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Abstract West Nile virus (WNV) and Usutu virus (USUV) are closely related flaviviruses with differing capacities to cause neurological disease in humans. WNV is thought to use a transneural route of neuroinvasion along motor neurons and causes severe motor deficits. The potential for use of transneural routes of neuroinvasion by USUV has not been investigated experimentally, and evidence from the few clinical case reports of USUV-associated neuroinvasive disease is lacking. We hypothesised that, compared with WNV, USUV is less able to infect motor neurons, and therefore determined the susceptibility of human induced pluripotent stem cell (iPSC)-derived spinal cord motor neurons to infection. Both viruses could grow to high titres in iPSC-derived neural cultures. However, USUV could not productively infect motor neurons due to restriction by the antiviral response, which was not induced upon WNV infection. Inhibition of the antiviral response allowed for widespread infection and transportation of USUV along motor neurons within a compartmented culture system. These results show a stark difference in the ability of these two viruses to evade initiation of intrinsic antiviral immunity. Our data suggests that USUV cannot infect motor neurons in healthy individuals but in case of immunodeficiency may pose a risk for motor-related neurological disease and transneural invasion.

Abstract Sewage metagenomics has risen to prominence in urban population surveillance of pathogens and antimicrobial resistance (AMR). Unknown species with similarity to known genomes cause database bias in reference-based metagenomics. To improve surveillance, we seek to recover sewage genomes and develop a quantification and correlation workflow for these genomes and AMR over time. We use longitudinal sewage sampling in seven treatment plants from five major European cities to explore the utility of catch-all sequencing of these population-level samples. Using metagenomic assembly methods, we recover 2332 metagenome-assembled genomes (MAGs) from prokaryotic species, 1334 of which were previously undescribed. These genomes account for ~69% of sequenced DNA and provide insight into sewage microbial dynamics. Rotterdam (Netherlands) and Copenhagen (Denmark) show strong seasonal microbial community shifts, while Bologna, Rome, (Italy) and Budapest (Hungary) have occasional blooms of Pseudomonas-dominated communities, accounting for up to ~95% of sample DNA. Seasonal shifts and blooms present challenges for effective sewage surveillance. We find that bacteria of known shared origin, like human gut microbiota, form communities, suggesting the potential for source-attributing novel species and their ARGs through network community analysis. This could significantly improve AMR tracking in urban environments.

Abstract West Nile virus (WNV) leads to thousands of cases of severe neurological disease in humans each year. Usutu virus (USUV) is closely related to WNV, but rarely induces disease in humans. We hypothesised that USUV is less able to cross the blood-brain barrier (BBB) and, consequently, is less likely to infect the brain. Therefore, we developed an in vitro BBB model consisting of primary human brain microvascular endothelial cells, pericytes and astrocytes. Both USUV and WNV invaded across the in vitro BBB via a transcellular mechanism in the absence of barrier disruption. USUV replicated to lower titres than WNV but induced a comparable cytokine and chemokine response, with modulation of key factors associated with barrier function and immune-cell migration. In conclusion, USUV appears attenuated in its ability to replicate at this interface compared with WNV, but further work must be done to identify key determinants underlying the differing clinical presentations.

Abstract Systematic monitoring of SARS-CoV-2 co-infections between different lineages and assessing the risk of intra-host recombinant emergence are crucial for forecasting viral evolution. Here we present a comprehensive analysis of more than 2 million SARS-CoV-2 raw read datasets submitted to the European COVID-19 Data Portal to identify co-infections and intra-host recombination. Co-infection was observed in 0.35% of the investigated cases. Two independent procedures were implemented to detect intra-host recombination. We show that sensitivity is predominantly determined by the density of lineage-defining mutations along the genome, thus we used an expanded list of mutually exclusive defining mutations of specific variant combinations to increase statistical power. We call attention to multiple challenges rendering recombinant detection difficult and provide guidelines for the reduction of false positives arising from chimeric sequences produced during PCR amplification. Additionally, we identify three recombination hotspots of Delta – Omicron BA.1 intra-host recombinants.

In October 2020, a new lineage of a clade 2.3.4.4b HPAI virus of the H5 subtype emerged in Europe, resulting in the largest global outbreak of HPAI to date, with unprecedented mortality in wild birds and poultry. The virus appears to have become enzootic in birds, continuously yielding novel HPAI virus variants. The recently increased abundance of infected birds worldwide increases the probability of bird–mammal contact, particularly in wild carnivores. Here, we performed molecular and serological screening of over 500 dead wild carnivores and sequencing of RNA positive materials. We show virological evidence for HPAI H5 virus infection in 0.8%, 1.4%, and 9.9% of animals tested in 2020, 2021, and 2022 respectively, with the highest proportion of positives in foxes, polecats and stone martens. We obtained near full genomes of 7 viruses and detected PB2 amino acid substitutions known to play a role in mammalian adaptation in three sequences. Infections were also found in without neurological signs or mortality. Serological evidence for infection was detected in 20% of the study population. These findings suggests that a high proportion of wild carnivores is infected but undetected in current surveillance programmes. We recommend increased surveillance in susceptible mammals, irrespective of neurological signs or encephalitis.

The health of humans, domestic and wild animals, plants, and the environment are inter-dependent. Global anthropogenic change is a key driver of disease emergence and spread and leads to biodiversity loss and ecosystem function degradation, which are themselves drivers of disease emergence. Pathogen spill-over events and subsequent disease outbreaks, including pandemics, in humans, animals and plants may arise when factors driving disease emergence and spread converge. One Health is an integrated approach that aims to sustainably balance and optimize human, animal and ecosystem health. Conventional disease surveillance has been siloed by sectors, with separate systems addressing the health of humans, domestic animals, cultivated plants, wildlife and the environment. One Health surveillance should include integrated surveillance for known and unknown pathogens, but combined with this more traditional disease-based surveillance, it also must include surveillance of drivers of disease emergence to improve prevention and mitigation of spill-over events. Here, we outline such an approach, including the characteristics and components required to overcome barriers and to optimize an integrated One Health surveillance system.

Understanding the emergence, evolution, and transmission of antibiotic resistance genes (ARGs) is essential to combat antimicrobial resistance. Here, Munk et al. analyse ARGs in hundreds of sewage samples from 101 countries and describe regional patterns, diverse genetic environments of common ARGs, and ARG-specific transmission patterns.

We detected a highly divergent SARS-CoV-2 Alpha variant in an immunocompromised person several months after the latest detection of the Alpha variant in the Netherlands. The patient was infected for 42 weeks despite several treatment regimens and disappearance of most clinical symptoms. We identified several potential immune escape mutations in the spike protein.

Multi-strain pathogens, such as influenza, present challenges for interpretation of seroprevalence data as estimates may vary by strain. Here, the authors develop a method for estimating age-specific seroprevalence based on principal components analysis and apply it to influenza data from Vietnam.

Rapid detection of infection is essential for stopping the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The Roche SD Biosensor rapid antigen test for SARS-CoV-2 was evaluated in a nonhospitalized symptomatic population. We rapid-tested a sample onsite and compared results with those from reverse transcription PCR and virus culture. We analyzed date of onset and symptoms using data from a clinical questionnaire. Overall test sensitivity was 84.9% (95% CI 79.1–89.4) and specificity was 99.5% (95% CI 98.7–99.8). Sensitivity increased to 95.8% (95% CI 90.5–98.2) for persons who sought care within 7 days of symptom onset. Test band intensity and time to result correlated strongly with viral load; thus, strong positive results could be read before the recommended time. Approximately 98% of all viable specimens with cycle threshold

The Mosquito Alert dataset includes occurrence records of adult mosquitoes collected worldwide in 2014–2020 through Mosquito Alert, a citizen science system for investigating and managing disease-carrying mosquitoes. Records are linked to citizen science-submitted photographs and validated by entomologists to determine the presence of five targeted European mosquito vectors: Aedes albopictus, Ae. aegypti, Ae. japonicus, Ae. koreicus, and Culex pipiens. Most records are from Spain, reflecting Spanish national and regional funding, but since autumn 2020 substantial records from other European countries are included, thanks to volunteer entomologists coordinated by the AIM-COST Action, and to technological developments to increase scalability. Among other applications, the Mosquito Alert dataset will help develop citizen science-based early warning systems for mosquito-borne disease risk. It can also be reused for modelling vector exposure risk, or to train machine-learning detection and classification routines on the linked images, to assist with data validation and establishing automated alert systems.

We report a new norovirus GII.4 variant, GII.4 Hong Kong, with low-level circulation in 4 Eurasia countries since mid-2017. Amino acid substitutions in key residues on the virus capsid associated with the emergence of pandemic noroviruses suggest that GII.4 Hong Kong has the potential to become the next pandemic variant.

SARS-CoV-2 is causing a global pandemic in which the implementation of serology can support decision making in different scenarios. Here, the authors compare the outcome of eight commercially available assays to virus neutralization and discuss their use in diagnostics and exposure assessment of SARS-CoV-2.

Identifying the origin of the rabies virus (RABV) infection may have significant implications for control measures. Here, we identified the source of a RABV infection of two Nepalese migrants in Qatar by comparing their RABV genomes with RABV genomes isolated from the brains of a RABV infected camel and fox from Qatar.

Summary: Biobanking infrastructures, which are crucial for responding early to new viral outbreaks, share pathogen genetic resources in an affordable, safe, and impartial manner and can provide expertise to address access and benefit-sharing issues. The European Virus Archive has had a crucial role in the global response to the COVID-19 pandemic by distributing EU-subsidised (free of charge) viral resources to users worldwide, providing non-monetary benefit sharing, implementing access and benefit-sharing compliance, and raising access and benefit-sharing awareness among members and users. All currently available SARS-CoV-2 material in the European Virus Archive catalogue, including variants of concern, are not access and benefit-sharing cases per se, but multilateral benefit-sharing has nevertheless occurred. We propose and discuss how a multilateral system enabling access and benefit-sharing from pathogen genetic resources, based on the European Virus Archive operational model, could help bridge the discrepancies between the current bilateral legal framework for pathogen genetic resources and actual pandemic response practices.

A Zika virus (ZIKV) infection during pregnancy can result in severe birth defects such as microcephaly. To date, it is incompletely understood how ZIKV can cross the human placenta. Furthermore, results from studies in pregnant mice and non-human primates are conflicting regarding the role of cross-reactive dengue virus (DENV) antibodies on transplacental ZIKV transmission. Elucidating how ZIKV can cross the placenta and which risk factors contribute to this is important for risk assessment and for potential intervention strategies for transplacental ZIKV transmission. In this study we use an ex vivo human placental perfusion model to study transplacental ZIKV transmission and the effect that cross-reactive DENV antibodies have on this transmission. By using this model, we demonstrate that DENV antibodies significantly increase ZIKV uptake in perfused human placentas and that this increased uptake is neonatal Fc-receptor-dependent. Furthermore, we show that cross-reactive DENV antibodies enhance ZIKV infection in term human placental explants and in primary fetal macrophages but not in primary trophoblasts. Our data supports the hypothesis that presence of cross-reactive DENV antibodies could be an important risk factor for transplacental ZIKV transmission. Furthermore, we demonstrate that the ex vivo placental perfusion model is a relevant and animal friendly model to study transplacental pathogen transmission.

An outbreak of SARS-CoV-2 Alpha variant (Pango lineage B.1.1.7) was detected at a primary school (School X) in Lansingerland, the Netherlands, in December 2020. The outbreak was studied retrospectively, and population-based screening was used to assess the extent of virus circulation and decelerate transmission. Cases were SARS-CoV-2 laboratory confirmed and were residents of Lansingerland (November 16th 2020 until February 22th 2021), or had an epidemiological link with School X or neighbouring schools. The SARS-CoV-2 variant was determined using variant PCR or whole genome sequencing. A questionnaire primarily assessed clinical symptoms. A total of 77 Alpha variant cases were found with an epidemiological link to School X, 16 Alpha variant cases linked to the neighbouring schools, and 146 Alpha variant cases among residents of Lansingerland without a link to the schools. The mean number of self-reported symptoms was not significantly different among Alpha variant infected individuals compared to non-Alpha infected individuals. The secondary attack rate (SAR) among Alpha variant exposed individuals in households was 52% higher compared to non-Alpha variant exposed individuals (p = 0.010), with the mean household age, and mean number of children and adults per household as confounders. Sequence analysis of 60 Alpha variant sequences obtained from cases confirmed virus transmission between School X and neighbouring schools, and showed that multiple introductions of the Alpha variant had already taken place in Lansingerland at the time of the study. The alpha variant caused a large outbreak at both locations of School X, and subsequently spread to neighbouring schools, and households. Population-based screening (together with other public health measures) nearly stopped transmission of the outbreak strain, but did not prevent variant replacement in the Lansingerland municipality.

During the 2015–2016 outbreak of Zika virus (ZIKV) in the Americas, a previously unknown severe complication of ZIKV infection during pregnancy resulting in birth defects was reported. Since the ZIKV outbreak occurred in regions that were highly endemic for the related dengue virus (DENV), it was speculated that antibody-dependent enhancement (ADE) of a ZIKV infection, caused by the presence of cross-reactive DENV antibodies, could contribute to ZIKV disease severity. Emerging evidence indicates that, while in vitro models can show ADE of ZIKV infection, ADE does not seem to contribute to congenital ZIKV disease severity in humans. However, the role of ADE of ZIKV infection during pregnancy and in vertical ZIKV transmission is not well studied. In this study, we hypothesized that pregnancy may affect the ability of myeloid cells to become infected with ZIKV, potentially through ADE. We first systematically assessed which cell lines and primary cells can be used to study ZIKV ADE in vitro, and we compared the difference in outcomes of (ADE) infection experiments between these cells. Subsequently, we tested the hypothesis that pregnancy may affect the ability of myeloid cells to become infected through ADE, by performing ZIKV ADE assays with primary cells isolated from blood of pregnant women from different trimesters and from age-matched non-pregnant women. We found that ADE of ZIKV infection can be induced in myeloid cell lines U937, THP-1, and K562 as well as in monocyte-derived macrophages from healthy donors. There was no difference in permissiveness for ZIKV infection or ADE potential of ZIKV infection in primary cells of pregnant women compared to non-pregnant women. In conclusion, no increased permissiveness for ZIKV infection and ADE of ZIKV infection was found using in vitro models of primary myeloid cells from pregnant women compared to age-matched non-pregnant women.

We analysed all proven cases of SARS-CoV-2 on sea-going vessels entering or present in the Port of Rotterdam to illustrate the burden of the corona pandemic on commercial shipping. We calculated the notification rate (NR) per arrival and attack rate per vessel and correlated outbreak sizes by vessel type over time during the pandemic using routinely gathered data from the port and public health (PH) authorities.

Late in 2020, two genetically-distinct clusters of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) with mutations of biological concern were reported, one in the United Kingdom and one in South Africa. Using a combination of data from routine surveillance, genomic sequencing and international travel we track the international dispersal of lineages B.1.1.7 and B.1.351 (variant 501Y-V2). We account for potential biases in genomic surveillance efforts by including passenger volumes from location of where the lineage was first reported, London and South Africa respectively. Using the software tool grinch (global report investigating novel coronavirus haplotypes), we track the international spread of lineages of concern with automated daily reports, Further, we have built a custom tracking website (cov-lineages.org/global_report.html) which hosts this daily report and will continue to include novel SARS-CoV-2 lineages of concern as they are detected.

Background: Previously, we conducted a systematic review and analyzed the respiratory kinetics of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (Chen et al., 2021). How age, sex, and coronavirus disease 2019 (COVID-19) severity interplay to influence the shedding dynamics of SARS-CoV-2, however, remains poorly understood. Methods: We updated our systematic dataset, collected individual case characteristics, and conducted stratified analyses of SARS-CoV-2 shedding dynamics in the upper (URT) and lower respiratory tract (LRT) across COVID-19 severity, sex, and age groups (aged 0–17 years, 18–59 years, and 60 years or older). Results: The systematic dataset included 1266 adults and 136 children with COVID-19. Our analyses indicated that high, persistent LRT shedding of SARS-CoV-2 characterized severe COVID-19 in adults. Severe cases tended to show slightly higher URT shedding post-symptom onset, but similar rates of viral clearance, when compared to nonsevere infections. After stratifying for disease severity, sex and age (including child vs. adult) were not predictive of respiratory shedding. The estimated accuracy for using LRT shedding as a prognostic indicator for COVID-19 severity was up to 81%, whereas it was up to 65% for URT shedding. Conclusions: Virological factors, especially in the LRT, facilitate the pathogenesis of severe COVID-19. Disease severity, rather than sex or age, predicts SARS-CoV-2 kinetics. LRT viral load may prognosticate COVID-19 severity in patients before the timing of deterioration and should do so more accurately than URT viral load. Funding: Natural Sciences and Engineering Research Council of Canada (NSERC) Discovery Grant, NSERC Senior Industrial Research Chair, and the Toronto COVID-19 Action Fund.

Abstract Background The European Commission (EC) Horizon 2020 (H2020)-funded ZIKAlliance Consortium designed a multicentre study including pregnant women (PW), children (CH) and natural history (NH) cohorts. Clinical sites were selected over a wide geographic range within Latin America and the Caribbean, taking into account the dynamic course of the ZIKV epidemic. Methods Recruitment to the PW cohort will take place in antenatal care clinics. PW will be enrolled regardless of symptoms and followed over the course of pregnancy, approximately every 4 weeks. PW will be revisited at delivery (or after miscarriage/abortion) to assess birth outcomes, including microcephaly and other congenital abnormalities according to the evolving definition of congenital Zika syndrome (CZS). After birth, children will be followed for 2 years in the CH cohort. Follow-up visits are scheduled at ages 1–3, 4–6, 12, and 24 months to assess neurocognitive and developmental milestones. In addition, a NH cohort for the characterization of symptomatic rash/fever illness was designed, including follow-up to capture persisting health problems. Blood, urine, and other biological materials will be collected, and tested for ZIKV and other relevant arboviral diseases (dengue, chikungunya, yellow fever) using RT-PCR or serological methods. A virtual, decentralized biobank will be created. Reciprocal clinical monitoring has been established between partner sites. Substudies of ZIKV seroprevalence, transmission clustering, disabilities and health economics, viral kinetics, the potential role of antibody enhancement, and co-infections will be linked to the cohort studies. Discussion Results of these large cohort studies will provide better risk estimates for birth defects and other developmental abnormalities associated with ZIKV infection including possible co-factors for the variability of risk estimates between other countries and regions. Additional outcomes include incidence and transmission estimates of ZIKV during and after pregnancy, characterization of short and long-term clinical course following infection and viral kinetics of ZIKV. Study registrations clinicaltrials.gov NCT03188731 (PW cohort), June 15, 2017; clinicaltrials.gov NCT03393286 (CH cohort), January 8, 2018; clinicaltrials.gov NCT03204409 (NH cohort), July 2, 2017.

Severe COVID-19 can be considered a systemic disease as many extrarespiratory complications occur. However, the systemic pathogenesis is poorly understood.

Late in 2020, two genetically-distinct clusters of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) with mutations of biological concern were reported, one in the United Kingdom and one in South Africa. Using a combination of data from routine surveillance, genomic sequencing and international travel we track the international dispersal of lineages B.1.1.7 and B.1.351 (variant 501Y-V2). We account for potential biases in genomic surveillance efforts by including passenger volumes from location of where the lineage was first reported, London and South Africa respectively. Using the software tool grinch (global report investigating novel coronavirus haplotypes), we track the international spread of lineages of concern with automated daily reports, Further, we have built a custom tracking website (cov-lineages.org/global_report.html) which hosts this daily report and will continue to include novel SARS-CoV-2 lineages of concern as they are detected.

Background: Which virological factors mediate overdispersion in the transmissibility of emerging viruses remains a long-standing question in infectious disease epidemiology. Methods: Here, we use systematic review to develop a comprehensive dataset of respiratory viral loads (rVLs) of SARS-CoV-2, SARS-CoV-1 and influenza A(H1N1)pdm09. We then comparatively meta-analyze the data and model individual infectiousness by shedding viable virus via respiratory droplets and aerosols. Results: The analyses indicate heterogeneity in rVL as an intrinsic virological factor facilitating greater overdispersion for SARS-CoV-2 in the COVID-19 pandemic than A(H1N1)pdm09 in the 2009 influenza pandemic. For COVID-19, case heterogeneity remains broad throughout the infectious period, including for pediatric and asymptomatic infections. Hence, many COVID-19 cases inherently present minimal transmission risk, whereas highly infectious individuals shed tens to thousands of SARS-CoV-2 virions/min via droplets and aerosols while breathing, talking and singing. Coughing increases the contagiousness, especially in close contact, of symptomatic cases relative to asymptomatic ones. Infectiousness tends to be elevated between 1 and 5 days post-symptom onset. Conclusions: Intrinsic case variation in rVL facilitates overdispersion in the transmissibility of emerging respiratory viruses. Our findings present considerations for disease control in the COVID-19 pandemic as well as future outbreaks of novel viruses. Funding: Natural Sciences and Engineering Research Council of Canada (NSERC) Discovery Grant program, NSERC Senior Industrial Research Chair program and the Toronto COVID-19 Action Fund.

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.

BackgroundPerformance of the SD Biosensor saliva antigen rapid test was evaluated at a large designated testing site in non-hospitalized patients, with or without symptoms.MethodAll eligible people over 18 years of age presenting for a booked appointment at the designated SARS-CoV-2 testing site were approached for inclusion and enrolled following verbal informed consent. One nasopharyngeal swab was taken to carry out the default antigen rapid test from which the results were reported back to the patient and one saliva sample was self-taken according to verbal instruction on site. This was used for the saliva antigen rapid test, the RT-PCR and for virus culture. Sensitivity of the saliva antigen rapid test was analyzed in two ways: i, compared to saliva RT-PCR; and ii, compared to virus culture of the saliva samples. Study participants were also asked to fill in a short questionnaire stating age, sex, date of symptom onset. Recommended time of ≥30mins since last meal, drink or cigarette if applicable was also recorded. The study was carried out in February-March 2021 for 4 weeks.ResultsWe could include 789 people with complete records and results. Compared to saliva RT-PCR, overall sensitivity and specificity of the saliva antigen rapid test was 66.1% and 99.6% which increased to 88.6% with Ct ≤30 cutoff. Analysis by days post onset did not result in higher sensitivities because the large majority of people were in the very early phase of disease ie ConclusionOverall, the potential benefits of saliva antigen rapid test, could outweigh the lower sensitivity compared to nasopharyngeal antigen rapid test in a comprehensive testing strategy, especially for home/self-testing and in vulnerable populations like elderly, disabled or children where in intrusive testing is either not possible or causes unnecessary stress.

Introduction Zika virus (ZIKV) infection in pregnancy has been associated with microcephaly and severe neurological damage to the fetus. Our aim is to document the risks of adverse pregnancy and birth outcomes and the prevalence of laboratory markers of congenital infection in deliveries to women experiencing ZIKV infection during pregnancy, using data from European Commission-funded prospective cohort studies in 20 centres in 11 countries across Latin America and the Caribbean.Methods and analysis We will carry out a centre-by-centre analysis of the risks of adverse pregnancy and birth outcomes, comparing women with confirmed and suspected ZIKV infection in pregnancy to those with no evidence of infection in pregnancy. We will document the proportion of deliveries in which laboratory markers of congenital infection were present. Finally, we will investigate the associations of trimester of maternal infection in pregnancy, presence or absence of maternal symptoms of acute ZIKV infection and previous flavivirus infections with adverse outcomes and with markers of congenital infection. Centre-specific estimates will be pooled using a two-stage approach.Ethics and dissemination Ethical approval was obtained at each centre. Findings will be presented at international conferences and published in peer-reviewed open access journals and discussed with local public health officials and representatives of the national Ministries of Health, Pan American Health Organization and WHO involved with ZIKV prevention and control activities.

The response of pathophysiological research to emerging epidemics often occurs after the epidemic and, as a consequence, has little to no impact on improving patient outcomes or on developing high-quality evidence to inform clinical management strategies during the epidemic. Rapid and informed guidance of epidemic (research) responses to severe infectious disease outbreaks requires quick compilation and integration of existing pathophysiological knowledge. As a case study we chose the Zika virus (ZIKV) outbreak that started in 2015 to develop a proof-of-concept knowledge repository. To extract data from available sources and build a computationally tractable and comprehensive molecular interaction map we applied generic knowledge management software for literature mining, expert knowledge curation, data integration, reporting and visualization. A multi-disciplinary team of experts, including clinicians, virologists, bioinformaticians and knowledge management specialists, followed a pre-defined workflow for rapid integration and evaluation of available evidence. While conventional approaches usually require months to comb through the existing literature, the initial ZIKV KnowledgeBase (ZIKA KB) was completed within a few weeks. Recently we updated the ZIKA KB with additional curated data from the large amount of literature published since 2016 and made it publicly available through a web interface together with a step-by-step guide to ensure reproducibility of the described use case. In addition, a detailed online user manual is provided to enable the ZIKV research community to generate hypotheses, share knowledge, identify knowledge gaps, and interactively explore and interpret data. A workflow for rapid response during outbreaks was generated, validated and refined and is also made available. The process described here can be used for timely structuring of pathophysiological knowledge for future threats. The resulting structured biological knowledge is a helpful tool for computational data analysis and generation of predictive models and opens new avenues for infectious disease research. ZIKV Knowledgebase is available at www.zikaknowledgebase.eu.

We tested samples collected from camels, camel workers, and other animals in Sudan and Qatar in 2015 and 2017 for evidence of Middle East respiratory syndrome coronavirus (MERS-CoV) infection. MERS-CoV antibodies were abundant in Sudan camels, but we found no evidence of MERS-CoV infection in camel workers, other livestock, or bats.

We evaluated the benefit of whole blood versus plasma to detect acute Zika virus infections. Comparison of Zika virus quantitative reverse transcription PCR results in single timepoint whole blood–plasma pairs from 227 patients with suspected Zika virus infection resulted in confirmation of 8 additional patients with Zika virus infection.

Abstract Although avian paramyxovirus type 1 is known to cause mild transient conjunctivitis in human beings, there are two recent reports of fatal respiratory disease in immunocompromised human patients infected with the pigeon lineage of the virus (PPMV-1). In order to evaluate the potential of PPMV-1 to cause respiratory tract disease, we inoculated a PPMV-1 isolate (hPPMV-1/Netherlands/579/2003) from an immunocompromised human patient into three healthy cynomolgus macaques (Macaca fascicularis) and examined them by clinical, virological, and pathological assays. In all three macaques, PPMV-1 replication was restricted to the respiratory tract and caused pulmonary consolidation affecting up to 30% of the lung surface. Both alveolar and bronchiolar epithelial cells expressed viral antigen, which co-localized with areas of diffuse alveolar damage. The results of this study demonstrate that PPMV-1 is a primary respiratory pathogen in cynomolgus macaques, and support the conclusion that PPMV-1 may cause fatal respiratory disease in immunocompromised human patients.

Abstract Seroepidemiological studies aim to understand population-level exposure and immunity to infectious diseases. Their results are normally presented as binary outcomes describing the presence or absence of pathogen-specific antibody, despite the fact that many assays measure continuous quantities. A population’s natural distribution of antibody titers to an endemic infectious disease may include information on multiple serological states – naiveté, recent infection, non-recent infection, childhood infection – depending on the disease in question and the acquisition and waning patterns of immunity. In this study, we investigate 20,152 general-population serum samples from southern Vietnam collected between 2009 and 2013 from which we report antibody titers to the influenza virus HA1 protein using a continuous titer measurement from a protein microarray assay. We describe the distributions of antibody titers to subtypes 2009 H1N1 and H3N2. Using a model selection approach to fit mixture distributions, we show that 2009 H1N1 antibody titers fall into four titer subgroups and that H3N2 titers fall into three subgroups. For H1N1, our interpretation is that the two highest-titer subgroups correspond to recent and historical infection, which is consistent with 2009 pandemic attack rates. Similar interpretations are available for H3N2, but right-censoring of titers makes these interpretations difficult to validate.

We screened squirrels in Germany and the Netherlands for the novel zoonotic variegated squirrel bornavirus 1 (VSBV-1). The detection of VSBV-1 in 11 squirrels indicates a considerable risk for transmission to humans handling those animals. Therefore, squirrels in contact with humans should routinely be tested for VSBV-1.

IntroductionZika virus (ZIKV) infection during pregnancy is a known cause of microcephaly and other congenital and developmental anomalies. In the absence of a ZIKV vaccine or prophylactics, principal investigators (PIs) and international leaders in ZIKV research have formed the ZIKV Individual Participant Data (IPD) Consortium to identify, collect and synthesise IPD from longitudinal studies of pregnant women that measure ZIKV infection during pregnancy and fetal, infant or child outcomes.Methods and analysisWe will identify eligible studies through the ZIKV IPD Consortium membership and a systematic review and invite study PIs to participate in the IPD meta-analysis (IPD-MA). We will use the combined dataset to estimate the relative and absolute risk of congenital Zika syndrome (CZS), including microcephaly and late symptomatic congenital infections; identify and explore sources of heterogeneity in those estimates and develop and validate a risk prediction model to identify the pregnancies at the highest risk of CZS or adverse developmental outcomes. The variable accuracy of diagnostic assays and differences in exposure and outcome definitions means that included studies will have a higher level of systematic variability, a component of measurement error, than an IPD-MA of studies of an established pathogen. We will use expert testimony, existing internal and external diagnostic accuracy validation studies and laboratory external quality assessments to inform the distribution of measurement error in our models. We will apply both Bayesian and frequentist methods to directly account for these and other sources of uncertainty.Ethics and disseminationThe IPD-MA was deemed exempt from ethical review. We will convene a group of patient advocates to evaluate the ethical implications and utility of the risk stratification tool. Findings from these analyses will be shared via national and international conferences and through publication in open access, peer-reviewed journals.Trial registration numberPROSPERO International prospective register of systematic reviews (CRD42017068915).

The emergence of the Middle East Respiratory Syndrome Corona Virus (MERS-CoV) in the Middle East in 2012 was associated with an overwhelming uncertainty about its epidemiological and clinical characteristics. Once dromedary camels (Camelus dromedarius) was found to be the natural reservoir of the virus, the public health systems across the Arabian Peninsula encountered an unprecedented pressure to control its transmission. This view point describes how the One Health approach was used in Qatar to manage the MERS-CoV outbreak during the period 2012–2017.One Health focuses on the association between the human, animals and environment sectors for total health and wellbeing of these three sectors. To manage the MERS outbreak in Qatar through a One Health approach, the Qatar National Outbreak Control Taskforce (OCT) was reactivated in November 2012. The animal health sector was invited to join the OCT. Later on, technical expertise was requested from the WHO, FAO, CDC, EMC, and PHE. Subsequently, a comprehensive One Health roadmap was delivered through leadership and coordination; surveillance and investigation; epidemiological studies and increase of local diagnostic capacity.The joint OCT, once trained had easy access to allocated resources and high risk areas to provide more evidence on the potential source of the virus and to investigate all reported cases within 24–48 h. Lack of sufficient technical guidance on veterinary surveillance and poor risk perception among the vulnerable population constituted major obstacles to maintain systematic One Health performance. Keywords: One Health, MERS-CoV, Qatar

Since March 2013, three waves of human infection with avian influenza A(H7N9) virus have been detected in China. To investigate virus transmission within and across epidemic waves, we used surveillance data and whole-genome analysis of viruses sampled in Guangdong during 2013–2015. We observed a geographic shift of human A(H7N9) infections from the second to the third waves. Live poultry market interventions were undertaken in epicenter cities; however, spatial phylogenetic analysis indicated that the third-wave outbreaks in central Guangdong most likely resulted from local virus persistence rather than introduction from elsewhere. Although the number of clinical cases in humans declined by 35% from the second to the third waves, the genetic diversity of third-wave viruses in Guangdong increased. Our results highlight the epidemic risk to a region reporting comparatively few A(H7N9) cases. Moreover, our results suggest that live-poultry market interventions cannot completely halt A(H7N9) virus persistence and dissemination.

Introduction: The increase in emerging human infectious diseases that have a zoonotic origin and the increasing resistance of microorganisms to antimicrobial drugs have shown the need for collaborations between the human, animal and environmental health sectors. The One Health concept increasingly receives recognition from policy makers and researchers all over the world. This overview compiled research and education activities in the area of One Health in Western Europe (Austria, Belgium, France, Germany, Italy, Iceland, Ireland, Liechtenstein, Luxembourg, Monaco, the Netherlands, Portugal, Scandinavia, Spain, Switzerland, and the United Kingdom (UK), with a focus on infectious diseases. It can serve as a starting point for future initiatives and collaborations. Material and methods: A literature search for ‘One Health’ was performed using National Center for Biotechnology Information and Google. Moreover, information from global and European policy documents was collected and a questionnaire was designed to gather current One Health research and training activities in Western Europe. Results: This overview shows that there is considerable recognition for One Health in Europe, although most educational initiatives are recent. In Europe, the One Health approach is currently mainly advocated in relation to antimicrobial resistance (AMR). Many countries have incorporated the One Health approach in their policy to fight AMR, and funding possibilities for AMR research increased significantly. The number of national and international multidisciplinary research networks in the area of zoonotic diseases and One Health is increasing. Discussion: Although One Health has gained recognition in Europe, often a One Health approach to research and education in the area of zoonotic diseases and AMR is not implemented. In many countries, collaboration between sectors is still lacking, and One Health activities are predominantly initiated by the veterinary sector. To facilitate the multidisciplinary approach that is needed to fight zoonotic diseases and AMR, exploring current barriers for collaboration is needed. Targeted funding can help address these boundaries and facilitate multidisciplinary research and training to fight both zoonotic diseases and AMR in Europe.

We characterized Middle East respiratory syndrome coronaviruses from a hospital outbreak in Jordan in 2015. The viruses from Jordan were highly similar to isolates from Riyadh, Saudi Arabia, except for deletions in open reading frames 4a and 3. Transmissibility and pathogenicity of this strain remains to be determined.

We report the rare event of two imported cases in the Netherlands presenting with renal syndrome caused by Dobrava (DOBV)/Saaremaa (SAAV) hantaviruses. DOBV/SAAV hantaviruses are not circulating in the Netherlands and their clinical manifestation is typically more severe than that of the endemic Puumala virus (PUUV). This report aims to increase awareness among healthcare professionals and diagnostic laboratories to consider different hantaviruses as a cause of renal failure.

Worldwide, noroviruses are a leading cause of gastroenteritis. They can be transmitted from person to person directly or indirectly through contaminated food, water, or environments. To estimate the proportion of foodborne infections caused by noroviruses on a global scale, we used norovirus transmission and genotyping information from multiple international outbreak surveillance systems (Noronet, CaliciNet, EpiSurv) and from a systematic review of peer-reviewed literature. The proportion of outbreaks caused by food was determined by genotype and/or genogroup. Analysis resulted in the following final global profiles: foodborne transmission is attributed to 10% (range 9%%–11%) of all genotype GII.4 outbreaks, 27% (25%–30%) of outbreaks caused by all other single genotypes, and 37% (24%%–52%) of outbreaks caused by mixtures of GII.4 and other noroviruses. When these profiles are applied to global outbreak surveillance data, results indicate that ≈14% of all norovirus outbreaks are attributed to food.

The largest outbreak of Ebola virus disease ever started in West Africa in December 2013; it created a pressing need to expand the workforce dealing with it. The aim of this study was to gain insight into the experiences of volunteers from the European Union who worked in deployable laboratories in West Africa during the outbreak. This study is part of the EMERGE project. We assessed the experiences of 251 volunteers with a 19-item online questionnaire. The questions asked about positive aspects of volunteering such as learning new skills, establishing a new path in life, and changing life values. Other questionnaire subjects were the compliance to follow-up measures, the extent to which volunteers felt these measures restricted their daily activities, the fear of stigmatization, and worries about becoming infected or infecting their families. The volunteers reported positive effects that reached far beyond their daily work, such as changes in life priorities and a greater appreciation of the value of their own lives. Although the volunteers did not feel that temperature monitoring restricted their daily activities, full compliance to temperature monitoring and reporting it to the authorities was low. The volunteers did not fear Ebola infection for themselves or their families and were not afraid of stigmatization. With respect to the burden on the families, 50% reported that their family members were worried that the volunteer would be infected with Ebola virus. Altogether, the positive experiences of the volunteers in this study far outweigh the negative implications and constitute an important argument for inspiring people who intend to join such missions and for motivating the hesitant ones.

Influenza A(H7N9) virus emerged in eastern China in February 2013 and continues to circulate in this region, but its ecology is poorly understood. In April 2013, the Guangdong Provincial Center for Disease Control and Prevention (CDC) implemented environmental and human syndromic surveillance for the virus. Environmental samples from poultry markets in 21 city CDCs (n = 8,942) and respiratory samples from persons with influenza-like illness or pneumonia (n = 32,342) were tested; viruses isolated from 6 environmental samples and 16 patients were sequenced. Sequence analysis showed co-circulation of 4 influenza A(H7N9) virus strains that evolved by reassortment with avian influenza A(H9N2) viruses circulating in this region. In addition, an increase in human cases starting in late 2013 coincided with an increase in influenza A H7 virus isolates detected by environmental surveillance. Co-circulation of multiple avian influenza viruses that can infect humans highlights the need for increased surveillance of poultry and potential environmental sources.

Which case-based intervention measures should be applied during an epidemic outbreak depends on how timely they can be applied and how effective they are. During the course of each individual's infection, the earlier control measures are applied on him/her the more effectively further disease spread can be prevented. However, quick implementation can lead to loss of efficacy or coverage, e.g., when individuals are targeted based on rapid but poorly sensitive diagnostic tests in place of slower but accurate PCR tests. To analyse this trade off between speed and coverage we used stochastic models considering how the individual reproduction density is modified by interventions. We took as example the case-based intervention strategy employed in the Netherlands during the beginning of the H1N1 pandemic. Suspected cases were isolated and samples were collected for PCR diagnosis. In case of positive diagnosis, antiviral drugs were provided to contacts as post-exposure prophylaxis. At the time there were also rapid influenza diagnostic tests (RIDTs) available which provided results within an hour after sample collection compared to a median of 2.7 days for PCR tests, but they were less sensitive. We studied how interventions based on RIDTs with various sensitivities affect the outbreak size and how these compare to PCR diagnosis based interventions. Using an intervention based on a bedside RIDT with 60% detection ratio or a laboratory RIDT with 70% detection ratio is as effective as the most effective PCR-diagnosis based intervention. Relative performances of interventions are not dependent on the basic reproduction number R0 but only on distributions of individual reproduction density and of delay periods. The individual reproduction density combines R0 and infection time distribution, both crucial in determining the impact of case-based interventions during epidemic outbreaks.