Nasopharyngeal microbiota in children is associated with severe asthma exacerbations.

BACKGROUND: The respiratory microbiome has been associated with the etiology and disease course of asthma. OBJECTIVE: We sought to assess the nasopharyngeal microbiota in children with a severe asthma exacerbation and their associations with medication, air quality, and viral infection. METHODS: A cross-sectional study was performed among children aged 2 to 18 years admitted to the medium care unit (MCU; n = 84) or intensive care unit (ICU; n = 78) with an asthma exacerbation. For case-control analyses, we matched all cases aged 2 to 6 years (n = 87) to controls in a 1:2 ratio. Controls were participants of either a prospective case-control study or a longitudinal birth cohort (n = 182). The nasopharyngeal microbiota was characterized by 16S-rRNA-gene sequencing. RESULTS: Cases showed higher Shannon diversity index (ICU and MCU combined; P = .002) and a distinct microbial community composition when compared with controls (permutational multivariate ANOVA R2 = 1.9%; P < .001). We observed significantly higher abundance of Staphylococcus and "oral" taxa, including Neisseria, Veillonella, and Streptococcus spp. and a lower abundance of Dolosigranulum pigrum, Corynebacterium, and Moraxella spp. (MaAsLin2; q < 0.25) in cases versus controls. Furthermore, Neisseria abundance was associated with more severe disease (ICU vs MCU MaAslin2, P = .03; q = 0.30). Neisseria spp. abundance was also related with fine particulate matter exposure, whereas Haemophilus and Streptococcus abundances were related with recent inhaled corticosteroid use. We observed no correlations with viral infection. CONCLUSIONS: Our results demonstrate that children admitted with asthma exacerbations harbor a microbiome characterized by overgrowth of Staphylococcus and "oral" microbes and an underrepresentation of beneficial niche-appropriate commensals. Several of these associations may be explained by (environmental or medical) exposures, although cause-consequence relationships remain unclear and require further investigations.

Experiences of adolescents and youth with HIV testing and linkage to care through the Red Carpet Program (RCP) in Kenya.

Adolescents and youth living with HIV (AYLHIV) experience worse health outcomes compared to adults. We aimed to understand the experiences of AYLHIV in care in the youth-focused Red-Carpet program in Kenya to assess the quality of service provision and identify programmatic areas for optimization. We conducted focus group discussions among 39 AYLHIV (15-24 years) and structured analysis into four thematic areas. Within the HIV testing theme, participants cited fear of positive results, confidentiality and stigma concerns, and suggested engaging the community and youth in HIV testing opportunities. Within the HIV treatment adherence theme, participants cited forgetfulness, stigma, adverse side effects, lack of family support, and treatment illiteracy as barriers to adherence. Most participants reported positive experiences with healthcare providers and peer support. In terms of the HIV status disclosure theme, AYLHIV cited concerns about their future capacity to conceive children and start families and discussed challenges with understanding HIV health implications and sharing their status with friends and partners. Youth voices informing service implementation are essential in strengthening our capacity to optimize the support for AYLHIV within the community, at schools and healthcare facilities.

Hemagglutinin stability as a key determinant of influenza A virus transmission via air.

To cause pandemics, zoonotic respiratory viruses need to adapt to replication in and spread between humans, either via (indirect or direct) contact or through the air via droplets and aerosols. To render influenza A viruses transmissible via air, three phenotypic viral properties must change, of which receptor-binding specificity and polymerase activity have been well studied. However, the third adaptive property, hemagglutinin (HA) acid stability, is less understood. Recent studies show that there may be a correlation between HA acid stability and virus survival in the air, suggesting that a premature conformational change of HA, triggered by low pH in the airways or droplets, may render viruses noninfectious before they can reach a new host. We here summarize available data from (animal) studies on the impact of HA acid stability on airborne transmission and hypothesize that the transmissibility of other respiratory viruses may also be impacted by an acidic environment in the airways.

Adoption is not associated with immunological and virological outcomes in children with perinatally acquired HIV infection in the Netherlands.

OBJECTIVES: To provide an overview of the demographics, treatment characteristics and long-term outcomes of children with perinatal HIV-1 infection (PHIV) living in the Netherlands (NL) and to specifically investigate whether outcomes differ by children's adoption status. DESIGN: A prospective population-based open cohort including children with PHIV in NL. METHODS: We included children with PHIV who had entered HIV care in NL since 2007, in view of a sharp increase in the number of adopted children with PHIV since that year. We compared the proportion with virologic suppression and CD4+T-cell count over time between the following groups of children with PHIV: adopted and born outside NL, non-adopted born in NL, and non-adopted born outside NL, using generalized estimating equations and linear mixed effects models, respectively. To account for the variation in cohort inclusion, we analyzed data of children exposed to at least one year of antiretroviral therapy (ART). RESULTS: We included 148 children (827.5 person-years of follow-up, 72% adopted, age at start care in NL 2.4 (0.5-5.3)). Under-18 mortality was zero. Over the years, a boosted PI-based regimen was most often prescribed. The use of integrase inhibitors increased since 2015. Non-adopted children born in NL were less likely to achieve virological suppression compared to adopted children (OR 0.66, 95%CI 0.51-0.86, p = 0.001), which disappeared after excluding one child with suspected treatment nonadherence (OR 0.85, 95%CI 0.57-1.25, p = 0.400). CD4+T-cell Z-score trajectories were not significantly different between groups. CONCLUSIONS: Despite considerable and increasing diversity of the population of children with PHIV in NL, geographical origin and adoption status do not seem to pose important challenges in achieving good immunological and virological outcomes.

Reduced Seasonal Coronavirus Antibody Responses in Children Following COVID-19 Mitigation Measures, The Netherlands.

SARS-CoV-2 prevention and control measures did not only impact SARS-CoV-2 circulation, but also the timing and prevalence of other seasonal respiratory viruses. Especially in children, information on exposure and infections to seasonal coronaviruses as well as SARS-CoV-2 in the first year of the pandemic is largely lacking. Therefore, we set up a one-year serological survey in a large tertiary hospital in the Netherlands. We show that seasonal coronavirus seroprevalence significantly decreased in 2021 in children less than one year, most likely due to COVID-19 control measures. The SARS-CoV-2 seroprevalence in children and adolescents increased from 0.4% to 11.3%, the highest in adolescents. This implies higher exposure rates in adolescents as compared to the general population (>18 years old). It is clear that there have been significant changes in the circulation and subsequent immunity against most respiratory pathogens as a result of the mitigation measures. The implications on shorter as well as longer term are still largely unknown, but the impact of the SARS-CoV-2 pandemic and subsequent control measures will continue to affect the dynamics of other pathogens.

Emergence and Potential Extinction of Genetic Lineages of Human Metapneumovirus between 2005 and 2021.

Human metapneumovirus (HMPV) is one of the leading causes of respiratory illness (RI), primarily in infants. Worldwide, two genetic lineages (A and B) of HMPV are circulating that are antigenically distinct and can each be further divided into genetic sublineages. Surveillance combined with large-scale whole-genome sequencing studies of HMPV are scarce but would help to identify viral evolutionary dynamics. Here, we analyzed 130 whole HMPV genome sequences obtained from samples collected from individuals hospitalized with RI and partial fusion (n = 144) and attachment (n = 123) protein gene sequences obtained from samples collected from patients with RI visiting general practitioners between 2005 and 2021 in the Netherlands. Phylogenetic analyses demonstrated that HMPV continued to group in the four sublineages described in 2004 (A1, A2, B1, and B2). However, one sublineage (A1) was no longer detected in the Netherlands after 2006, while the others continued to evolve. No differences were observed in dominant (sub)lineages between samples obtained from patients with RI being hospitalized and those consulting general practitioners. In both populations, viruses of lineage A2 carrying a 180-nucleotide or 111-nucleotide duplication in the attachment protein gene became the most frequently detected genotypes. In the past, different names for the newly energing lineages have been proposed, demonstrating the need for a consistent naming convention. Here, criteria are proposed for the designation of new genetic lineages to aid in moving toward a systematic HMPV classification. IMPORTANCE Human metapneumovirus (HMPV) is one of the major causative agents of human respiratory tract infections. Monitoring of virus evolution could aid toward the development of new antiviral treatments or vaccine designs. Here, we studied HMPV evolution between 2005 and 2021, with viruses obtained from samples collected from hospitalized individuals and patients with respiratory infections consulting general practitioners. Phylogenetic analyses demonstrated that HMPV continued to group in the four previously described sublineages (A1, A2, B1, and B2). However, one sublineage (A1) was no longer detected after 2006, while the others continued to evolve. No differences were observed in dominant (sub)lineages between patients being hospitalized and those consulting general practitioners. In both populations, viruses of lineage A2 carrying a 180-nucleotide or 111-nucleotide duplication in the attachment protein gene became the most frequently detected genotypes. These data were used to propose criteria for the designation of new genetic lineages to aid toward a systematic HMPV classification.

SARS-CoV-2 outbreaks in secondary school settings in the Netherlands during fall 2020; silent circulation.

BACKGROUND: In fall 2020 when schools in the Netherlands operated under a limited set of COVID-19 measures, we conducted outbreaks studies in four secondary schools to gain insight in the level of school transmission and the role of SARS-CoV-2 transmission via air and surfaces. METHODS: Outbreak studies were performed between 11 November and 15 December 2020 when the wild-type variant of SARS-CoV-2 was dominant. Clusters of SARS-CoV-2 infections within schools were identified through a prospective school surveillance study. All school contacts of cluster cases, irrespective of symptoms, were invited for PCR testing twice within 48 h and 4-7 days later. Combined NTS and saliva samples were collected at each time point along with data on recent exposure and symptoms. Surface and active air samples were collected in the school environment. All samples were PCR-tested and sequenced when possible. RESULTS: Out of 263 sampled school contacts, 24 tested SARS-CoV-2 positive (secondary attack rate 9.1%), of which 62% remained asymptomatic and 42% had a weakly positive test result. Phylogenetic analysis on 12 subjects from 2 schools indicated a cluster of 8 and 2 secondary cases, respectively, but also other distinct strains within outbreaks. Of 51 collected air and 53 surface samples, none were SARS-CoV-2 positive. CONCLUSION: Our study confirmed within school SARS-CoV-2 transmission and substantial silent circulation, but also multiple introductions in some cases. Absence of air or surface contamination suggests environmental contamination is not widespread during school outbreaks.

HIV-1 resistance against dolutegravir fluctuates rapidly alongside erratic treatment adherence: a case report.

OBJECTIVES: We report a case of incomplete HIV-1 suppression on a dolutegravir, lamivudine, and abacavir single-tablet regimen with the emergence of the H51Y and G118R integrase resistance mutations. METHODS: Integrase sequencing was performed retrospectively by Sanger and next-generation sequencing. Rates of emergence and decline of resistance mutations were calculated using next-generation sequencing data. Dolutegravir plasma concentrations were measured by ultra-performance liquid chromatography-tandem mass spectrometry. The effects of H51Y and G118R on infectivity, fitness, and susceptibility to dolutegravir were quantified using cell-based assays. RESULTS: During periods of non-adherence to treatment, mutations were retrospectively documented only by next-generation sequencing. Misdiagnosis by Sanger sequencing was caused by the rapid decline of mutant strains within the retroviral population. This observation was also true for a M184V lamivudine-resistant reverse transcriptase mutation found in association with integrase mutations on single HIV genomes. Resistance rebound upon treatment re-initiation was swift (>8000 copies per day). Next-generation sequencing indicated cumulative adherence to treatment. Compared to WT HIV-1, relative infectivity was 73%, 38%, and 43%; relative fitness was 100%, 35%, and 10% for H51Y, G118R, and H51Y+G118R viruses, respectively. H51Y did not change the susceptibility to dolutegravir, but G188R and H51Y+G118R conferred 7- and 28-fold resistance, respectively. CONCLUSION: This case illustrates how poorly-fit drug-resistant viruses wax and wane alongside erratic treatment adherence and are easily misdiagnosed by Sanger sequencing. We recommend next-generation sequencing to improve the clinical management of incomplete virological suppression with dolutegravir.

Adequate exposure of 50 mg dolutegravir in children weighing 20 to 40 kg outside of sub-Sahara Africa.

Dolutegravir 50 mg is registered for use in children weighing 20-40 kg. This approval is based on data from an African paediatric cohort, and no pharmacokinetic data was available from children outside of Africa. This study provides further evidence of the effective use of dolutegravir 50 mg in children weighing 20 to 40 kg by showing that concentration data gathered in clinical practice shows adequate concentration levels in Dutch children without a safety signal.

Profiling of humoral immune responses to norovirus in children across Europe.

Norovirus is a leading cause of epidemic acute gastroenteritis. More than 30 genotypes circulate in humans, some are common, and others are only sporadically detected. Here, we investigated whether serology can be used to determine which genotypes infect children. We established a multiplex protein microarray with structural and non-structural norovirus antigens that allowed simultaneous antibody testing against 30 human GI and GII genotypes. Antibody responses of sera obtained from 287 children aged < 1 month to 5.5 years were profiled. Most specific IgG and IgA responses were directed against the GII.2, GII.3, GII.4, and GII.6 capsid genotypes. While we detected antibody responses against rare genotypes, we found no evidence for wide circulation. We also detected genotype-specific antibodies against the non-structural proteins p48 and p22 in sera of older children. In this study, we show the age-dependent antibody responses to a broad range of norovirus capsid and polymerase genotypes, which will aid in the development of vaccines.

Case numbers of acute hepatitis of unknown aetiology among children in 24 countries up to 18 April 2022 compared to the previous 5 years.

An increase of acute hepatitis of unknown aetiology has been reported among children in multiple countries worldwide. With a rapid online survey among hospitals in and outside of Europe, we describe case numbers recorded from 1 January to 18 April 2022 vs the previous 5 years. Of 24 countries that responded, we identified 5/17 European and 1/7 non-European countries with an elevation in probable cases of unexplained acute hepatitis, and severe cases were elevated in five European countries.

Prevalence and Impact of Fatigue in Children with Primary Immunodeficiency Disorders: a Quantitative Single-Center Study.

Although fatigue is a common symptom in adult patients with primary immunodeficiencies (PID), data in pediatric patients are limited. The goal of this study is to estimate the prevalence and impact of fatigue in children with PID as reported by patients, parents, and health-care providers. A retrospective single-center observational study was performed. Prevalence of fatigue was measured by reviewing medical charts of 54 children in our department who are on immunoglobulin replacement therapy. Both prevalence and impact were also measured by the PedsQL-Multidimensional Fatigue Scale (MFS) in 27 patients and 32 of their parents. This is an age-appropriate questionnaire for self-report of fatigue symptoms in patients aged 5-18 years and for parent proxy reports for patients aged 2-18 years. General, cognitive, and sleep-rest fatigue was measured, and a total fatigue score was calculated. Means, standard deviation and Z scores were calculated using age-specific reference values. Intraclass correlation coefficients (ICC) were calculated for comparison of scores provided by parents vs children's self-reported scores. Both chart review data and PedsQL-MFS showed fatigue rates of 65%. Pediatric PID patients of all ages had significantly lower scores on all subscales and total score of the PedsQL-MFS compared to healthy children, indicating greater perceived symptoms of fatigue. General fatigue was the most affected subscale in PID patients, suggesting that fatigue in these patients is mainly physical. Seventy-four percent of PID patients had a Z score lower than - 1 on the general fatigue subscale indicating severe fatigue. Child-parent concordance varied between 0.24 and 0.93. Our results show the feasibility of the PedsQL-MFS survey to evaluate the prevalence and severity of fatigue in children with PID and underscore the importance of this issue in our patient care.

Clinical and In Vitro Evidence Favoring Immunoglobulin Treatment of a Chronic Norovirus Infection in a Patient With Common Variable Immunodeficiency.

BACKGROUND: Immunocompromised individuals can become chronically infected with norovirus, but effective antiviral therapies are not yet available. METHODS: Treatments with nitazoxanide, ribavirin, interferon alpha-2a, and nasoduodenally administered immunoglobulins were evaluated sequentially in an immunocompromised patient chronically infected with norovirus. In support, these components were also applied to measure norovirus inhibition in intestinal enteroid cultures in vitro. Viral RNA levels were determined in fecal and plasma samples during each treatment and viral genomes were sequenced. RESULTS: None of the antivirals resulted in a reduction of viral RNA levels in feces or plasma. However, during ribavirin treatment, there was an increased accumulation of virus genome mutations. In vitro, an effect of interferon alpha-2a on virus replication was observed and a genetically related strain was neutralized effectively in vitro using immunoglobulins and post-norovirus-infection antiserum. In agreement, after administration of immunoglobulins, the patient cleared the infection. CONCLUSIONS: Intestinal enteroid cultures provide a relevant system to evaluate antivirals and the neutralizing potential of immunoglobulins. We successfully treated a chronically infected patient with immunoglobulins, despite varying results reported by others. This case study provides in-depth, multifaceted exploration of norovirus treatment that can be used as a guidance for further research towards norovirus treatments.

Supporting adolescents living with HIV within boarding schools in Kenya.

Adolescents and youth living with HIV (AYLHIV) are a uniquely vulnerable population facing challenges around adherence, disclosure of HIV status and stigma. Providing school-based support for AYLHIV offers an opportunity to optimize their health and wellbeing. The purpose of this study was to evaluate the feasibility of school-based supportive interventions for AYLHIV in Kenya. From 2016-2019, with funding from ViiV Healthcare, the Elizabeth Glaser Pediatric AIDS Foundation implemented the innovative Red Carpet Program (RCP) for AYLHIV in participating public healthcare facilities and boarding schools in Homa Bay and Turkana Counties in Kenya. In this analysis, we report the implementation of the school-based interventions for AYLHIV in schools, which included: a) capacity building for overall in-school HIV, stigma and sexual and reproductive health education; b) HIV care and treatment support; c) bi-directional linkages with healthcare facilities; and d) psychosocial support (PSS). Overall, 561 school staff and 476 school adolescent health advocates received training to facilitate supportive environments for AYLHIV and school-wide education on HIV, stigma, and sexual and reproductive health. All 87 boarding schools inter-linked to 66 regional healthcare facilities to support care and treatment of AYLHIV. Across all RCP schools, 546 AYLHIV had their HIV status disclosed to school staff and received supportive care within schools, including treatment literacy and adherence counselling, confidential storage and access to HIV medications. School-based interventions to optimize care and treatment support for AYLHIV are feasible and contribute to advancing sexual and reproductive health within schools.

Neuropsychological and Psychosocial Functioning of Children with Perinatal HIV-Infection in The Netherlands.

Advances in antiretroviral treatment improved the life expectancy of perinatally HIV-infected children. However, growing up with HIV provides challenges in daily functioning. This cross-sectional cohort study investigated the neuropsychological and psychosocial functioning of a group of perinatally HIV-infected children in the Netherlands and compared their outcomes with Dutch normative data and outcomes of a control group of uninfected siblings. The children's functioning was assessed with internationally well-known and standardized questionnaires, using a multi-informant approach, including the perspectives of caregivers, teachers, and school-aged children. In addition, we explored the associations of socio-demographic and medical characteristics of the HIV-infected children with their neuropsychological and psychosocial functioning. Caregivers reported compromised functioning when compared to Dutch normative data for HIV-infected children in the areas of attention, sensory processing, social-emotional functioning, and health-related quality of life. Teachers reported in addition compromised executive functioning for HIV-infected children. A comparison with siblings revealed differences in executive functioning, problems with peers, and general health. The concurrent resemblance between HIV-infected children and siblings regarding problems in other domains implies that social and contextual factors may be of influence. A family-focused approach with special attention to the child's socio-environmental context and additional attention for siblings is recommended.

Unique Severe COVID-19 Placental Signature Independent of Severity of Clinical Maternal Symptoms.

BACKGROUND: Although the risk for transplacental transmission of SARS-CoV-2 is rare, placental infections with adverse functional consequences have been reported. This study aims to analyse histological placental findings in pregnancies complicated by SARS-CoV-2 infection and investigate its correlation with clinical symptoms and perinatal outcomes. We want to determine which pregnancies are at-risk to prevent adverse pregnancy outcomes related to COVID-19 in the future. METHODS: A prospective, longitudinal, multicentre, cohort study. All pregnant women presenting between April 2020 and March 2021 with a nasopharyngeal RT-PCR-confirmed SARS-CoV-2 infection were included. Around delivery, maternal, foetal and placental PCR samples were collected. Placental pathology was correlated with clinical maternal characteristics of COVID-19. RESULTS: Thirty-six patients were included, 33 singleton pregnancies (n = 33, 92%) and three twin pregnancies (n = 3, 8%). Twenty-four (62%) placentas showed at least one abnormality. Four placentas (4/39, 10%) showed placental staining positive for the presence of SARS-CoV-2 accompanied by a unique combination of diffuse, severe inflammatory placental changes with massive perivillous fibrin depositions, necrosis of syncytiotrophoblast, diffuse chronic intervillositis, and a specific, unprecedented CD20+ B-cell infiltration. This SARS-CoV-2 placental signature seems to correlate with foetal distress (75% vs. 15.6%, p = 0.007) but not with the severity of maternal COVID-19 disease. CONCLUSION: We describe a unique placental signature in pregnant patients with COVID-19, which has not been reported in a historical cohort. We show that the foetal environment can be seriously compromised by disruption of placental function due to local, devastating SARS-CoV-2 infection. Maternal clinical symptoms did not predict the severity of the SARS-CoV-2-related placental signature, resulting in a lack of adequate identification of maternal criteria for pregnancies at risk. Close foetal monitoring and pregnancy termination in case of foetal distress can prevent adverse pregnancy outcomes due to COVID-19 related placental disease.

Small quantities of respiratory syncytial virus RNA only in large droplets around infants hospitalized with acute respiratory infections.

BACKGROUND: Respiratory syncytial virus (RSV) is a major cause of respiratory tract infections in young children. The predominant transmission routes for RSV are still a matter of debate. Specifically, it remains unclear if RSV can be transmitted through the air and what the correlation is between the amount of RSV in nasopharynx samples and in the air. METHODS: The amount of RSV in the air around hospitalized RSV infected infants in single-patient rooms was quantified using a six-stage Andersen cascade impactor that collects and fractionates aerosols and droplets according to size. RSV shedding in the nasopharynx of patients was followed longitudinally by quantifying RSV RNA levels and infectious virus in nasopharyngeal aspirates. Nose and throat swabs of parents and swabs of the patient's bedrail and a datalogger were also collected. RESULTS: Patients remained RSV positive during the air sampling period and infectious virus was isolated up to 9 days post onset of symptoms. In three out of six patients, low levels of RSV RNA, but no infectious virus, were recovered from impactor collection plates that capture large droplets > 7 µm. For four of these patients, one or both parents were also positive for RSV. All surface swabs were RSV-negative. CONCLUSIONS: Despite the prolonged detection of infectious RSV in the nasopharynx of patients, only small amounts of RSV RNA were collected from the air around three out of six patients, which were primarily contained in large droplets which do not remain suspended in the air for long periods of time.

Aetiology of acute respiratory infection in preschool children requiring hospitalisation in Europe-results from the PED-MERMAIDS multicentre case-control study.

BACKGROUND: Both pathogenic bacteria and viruses are frequently detected in the nasopharynx (NP) of children in the absence of acute respiratory infection (ARI) symptoms. The aim of this study was to estimate the aetiological fractions for ARI hospitalisation in children for respiratory syncytial virus (RSV) and influenza virus and to determine whether detection of specific respiratory pathogens on NP samples was associated with ARI hospitalisation. METHODS: 349 children up to 5 years of age hospitalised for ARI (following a symptom-based case definition) and 306 hospital controls were prospectively enrolled in 16 centres across seven European Union countries between 2016 and 2019. Admission day NP swabs were analysed by multiplex PCR for 25 targets. RESULTS: RSV was the leading single cause of ARI hospitalisations, with an overall population attributable fraction (PAF) of 33.4% and high seasonality as well as preponderance in younger children. Detection of RSV on NP swabs was strongly associated with ARI hospitalisation (OR adjusted for age and season: 20.6, 95% CI: 9.4 to 45.3). Detection of three other viral pathogens showed strong associations with ARI hospitalisation: influenza viruses had an adjusted OR of 6.1 (95% CI: 2.5 to 14.9), parainfluenza viruses (PIVs) an adjusted OR of 4.6 (95% CI: 1.8 to 11.3) and metapneumoviruses an adjusted OR of 4.5 (95% CI: 1.3 to 16.1). Influenza viruses had a PAF of 7.9%, PIVs of 6.5% and metapneumoviruses of 3.0%. In contrast, most other pathogens were found in similar proportions in cases and controls, including Streptococcus pneumoniae, which was weakly associated with case status, and endemic coronaviruses. CONCLUSION: RSV is the predominant cause of ARI hospitalisations in young children in Europe and its detection, as well as detection of influenza virus, PIV or metapneumovirus, on NP swabs can establish aetiology with high probability. PAFs for RSV and influenza virus are highly seasonal and age dependent.

Whole genome sequencing of human metapneumoviruses from clinical specimens using MinION nanopore technology.

Human metapneumovirus (HMPV), a member of the Pneumoviridae family, is a causative agent of respiratory illness in young children, the elderly, and immunocompromised individuals. Globally, viruses belonging to two main genetic lineages circulate, A and B, which are further divided into four genetic sublineages (A1, A2, B1, B2). Classical genotyping of HMPV is based on the sequence of the fusion (F) and attachment (G) glycoprotein genes, which are under direct antibody-mediated immune pressure. Whole genome sequencing provides more information than sequencing of subgenomic fragments and is therefore a powerful tool for studying virus evolution and disease epidemiology and for identifying transmission events and nosocomial outbreaks. Here, we report a robust method to obtain whole genome sequences for HMPV using MinION Nanopore technology. This assay is able to generate HMPV whole genome sequences from clinical specimens with good coverage of the highly variable G gene and is equally sensitive for strains of all four genetic HMPV sublineages. This method can be used for studying HMPV genetics, epidemiology, and evolutionary dynamics.