT-PCR targeting RNAdependent RNA polymerase and E genes. Virus isolation from positive samples was attempted in vitro on Vero E6 cells. Results: Twenty-six samples were collected and only two were positive for low-level SARS-CoV-2 RNA, both collected on the external surface of continuous positive airway pressure helmets. All transport media were inoculated onto susceptible cells, but none induced a cytopathic effect on day 7 of culture. Conclusions: Even though daily contact with inanimate surfaces and patient fomites in contaminated areas may be a medium of infection, our data obtained in real-life conditions suggest that it might be less extensive than hitherto recognized.

In order to investigate the genetic diversity and patterns of the co-circulating genotypes of respiratory syncytial virus (RSV) and their possible relationships with the severity of RSV infection, we studied all of the RSV-positive nasopharyngeal samples collected from children during five consecutive winters (2009–2010, 2010–2011, 2011–2012, 2012–2013 and 2013–2014). The RSVs were detected using the respiratory virus panel fast assay and single-tube RT-PCR, their nucleotides were sequenced, and they were tested for positive selection. Of the 165 positive samples, 131 (79.4%) carried RSV-A and 34 (20.6%) RSV-B; both groups co-circulated in all of the study periods, with RSV-A predominating in all the seasons except for winter 2010–2011, which had a predominance of RSV-B. Phylogenetic analysis of the RSV-A sequences identified genotypes NA1 and ON1, the second replacing the first during the last two years of the study period. The RSV-B belonged to genotypes BA9 and BA10. BA9 was detected in all the years of the study whereas BA only desultorily. Comparison of the subjects infected by RSV-A and RSV-B types did not reveal any significant differences, but the children infected by genotype A/NA1 more frequently had lower respiratory tract infections (p<0.0001) and required hospitalisation (p = 0.007) more often than those infected by genotype A/ON1. These findings show that RSV has complex patterns of circulation characterised by the periodical replacement of the predominant genotypes, and indicate that the circulation and pathogenic role of the different RSV strains should be investigated as each may have a different impact on the host. A knowledge of the correlations between types, genotypes and disease severity may also be important in order to be able to include the more virulent strains in future vaccines.

We describe clinical characteristics, treatments and outcomes of 44 Caucasian patients with coronavirus disease (COVID-19) at a single hospital in Pavia, Italy, from 21–28 February 2020, at the beginning of the outbreak in Europe. Seventeen patients developed severe disease, two died. After a median of 6 days, 14 patients were discharged from hospital. Predictors of lower odds of discharge were age > 65 years, antiviral treatment and for severe disease, lactate dehydrogenase > 300 mg/dL.

HKU1 hCoV strains circulated in northern Italy during the winter-spring season 2005-2006. Both HKU1 genotypes were detected. HKU1-specific MAb may contribute to the rapid diagnosis of HKU1 infections currently performed by RT-PCR.