The emergence, impact, and evolution of human metapneumovirus variants from 2014 to 2021 in Spain.

BACKGROUND: Human metapneumovirus (HMPV) is an important aetiologic agent of respiratory tract infection (RTI). This study aimed to describe the prevalence, genetic diversity, and evolutionary dynamics of HMPV. METHODS: Laboratory-confirmed HMPV were characterised based on partial-coding G gene sequences with MEGA.v6.0. WGS was performed with Illumina, and evolutionary analyses with Datamonkey and Nextstrain. RESULTS: HMPV prevalence was 2.5%, peaking in February-April and with an alternation in the predominance of HMPV-A and -B until the emergence of SARS-CoV-2, not circulating until summer and autumn-winter 2021, with a higher prevalence and with the almost only circulation of A2c111dup. G and SH proteins were the most variable, and 70% of F protein was under negative selection. Mutation rate of HMPV genome was 6.95 × 10-4 substitutions/site/year. CONCLUSION: HMPV showed a significant morbidity until the emergence of SARS-CoV-2 pandemic in 2020, not circulating again until summer and autumn 2021, with a higher prevalence and with almost the only circulation of A2c111dup, probably due to a more efficient immune evasion mechanism. The F protein showed a very conserved nature, supporting the need for steric shielding. The tMRCA showed a recent emergence of the A2c variants carrying duplications, supporting the importance of virological surveillance.

Emergence of Delta and Omicron variants carrying resistance-associated mutations in immunocompromised patients undergoing sotrovimab treatment with long-term viral excretion.

OBJECTIVES: To monitor the early emergence of genetic mutations related to reduced susceptibility to monoclonal anti-body (mAb)-based treatment in immunocompromised patients with long-term viral excretion using whole-genome sequencing at a tertiary university hospital in Barcelona, Spain. METHODS: Serial severe acute respiratory syndrome coronavirus 2-positive samples (mid-December 2021-mid-March 2022) from eight immunosuppressed, fully vaccinated patients (for solid-organ transplantation or haematologic malignancies) with long-term viral excretion despite undergoing mAb therapy (sotrovimab) for coronavirus disease 2019 were selected. Whole-genome sequencing was performed following the ARTIC, version 4.1, protocol on the MiSeq platform. Mutations in the coding sequence of the spike protein with a frequency of ≥5% were studied. RESULTS: A total of 37 samples from the studied cases were analysed. All the cases, except one, were confirmed to have the Omicron variant BA.1; one had Delta (AY.100). Thirty-four different mutations were detected within the receptor-binding domain of the spike protein in 62.5% of patients, eight of which were not lineage related and located in the sotrovimab target epitope (P337L, E340D, E340R, E340K, E340V, E340Q, R346T and K356T). Except for P337L, all changes showed a significant increase in frequency or fixation after the administration of sotrovimab. Some of them have been associated with either reduced susceptibility to mAb therapy, such as those at position 340, or the acquisition of a new glycosylation site (346 and 356 positions). CONCLUSIONS: This study highlights the importance of monitoring for early in vivo selection of mutations associated with reduced susceptibility to mAb therapy, especially in immunocompromised patients receiving anti-viral drugs, whose immune response is not able to control viral replication, resulting in long-term viral shedding, and those receiving selective evolution pressure. Virologic surveillance of genetically resistant viruses to available anti-viral therapies is considered a priority for both patients and the community.

The frequency of defective genomes in Omicron differs from that of the Alpha, Beta and Delta variants.

The SARS-CoV-2 Omicron variant emerged showing higher transmissibility and possibly higher resistance to current COVID-19 vaccines than other variants dominating the global pandemic. In March 2020 we performed a study in clinical samples, where we found that a portion of genomes in the SARS-CoV-2 viral population accumulated deletions immediately before the S1/S2 cleavage site (furin-like cleavage site, PRRAR/S) of the spike gene, generating a frameshift and appearance of a premature stop codon. The main aim of this study was to determine the frequency of defective deletions in prevalent variants from the first to sixth pandemic waves in our setting and discuss whether the differences observed might support epidemiological proposals. The complete SARS-CoV-2 spike gene was deeply studied by next-generation sequencing using the MiSeq platform. More than 90 million reads were obtained from respiratory swab specimens of 78 COVID-19 patients with mild infection caused by the predominant variants circulating in the Barcelona city area during the six pandemic waves: B.1.5, B.1.1, B.1.177, Alpha, Beta, Delta, and Omicron. The frequency of defective genomes found in variants dominating the first and second waves was similar to that seen in Omicron, but differed from the frequencies seen in the Alpha, Beta and Delta variants. The changing pattern of mutations seen in the various SARS-CoV-2 variants driving the pandemic waves over time can affect viral transmission and immune escape. Here we discuss the putative biological effects of defective deletions naturally occurring before the S1/S2 cleavage site during adaption of the virus to human infection.

Characteristics of 24 SARS-CoV-2-Sequenced Reinfection Cases in a Tertiary Hospital in Spain.

Background: Since the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the main concern is whether reinfections are possible, and which are the associated risk factors. This study aims to describe the clinical and molecular characteristics of 24 sequence-confirmed reinfection SARS-CoV-2 cases over 1 year in Barcelona (Catalonia, Spain). Methods: Patients with > 45 days between two positive PCR tests regardless of symptoms and negative tests between episodes were initially considered as suspected reinfection cases from November 2020 to May 2021. Whole-genome sequencing (WGS) was performed to confirm genetic differences between consensus sequences and for phylogenetic studies based on PANGOLIN nomenclature. Reinfections were confirmed by the number of mutations, change in lineage, or epidemiological criteria. Results: From 39 reported suspected reinfection cases, complete viral genomes could be sequenced from both episodes of 24 patients, all were confirmed as true reinfections. With a median age of 44 years (interquartile range [IQR] 32-65), 66% were women and 58% were healthcare workers (HCWs). The median days between episodes were 122 (IQR 72-199), occurring one-third within 3 months. Reinfection episodes were frequently asymptomatic and less severe than primary infections. The absence of seroconversion was associated with symptomatic reinfections. Only one case was reinfected with a variant of concern (VOC). Conclusion: Severe acute respiratory syndrome coronavirus 2 reinfections can occur in a shorter time than previously reported and are mainly found in immunocompetent patients. Surveillance through WGS is useful to identify viral mutations associated with immune evasion.

Enterovirus D68 in Hospitalized Children, Barcelona, Spain, 2014-2021.

To determine molecular epidemiology and clinical features of enterovirus D68 (EV-D68) infections, we reviewed EV-D68-associated respiratory cases at a hospital in Barcelona, Spain, during 2014-2021. Respiratory samples were collected from hospitalized patients or outpatients with symptoms of acute respiratory tract infection or suggestive of enterovirus infection. Enterovirus detection was performed by real-time multiplex reverse transcription PCR and characterization by phylogenetic analysis of the partial viral protein 1 coding region sequences. From 184 patients with EV-D68 infection, circulating subclades were B3 (80%), D1 (17%), B2 (1%), and A (<1%); clade proportions shifted over time. EV-D68 was detected mostly in children (86%) and biennially (2016, 2018, 2021). In patients <16 years of age, the most common sign/symptom was lower respiratory tract infection, for which 11.8% required pediatric intensive care unit admission and 2.3% required invasive mechanical ventilation; neurologic complications developed in 1. The potential neurotropism indicates that enterovirus surveillance should be mandatory.

A year living with SARS-CoV-2: an epidemiological overview of viral lineage circulation by whole-genome sequencing in Barcelona city (Catalonia, Spain).

Herein, we describe the genetic diversity of circulating SARS-CoV-2 viruses by whole-genome sequencing (WGS) in Barcelona city (Catalonia, Spain) throughout the first four pandemic waves. From weeks 11/2020-24/2021, SARS-CoV-2-positive respiratory samples were randomly selected per clinical setting (80% from primary care or 20% from the hospital), age group, and week. WGS was performed following the ARTICv3 protocol on MiSeq or NextSeq2000 Illumina platforms. Nearly complete consensus sequences were used for genetic characterization based on GISAID and PANGOLIN nomenclatures. From 2475 samples, 2166 (87%) were fully sequenced (78% from primary care and 22% from hospital settings). Multiple genetic lineages were co-circulating, but four were predominant at different periods. While B.1.5 (50.68%) and B.1.1 (32.88%) were the major lineages during the first pandemic wave, B.1.177 (66.85%) and B.1.1.7 (83.80%) were predominant during the second, third, and fourth waves, respectively. Almost all (96.4%) were carrying D614G mutation in the S protein, with additional mutations that define lineages or variants. But some mutations of concern, such as E484K from B.1.351 and P.1 lineages are currently under monitoring, together with those observed in the receptor-binding domain or N-terminal domain, such as L452R and T478K from B.1.617.2 lineage. The fact that a predominant lineage was observed in each pandemic wave suggests advantageous properties over other contemporary co-circulating variants. This genetic variability should be monitored, especially when a massive vaccination campaign is ongoing because the potential selection and emergence of novel antigenic SARS-CoV-2 strains related to immunological escapement events.

Host-dependent editing of SARS-CoV-2 in COVID-19 patients.

A common trait among RNA viruses is their high capability to acquire genetic variability due to viral and host mechanisms. Next-generation sequencing (NGS) analysis enables the deep study of the viral quasispecies in samples from infected individuals. In this study, the viral quasispecies complexity and single nucleotide polymorphisms of the SARS-CoV-2 spike gene of coronavirus disease 2019 (COVID-19) patients with mild or severe disease were investigated using next-generation sequencing (Illumina platform). SARS-CoV-2 spike variability was higher in patients with long-lasting infection. Most substitutions found were present at frequencies lower than 1%, and had an A → G or T → C pattern, consistent with variants caused by adenosine deaminase acting on RNA-1 (ADAR1). ADAR1 affected a small fraction of replicating genomes, but produced multiple, mainly non-synonymous mutations. ADAR1 editing during replication rather than the RNA-dependent RNA polymerase (nsp12) was the predominant mechanism generating SARS-CoV-2 genetic variability. However, the mutations produced are not fixed in the infected human population, suggesting that ADAR1 may have an antiviral role, whereas nsp12-induced mutations occurring in patients with high viremia and persistent infection are the main source of new SARS-CoV-2 variants.

Coordinated Response to Imported Vaccine-Derived Poliovirus Infection, Barcelona, Spain, 2019-2020.

In 2019, the Public Health Agency of Barcelona, Spain, was notified of a vaccine-derived poliovirus infection. The patient had an underlying common variable immunodeficiency and no signs of acute flaccid paralysis. We describe the ongoing coordinated response to contain the infection, which included compassionate-use treatment with pocapavir.

Insights into immune evasion of human metapneumovirus: novel 180- and 111-nucleotide duplications within viral G gene throughout 2014-2017 seasons in Barcelona, Spain.

BACKGROUND: Human metapneumovirus (HMPV) is an important aetiologic agent of respiratory tract infection (RTI). This study aimed to describe its genetic diversity and clinical impact in patients attended at a tertiary university hospital in Barcelona from the 2014-2015 to the 2016-2017 seasons, focusing on the emerging duplications in G gene and their structural properties. METHODS: Laboratory-confirmed HMPV were characterised based on partial-coding F and G gene sequences with MEGA.v6.0. Computational analysis of disorder propensity, aggregation propensity and glycosylation sites in viral G predicted protein sequence were carried out. Clinical data was retrospectively reviewed and further associated to virological features. RESULTS: HMPV prevalence was 3%. The 180- and 111-nucleotide duplications occurred in A2c lineage G protein increased in prevalence throughout the study, in addition to short genetic changes observed in other HMPV lineages. The A2c G protein without duplications was calculated to protrude over F protein in 23% of cases and increased to a 39% and a 46% with the 111- and 180-nucleotide duplications, respectively. Children did not seem to be more affected by these mutant viruses, but there was a strong association of these variants to LRTI in adults. DISCUSSION: HMPV presents a high genetic diversity in all lineages. Novel variants carrying duplications might present an evolutionary advantage due to an improved steric shielding, which would have been responsible for the reported increasing prevalence and the association to LRTI in adults.

Naturally occurring SARS-CoV-2 gene deletions close to the spike S1/S2 cleavage site in the viral quasispecies of COVID19 patients.

The SARS-CoV-2 spike (S) protein, the viral mediator for binding and entry into the host cell, has sparked great interest as a target for vaccine development and treatments with neutralizing antibodies. Initial data suggest that the virus has low mutation rates, but its large genome could facilitate recombination, insertions, and deletions, as has been described in other coronaviruses. Here, we deep-sequenced the complete SARS-CoV-2 S gene from 18 patients (10 with mild and 8 with severe COVID-19), and found that the virus accumulates deletions upstream and very close to the S1/S2 cleavage site (PRRAR/S), generating a frameshift with appearance of a stop codon. These deletions were found in a small percentage of the viral quasispecies (2.2%) in samples from all the mild and only half the severe COVID-19 patients. Our results suggest that the virus may generate free S1 protein released to the circulation. We suggest that natural selection has favoured a "Don't burn down the house" strategy, in which free S1 protein may compete with viral particles for the ACE2 receptor, thus reducing the severity of the infection and tissue damage without losing transmission capability.

The high genetic similarity between rhinoviruses and enteroviruses remains as a pitfall for molecular diagnostic tools: A three-year overview.

BACKGROUND: Enteroviruses (EVs) and rhinoviruses (RVs) belong to the Enterovirus genus within the Picornaviridae family, and show genetic similarities. These viruses are related to mild diseases, but EVs infections can sometimes lead to more severe complications. Current diagnostic molecular techniques should discriminate between the four EV and the three RV species that infect humans. The aim was to revise the EV and RV PCR-confirmed specimens by sequencing for genetic characterisation. MATERIAL AND METHODS: Respiratory tract specimens were collected from patients with suspicion of respiratory infection. Respiratory viruses' laboratory-confirmation was performed by commercial multiplex real-time RT-PCR assays. Genetic characterisation of all EV and in a selection of RV was performed based on the phylogenetic analyses of partial VP1 and VP4/2 sequences, respectively. RESULTS: From 19,957 tested specimens, 309 (1.5%) were EV-positive, 2546 (12%) were RV-positive, and 233 (1%) were EV/RV co-detections. The phylogenetic analyses revealed that: among single EV detections, 177/309 (57%) were characterised as EV, 2/309 (1%) as RV, and 130/309 (42%) could not be typed; among single 1771 RV detections (Ct < 35), 1651/1771 (93%) were characterised as RV, 3/1771 (0.3%) as EV and 117/1771 (6.7%) could not be typed. Among EV/RV co-detections, 62/233 (27%) were characterised as EV, 130/233 (56%) as RV and 41/233 (18%) could not be typed. CONCLUSIONS: A diagnostic method well considered for routine laboratory-confirmation of respiratory viruses should discriminate EV and RV targets. RVs are usually associated with mild respiratory disease, but the potential relatedness of EVs to neurological complications makes their monitoring mandatory. Therefore, an accurate detection and differentiation should be required in commercial diagnostic solutions.

Recombinant CV-A6 strains related to hand-foot-mouth disease and herpangina at primary care centers (Barcelona, Spain).

Aim: To describe the genetic diversity of enteroviruses (EV) causing hand, foot and mouth disease (HFMD) and herpangina, especially of coxsackievirus (CV)-A6, from patients attended at pediatric primary care centers during the 2017-2018 season. Methods: Phylogenetic analysis of partial VP1 region was performed for genetic characterization. The complete VP1 and 3Dpol proteins were sequenced for lineage determination and detection of recombination events. Results: An 80% of samples were EV laboratory-confirmed. CV-A6 was the most detected (70%) and associated with atypical HFMD (78%). The comparison of VP1 and 3Dpol phylogenies showed evidence of recombination in three strains, in which two shifted to CV-A16 3Dpol. Conclusion: The study provides recent information regarding the nonrecombinant and recombinant EVs related to HFMD at primary care centers.

Molecular influenza surveillance at a tertiary university hospital during four consecutive seasons (2012-2016) in Catalonia, Spain.

BACKGROUND: Influenza viruses (FLUV) are continuously evolving, which explain the occurrence of seasonal influenza epidemics and the need to review the vaccine strain composition annually. The aim is to describe the genetic diversity and clinical outcomes of FLUV detected at a tertiary university hospital in Barcelona (Spain) during the 2012-2016 seasons. METHODS: The detection of FLUV from patients attended at the Emergency Department or admitted to the hospital was performed by either immunofluorescence or PCR-based assays. A specific real-time one-step multiplex RT-PCR was performed for influenza A (FLUAV) subtyping. The complete coding haemagglutinin domain 1 (HA1) and neuraminidase (NA) (2015-2016) protein sequences from a representative sampling were molecular characterised. RESULTS: A total 1774 (66.1%) FLUAV and 910 (33.9%) influenza B (FLUBV) cases were laboratory-confirmed. The hospitalisation rate was different between seasons, being the highest (81.4%) during the 2014-2015 season. FLUV were genetically close to vaccine strains except to the 2014-2015, in which most characterised A(H3N2) viruses belonged to a genetic group different from the vaccine strain. During the 2015-2016 season, B/Victoria-like viruses were the most predominant, but this component was not included in the trivalent vaccine used. Mutations D222G or D222N in HA1-domain were found in 3 A(H1N1)pdm09 strains from ICU-admitted cases. Three A(H1N1)pdm09 strains carried the NA H275Y (2) and S247N (1) mutations, respectively related to resistance or decreased susceptibility to oseltamivir. CONCLUSIONS: The circulation of drifted A(H3N2) strains during the 2014-2015 season was related to the high hospitalisation rate due to the mismatch with the vaccine strains. The predominance of a FLUBV lineage not included in the trivalent influenza vaccine during the 2015-2016 season highlights the need to use a tetravalent influenza vaccine. Virological surveillance of viral variants carrying protein changes that alter tropism and susceptibility to antivirals features should be strengthened in hospital settings.

Surveillance of enteroviruses from paediatric patients attended at a tertiary hospital in Catalonia from 2014 to 2017.

BACKGROUND: Enterovirus (EV) infections are usually asymptomatic or mild, but symptomatic infections can evolve to severe complications. Outbreaks of EV-A71 and EV-D68 have been recently reported worldwide, sometimes related to severe clinical outcomes. OBJECTIVE: To describe EV genetic diversity and the clinical outcomes from paediatric patients attended at a tertiary university hospital in Barcelona (Catalonia, Spain) from 2014 to 2017. STUDY DESIGN: Specimens were collected from paediatric (<17 years old) cases with suspicion of respiratory tract infection or EV infection. EV laboratory-confirmation was performed by specific real-time multiplex RT-PCR assay. Partial viral VP1 protein was sequenced for genetic characterisation by phylogenetic analyses. RESULTS: A total of 376 (7%) from 5703 cases were EV laboratory-confirmed. Phylogenetic analyses of VP1 (210; 81%) sequences distinguished up to 27 different EV types distributed within EV-A (82; 40%), EV-B (90; 42%), EV-C (5; 2%), and EV-D (33; 15%), in addition to 50 (19%) rhinoviruses. The most predominant were EV-A71 (37; 45%) and EV-D68 (32; 99%). EV-A71 was highly related to neurological complications (25/39, 63%), of which 20/39 were rhombencephalitis, and most EV-D68 (28/32, 88%) were associated with lower respiratory tract infections (LRTI), and exceptionally one (3%) with acute flaccid paralysis. CONCLUSIONS: EV-A71 and EV-D68 were the most detected EV in respiratory specimens. EV-A71 was highly related to neurological disease and EV-D68 was often associated with LRTI. However, both potential relatedness to neurological diseases makes the monitoring of EV circulation obligatory.

Genetic diversity of rhinoviruses detected at a tertiary hospital in Catalonia (Spain) during the 2014-2017 seasons.

AIM: To describe the genetic diversity of rhinovirus (RV) from patients attended at a tertiary hospital in Barcelona (Spain) from October 2014 to May 2017. METHODS: RV detection was performed by real-time multiplex RT-PCR. A specific real-time quantitive retrotranscription PCR (qRT-PCR) was carried out to select those samples (Ct < 35) for molecular characterization based on partial VP4/2 protein. RESULTS: Phylogenetic characterization revealed proportions of 63% RV-A, 6% RV-B and 31% RV-C (119 different types). RV-A circulated throughout all the study period, with a minor circulation during winter, just when RV-C prevailed. Differences between age medians by RV-specie were reported. CONCLUSION: The large genetic diversity of RV detected in our area is described here. The variable cocirculation of multiple RV types is also reported, showing differences by age.

Evaluation of Seegene Allplex Respiratory Panel 1 kit for the detection of influenza virus and human respiratory syncytial virus.

BACKGROUND: Influenza (FLUV) and human respiratory syncytial (HRSV) viruses are etiological agents of respiratory infections that cause a significant morbidity and mortality worldwide. A rapid and accurate diagnosis of these respiratory viruses is essential for an appropriate patient management. Molecular tests are the best detection option due to their high sensitivity and specificity. Seegene's Allplex™ Respiratory Panel 1 (Allplex RP1) is a real-time one-step RT-PCR assay for the simultaneous detection of FLUAV, FLUBV, HRSV-A and HRSV-B. In addition, it allows the determination of FLUAV subtype (H1, H3 and H1pdm09). OBJECTIVES: This study aims to evaluate Allplex RP1 as a rapid molecular test for the detection of FLUAV, FLUBV, HRSV-A and HRSV-B viruses. STUDY DESIGN: The Allplex RP1 assay will be compared with other two commercial molecular assays, Prodesse ProFlu+ and ProFAST+ (Hologic, Madison, WI, USA), and GeneXpert Flu/RSV XC (Cepheid, USA). RESULTS: Allplex RP1, ProFlu+ and GeneXpert tests showed 95%, 91% and 96% of accuracy; and 94%, 88% and 95% of sensitivity, respectively. Moreover, Allplex RP1 showed a FLUAV subtype sensitivity of 91% and 88% for FLUAV-H1pdm09 and FLUAV-H3 respectively, and ProFAST+ assay showed sensitivities of 100% for both targets. The three assays showed a 100% of specificity and PPV, while the NPV were 84%, 73% and 86% for Allplex RP1, Prodesse and GeneXpert, respectively. CONCLUSIONS: In this study, Seegene's Allplex RP1 assay showed to be highly sensitive, specific, and suitable for detection of FLUV and HRSV, including FLUAV subtyping. In addition, it is also a hands-on-time saving assay due to the automated nucleic acid extraction and PCR setup.

Novel human metapneumovirus with a 180-nucleotide duplication in the G gene.

AIM: To describe the circulation, genetic diversity and clinical features of human metapneumovirus (HMPV) in pediatric patients that attended the Hospital Universitari Vall d'Hebron, Spain from 2014 to 2016. MATERIALS & METHODS: Partial G gene was sequenced from laboratory-confirmed HMPV respiratory specimens for subsequent phylogenetic analysis. RESULTS: A total of 121 different samples were HMPV laboratory-confirmed out of 6658 specimens received. The highest circulation was from February to April, with a prevalence of 3%. Different genetic groups within both genotypes were detected at variable levels. A 180-nucleotide duplication was first characterized within the G gene in nine cases, mostly related to lower respiratory-tract infection. CONCLUSION: This study reported on the circulation of a novel HMPV with a 180-nucleotide duplication in the G gene, but no clinical changes in related cases were observed. Their prevalence increased during the last season suggesting changes in viral features.

A molecular epidemiological study of human parainfluenza virus type 3 at a tertiary university hospital during 2013-2015 in Catalonia, Spain.

Human parainfluenza virus type 3 (HPIV-3) is one of the most common respiratory viruses particularly among young children and immunocompromised patients. The seasonality, prevalence and genetic diversity of HPIV-3 at a Spanish tertiary-hospital from 2013 to 2015 are reported. HPIV-3 infection was laboratory-confirmed in 102 patients (76%, under 5 years of age). Among <5 years-old patients, 9 (11.5%) were under any degree of immunosuppression, whereas this percentage was significantly higher (19; 79.2%) among patients older than 5 years. HPIV-3 was detected at varying levels, but mainly during spring and summer. All characterized HN/F sequences fell within C1b, C5 and in other two closely C3a-related groups. Furthermore, a new genetic lineage (C1c) was described. Genetic similarity and epidemiological data confirmed some nosocomial infections, highlighting the importance of the HPIV-3 surveillance, particularly in high-risk patients. This study provides valuable information on HPIV-3 diversity due to the scarce information in Europe.

First Enterovirus D68 (EV-D68) cases detected in hospitalised patients in a tertiary care university hospital in Spain, October 2014 / Primeras detecciones de Enterovirus D68 (EV-D68) en población hospitalizada en España, Octubre 2014

Several outbreaks of Enterovirus 68 (EV-D68) have recently been reported in the USA and Canada, causing substantial hospitalisation of children with severe respiratory disease. The acute flaccid paralysis detected in the USA and Canada among children with EV-D68 infection has raised concerns about the aetiological role of this EV serotype in severe neurological disease. The circulation of EV-D68 in the general European population seems to be low, but European Centre for Disease Prevention and Control (ECDC) recommends being vigilant to new cases, particularly in severely ill hospitalised patients. In October 2014, enteroviruses were detected in respiratory samples collected from five hospitalised patients, children and adults. Phylogenetic analysis of partial VP1 sequences confirmed that the detected enteroviruses belonged to the D68 serotype, which were also similar to strains reported in USA (2014). However, all five patients developed respiratory symptoms, but only one required ICU admission. None of the patients described had symptoms of neurological disease. Other considerations related to the detection methods used for the diagnosis of respiratory enteroviruses are also discussed. In conclusion, additional evidence has been provided that supports the role of EV-D68 in respiratory infections in hospitalised patients

En EEUU y Canadá, desde el pasado verano, se han descrito varios brotes causados por EV-D68 afectando a pacientes, principalmente niños, con enfermedad respiratoria grave. En algunos de estos casos la infección por EV-D68 se ha asociado a enfermedad neurológica grave. Aunque en población europea la circulación de este serotipo parece ser baja, el ECDC recuerda la necesidad de reforzar la vigilancia, especialmente en pacientes hospitalizados. En octubre de 2014, en las muestras respiratorias de 5 pacientes ingresados en el Hospital Universitario Vall d’Hebron de Barcelona, se pudieron detectar enterovirus. Estos fueron caracterizados como EV-D68 mediante análisis filogenético de las secuencias parciales codificantes para la proteína viral VP1. Estas secuencias eran además similares a las de las cepas aisladas en los últimos meses en EEUU. Estos 5 pacientes presentaron síntomas respiratorios, pero sólo uno requirió ingreso en la Unidad de Cuidados Intensivos. Sin embargo, ninguno de los pacientes presentó síntomas de enfermedad neurológica. En este trabajo se comentan también consideraciones relacionadas con los métodos de diagnóstico para enterovirus, especialmente para este serotipo. En conclusión, en este trabajo se confirma el posible papel etiológico del EV-D68 en la infección respiratoria del paciente ingresado

Molecular epidemiology and molecular characterization of respiratory syncytial viruses at a tertiary care university hospital in Catalonia (Spain) during the 2013-2014 season.

BACKGROUND: Human respiratory syncytial virus (HRSV) is the main cause of lower respiratory tract infections among infants and young children. OBJECTIVES: The molecular epidemiology and characterization of HRSV strains detected at a Spanish tertiary hospital during the 2013-2014 season is reported. STUDY DESIGN: Phylogenetic analysis and molecular characterization of HRSV laboratory-confirmed respiratory samples were performed, based on coding sequences of G and F proteins. RESULTS: HRSV infection was laboratory-confirmed in respiratory samples from 320 patients of which 223 (70%) were less than 2 years of age and none undergoing Palivizumab treatment. HRSV was detected at varying levels throughout the season with a maximum rate in the week 52/2013, right before the beginning of the influenza epidemic. Whilst both HRSV groups were found co-circulating, HRSV-B group clearly predominated. The phylogenetic analyses from 139 HVR-2 sequences revealed that most characterized strains belonged to ON1 and BA9 genotypes. Three different phylogenetic subgroups could be distinguished within these genotypes. In addition, three strains (out of the 52 randomly selected) were carrying amino acid substitutions in the epitope A of the F protein, one of them previously related to Palivizumab resistance. CONCLUSIONS: The results of the present study highlight the importance of a continuous HRSV surveillance to monitor not only the introduction of new genotypes on circulation but also the emergence of viral variants with new genetic characteristics that can affect the antigenicity features and the susceptibility to the only current prophylaxis treatment and also for the future development of HRSV vaccines.