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.

Virological surveillance of human respiratory syncytial virus A and B at a tertiary hospital in Catalonia (Spain) during five consecutive seasons (2013-2018).

AIM: Human respiratory syncytial virus (HRSV) is the main cause of respiratory tract infections among infants. MATERIALS & METHODS: In the present study, the molecular epidemiology of HRSV detected from 2013 to 2017 has been described. RESULTS: A 10% of collected samples were laboratory confirmed for HRSV. Patients under 2 years of age were the main susceptible population to respiratory syncytial virus disease, but an increasingly number of confirmed patients over 65 years of age was reported. Epidemics usually started in autumn and ended in spring. Both HRSV groups co-circulated every season, but the HRSV-B was the most predominant. HRSV-A and HRSV-B strains mainly belonged to ON1 and BA9 genotypes, respectively. CONCLUSION: The present study reports recent data about the genetic diversity of circulating HRSV in Spain.

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.

Novel tp0548 Sequence-Type of Treponema pallidum Identified in Barcelona, Spain.

A novel tp0548 sequence-type of Treponema pallidum has been identified in a genital ulcer sample collected from a patient diagnosed with primary syphilis at the Hospital Universitari Vall d'Hebron in Barcelona. Following the nomenclature used in the Enhanced Centers for Disease Control and Prevention Typing methodology, letter "z" has been assigned to the new sequence type.

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.

Evaluation of the novel DiaSorin LIAISON(R) Campylobacter assay for the rapid detection of Campylobacter spp / Evaluación de la nueva plataforma DiaSorin LIAISON(R) Campylobacter assay para la detección rápida de Campylobacter spp

INTRODUCTION: Campylobacter spp. infection is one of the leading causes of foodborne diarrhoeal illness in humans worldwide. The purpose of this study was to evaluate the DiaSorin LIAISON(R) Campylobacter assay for human campylobacteriosis diagnosis. METHODOLOGY: A total of 645 stool samples from 640 patients suspected of having gastrointestinal infection were included. A stool culture was simultaneously performed with the DiaSorin LIAISON(R) Campylobacter assay to detect the presence of Campylobacter spp. RESULTS: Taking the conventional culture to be the perfect gold standard, sensitivity and specificity rates of the DiaSorin LIAISON(R)Campylobacter assay were 100% and 97.7%, respectively; and 99.1% and 98.6%, respectively, when taking the culture to be the imperfect gold standard (Bayesian Model). CONCLUSION: This new assay might be a useful tool especially for the screening of negative results

INTRODUCCIÓN: La infección por Campylobacter spp. es una de las principales causas de enfermedades diarreicas de transmisión alimentaria en el ser humano. Este estudio tuvo como objetivo evaluar la plataforma DiaSorin LIAISON(R) Campylobacter assay para el diagnóstico de la campilobacteriosis humana. METODOLOGÍA: Se incluyeron un total de 645 muestras de heces de 640 pacientes con sospecha de infección gastrointestinal. Se realizaron simultáneamente coprocultivo y DiaSorin LIAISON(R) Campylobacter assay para detectar la presencia de Campylobacter spp. RESULTADOS: Asumiendo el cultivo convencional como el método de referencia perfecto, DiaSorin LIAISON(R) Campylobacter assay obtuvo una sensibilidad y una especificidad del 100% y 97,7%, respectivamente; y del 99,1% y 98,6%, respectivamente, asumiendo el cultivo como método de referencia imperfecto (modelo bayesiano)

Evaluation of the novel DiaSorin LIAISON®Campylobacter assay for the rapid detection of Campylobacter spp.

INTRODUCTION: Campylobacter spp. infection is one of the leading causes of foodborne diarrhoeal illness in humans worldwide. The purpose of this study was to evaluate the DiaSorin LIAISON®Campylobacter assay for human campylobacteriosis diagnosis. METHODOLOGY: A total of 645 stool samples from 640 patients suspected of having gastrointestinal infection were included. A stool culture was simultaneously performed with the DiaSorin LIAISON®Campylobacter assay to detect the presence of Campylobacter spp. RESULTS: Taking the conventional culture to be the perfect gold standard, sensitivity and specificity rates of the DiaSorin LIAISON®Campylobacter assay were 100% and 97.7%, respectively; and 99.1% and 98.6%, respectively, when taking the culture to be the imperfect gold standard (Bayesian Model). CONCLUSION: This new assay might be a useful tool especially for the screening of negative results.

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.

Imported cases of Chikungunya in Barcelona in relation to the current American outbreak.

BACKGROUND: The Chikungunya virus (CKIKV) is currently present in America. Travel between America and Europe is particularly intense and one of the main vectors of CHIKV, Aedes albopictus, is well established in the Mediterranean basin. We describe a series of imported cases that could originate a European outbreak. METHODS: We retrospectively studied cases of CHIKV originating in America and diagnosed in the last year in three Tropical Medicine Units of Barcelona of the International Health Program of the Catalan Health Institute (PROSICS). Clinical, microbiological and epidemiological data were analyzed. RESULTS: Forty-two CHIKV cases who had returned from 11 American countries were included. Fever was the most common symptom at onset (96.1%). Three months after symptom onset 50% continued with arthralgias, 35.3% fatigue and 11.8% arthritis. Three patients were viremic at the time of diagnosis by RT-PCR, and the remaining were diagnosed by serology (CHIKV IgM or IgG). Five (11.9%) patients had positive IgM for both dengue virus and CHIKV. CONCLUSIONS: The origin of the cases was diverse, the most frequent being initially the Dominican Republic, followed later by Venezuela and Colombia. Symptoms were not severe but persisted, accompanied by unremitting positive IgM. Diagnosis was mainly based on serology and RT-PCR, with the performance of the rapid immunochromatographic test being low. Phylogenetic studies showed that two viremic cases were caused by a strain of Asian lineage with a lower adaptability to Aedes albopictus. Co-infection with the dengue virus was common, but the clinical course was not affected by coinfection. Non-steroidal anti-inflammatory drugs were administered to 71.4% and steroids to 21.4%. The number of imported cases of CHIKV in Spain is rising due to introduction of this virus in America, and this could lead to an autochthonous outbreak if Public Health measures are not taken.

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.

First Enterovirus D68 (EV-D68) cases detected in hospitalised patients in a tertiary care university hospital in Spain, October 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.