SARS-CoV-2 variants reveal features critical for replication in primary human cells.

Since entering the human population, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2; the causative agent of Coronavirus Disease 2019 [COVID-19]) has spread worldwide, causing >100 million infections and >2 million deaths. While large-scale sequencing efforts have identified numerous genetic variants in SARS-CoV-2 during its circulation, it remains largely unclear whether many of these changes impact adaptation, replication, or transmission of the virus. Here, we characterized 14 different low-passage replication-competent human SARS-CoV-2 isolates representing all major European clades observed during the first pandemic wave in early 2020. By integrating viral sequencing data from patient material, virus stocks, and passaging experiments, together with kinetic virus replication data from nonhuman Vero-CCL81 cells and primary differentiated human bronchial epithelial cells (BEpCs), we observed several SARS-CoV-2 features that associate with distinct phenotypes. Notably, naturally occurring variants in Orf3a (Q57H) and nsp2 (T85I) were associated with poor replication in Vero-CCL81 cells but not in BEpCs, while SARS-CoV-2 isolates expressing the Spike D614G variant generally exhibited enhanced replication abilities in BEpCs. Strikingly, low-passage Vero-derived stock preparation of 3 SARS-CoV-2 isolates selected for substitutions at positions 5/6 of E and were highly attenuated in BEpCs, revealing a key cell-specific function to this region. Rare isolate-specific deletions were also observed in the Spike furin cleavage site during Vero-CCL81 passage, but these were rapidly selected against in BEpCs, underscoring the importance of this site for SARS-CoV-2 replication in primary human cells. Overall, our study uncovers sequence features in SARS-CoV-2 variants that determine cell-specific replication and highlights the need to monitor SARS-CoV-2 stocks carefully when phenotyping newly emerging variants or potential variants of concern.

Nosocomial COVID-19 Incidence and Secondary Attack Rates among Patients of Tertiary Care Center, Zurich, Switzerland.

Of 1,118 patients with COVID-19 at a university hospital in Switzerland during October 2020-June 2021, we found 83 (7.4%) had probable or definite healthcare-associated COVID-19. After in-hospital exposure, we estimated secondary attack rate at 23.3%. Transmission was associated with longer contact times and with lower cycle threshold values among index patients.

External Quality Assessment of SARS-CoV-2 Sequencing: an ESGMD-SSM Pilot Trial across 15 European Laboratories.

This first pilot trial on external quality assessment (EQA) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) whole-genome sequencing, initiated by the European Society of Clinical Microbiology and Infectious Diseases (ESCMID) Study Group for Genomic and Molecular Diagnostics (ESGMD) and the Swiss Society for Microbiology (SSM), aims to build a framework between laboratories in order to improve pathogen surveillance sequencing. Ten samples with various viral loads were sent out to 15 clinical laboratories that had free choice of sequencing methods and bioinformatic analyses. The key aspects on which the individual centers were compared were the identification of (i) single nucleotide polymorphisms (SNPs) and indels, (ii) Pango lineages, and (iii) clusters between samples. The participating laboratories used a wide array of methods and analysis pipelines. Most were able to generate whole genomes for all samples. Genomes were sequenced to various depths (up to a 100-fold difference across centers). There was a very good consensus regarding the majority of reporting criteria, but there were a few discrepancies in lineage and cluster assignments. Additionally, there were inconsistencies in variant calling. The main reasons for discrepancies were missing data, bioinformatic choices, and interpretation of data. The pilot EQA was overall a success. It was able to show the high quality of participating laboratories and provide valuable feedback in cases where problems occurred, thereby improving the sequencing setup of laboratories. A larger follow-up EQA should, however, improve on defining the variables and format of the report. Additionally, contamination and/or minority variants should be a further aspect of assessment.

Response of European grayling, Thymallus thymallus, to multiple stressors in hydropeaking rivers.

Rivers of the large Alpine valleys constitute iconic ecosystems that are highly threatened by multiple anthropogenic stressors. This stressor mix, however, makes it difficult to develop and refine conservation and restoration strategies. It is, therefore, urgent to acquire more detailed knowledge on the consequences and interactions of prevalent stressors on fish populations, in particular, on indicator species such as the European grayling Thymallus thymallus. Here, we conducted a multi-river, multi-stressor investigation to analyze the population status of grayling. Using explorative decision-tree approaches, we disentangled the main and interaction effects of four prevalent stressor groups: flow modification (i.e., hydropeaking), channelization, fragmentation, and water quality alteration. Moreover, using a modified variant of the bootstrapping method, pooled bootstrapping, we determined the optimal number of characteristics that adequately describe fish population status. In our dataset, hydropeaking had the strongest single effect on grayling populations. Grayling biomass at hydrological control sites was around eight times higher than at sites affected by hydropeaking. The primary parameters for predicting population status were downramping rate and peak amplitude, with critical ranges of 0.2-0.4 cm min-1 and 10-25 cm. In hydropeaking rivers, river morphology and connectivity were the preceding subordinated parameters. Repeating the procedure with pooled bootstrapping datasets strengthened the hypothesis that the identified parameters are most relevant in predicting grayling population status. Hence, hydropeaking mitigation based on ecological thresholds is key to protect and restore already threatened grayling populations. In hydropeaking rivers, high river network connectivity and heterogenous habitat features can dampen the adverse effects of pulsed-flow releases by offering shelter and habitats for all life cycle stages of fish. The presented approach of explorative tree analysis followed by post-hoc tests of identified effects, as well as the pooled bootstrapping method, offers a simple framework for researchers and managers to analyze multi-factorial datasets and draw solid management conclusions.

Metagenomic Virome Sequencing in Living Donor and Recipient Kidney Transplant Pairs Revealed JC Polyomavirus Transmission.

BACKGROUND: Before kidney transplantation, donors and recipients are routinely screened for viral pathogens using specific tests. Little is known about unrecognized viruses of the urinary tract that potentially result in transmission. Using an open metagenomic approach, we aimed to comprehensively assess virus transmission in living-donor kidney transplantation. METHODS: Living kidney donors and their corresponding recipients were enrolled at the time of transplantation. Follow-up study visits for recipients were scheduled 4-6 weeks and 1 year thereafter. At each visit, plasma and urine samples were collected and transplant recipients were evaluated for signs of infection or other transplant-related complications. For metagenomic analysis, samples were enriched for viruses, amplified by anchored random polymerase chain reaction (PCR), and sequenced using high-throughput metagenomic sequencing. Viruses detected by sequencing were confirmed using real-time PCR. RESULTS: We analyzed a total of 30 living kidney donor and recipient pairs, with a follow-up of at least 1 year. In addition to viruses commonly detected during routine post-transplant virus monitoring, metagenomic sequencing detected JC polyomavirus (JCPyV) in the urine of 7 donors and their corresponding recipients. Phylogenetic analysis confirmed infection with the donor strain in 6 cases, suggesting transmission from the transplant donor to the recipient, despite recipient seropositivity for JCPyV at the time of transplantation. CONCLUSIONS: Metagenomic sequencing identified frequent transmission of JCPyV from kidney transplant donors to recipients. Considering the high incidence rate, future studies within larger cohorts are needed to define the relevance of JCPyV infection and the donor's virome for transplant outcomes.

Meningitis and epididymitis caused by Toscana virus infection imported to Switzerland diagnosed by metagenomic sequencing: a case report.

BACKGROUND: We report a rare case of Toscana virus infection imported into Switzerland in a 23-year old man who travelled to Imperia (Italy) 10 days before onset of symptoms. Symptoms included both meningitis and as well epididymitis. This is only the fourth case of Toscana virus reported in Switzerland. CASE PRESENTATION: The patient presented with lymphocytic meningitis and scrotal pain due to epididymitis. Meningitis was initially treated with ceftriaxone. Herpes simplex, tick-borne encephalitis, enterovirus, measles, mumps, rubella and Treponema pallidum were excluded with specific polymerase chain reaction (PCR) or serology. In support of routine diagnostic PCR and serology assays, unbiased viral metagenomic sequencing was performed of cerebrospinal fluid and serum. Toscana virus infection was identified in cerebrospinal fluid and the full coding sequence could be obtained. Specific PCR in cerebrospinal fluid and blood and serology with Immunoglobulin (Ig) M and IgG against Toscana virus confirmed our diagnosis. Neurological symptoms recovered spontaneously after 5 days. CONCLUSIONS: This case of Toscana virus infection highlights the benefits of unbiased metagenomic sequencing to support routine diagnostics in rare or unexpected viral infections. With increasing travel histories of patients, physicians should be aware of imported Toscana virus as the agent for viral meningitis and meningoencephalitis.

Low prevalence of transmitted HIV-1 drug resistance detected by a dried blood spot (DBS)-based next-generation sequencing (NGS) method in newly diagnosed individuals in Cameroon in the years 2015-16.

Objectives: To determine the most recent prevalence, transmission patterns and risk factors of transmitted drug-resistance mutations (TDRMs) in Cameroon, we initiated a multicentre study monitoring HIV-1 drug resistance in newly HIV-1-diagnosed individuals using a novel next-generation sequencing (NGS) assay applicable to fingerprick dried blood spot (DBS) samples. Methods: Fingerprick DBS samples and questionnaires were collected from 360 newly HIV-1-diagnosed individuals in four hospitals in urban areas in Cameroon in the years 2015-16. We developed an HIV-1 protease and reverse transcriptase drug resistance genotyping assay applicable to DBS samples and HIV-1 genomes of groups M, N and O. The WHO 2009 list of mutations for surveillance of transmitted drug-resistant HIV strains was used to analyse TDRMs. Results: Applying our 'DBS-NGS-genotypic resistance test', baseline HIV-1 drug resistance data were successfully obtained from 82.8% (298/360) of newly diagnosed individuals. At nucleotide frequencies >15%, TDRMs to NRTIs were observed in 3.0% (9/298), to NNRTIs in 4.0% (12/298) and to PIs in 1.3% (3/240). The NNRTI mutation K103N was most commonly detected (2.7%). Expanding the analysis to low-abundance TDRMs, i.e. 3%-15%, 12 additional individuals (4.0%) harbouring TDRMs were identified. Having unprotected sex with a known HIV-1-positive person was significantly associated with the transmission of DRMs (adjusted OR 9.6; 95% CI 1.79-51.3). Conclusions: The prevalence of transmitted HIV-1 drug resistance is currently low in the study sites in Cameroon. Evidence of some risky sexual behaviours depicts a public health problem with possible implications for the prevention of new HIV-1 infections.

Full-length haplotype reconstruction to infer the structure of heterogeneous virus populations.

Next-generation sequencing (NGS) technologies enable new insights into the diversity of virus populations within their hosts. Diversity estimation is currently restricted to single-nucleotide variants or to local fragments of no more than a few hundred nucleotides defined by the length of sequence reads. To study complex heterogeneous virus populations comprehensively, novel methods are required that allow for complete reconstruction of the individual viral haplotypes. Here, we show that assembly of whole viral genomes of ∼8600 nucleotides length is feasible from mixtures of heterogeneous HIV-1 strains derived from defined combinations of cloned virus strains and from clinical samples of an HIV-1 superinfected individual. Haplotype reconstruction was achieved using optimized experimental protocols and computational methods for amplification, sequencing and assembly. We comparatively assessed the performance of the three NGS platforms 454 Life Sciences/Roche, Illumina and Pacific Biosciences for this task. Our results prove and delineate the feasibility of NGS-based full-length viral haplotype reconstruction and provide new tools for studying evolution and pathogenesis of viruses.

A new fish-based multi-metric assessment index for cyprinid streams in the Iranian Caspian Sea Basin.

A major issue for water resource management is the assessment of environmental degradation of lotic ecosystems. The overall aim of this study is to develop a multi-metric fish index for the cyprinid streams of the Caspian Sea Basin (MMICS) in Iran. As species diversity and composition as well as population structure in the studied streams are different to other regions, there is a substantial need to develop a new fish index. We sampled fish and environmental data of 102 sites in medium sized streams. We analysed human pressures at different spatial scales and determined applicable fish metrics showing a response to human pressures. In total, five structural and functional types of metrics (i.e. biodiversity, habitat, reproduction, trophic level and water quality sensitivity) were considered. In addition, we used 29 criteria describing major anthropogenic human pressures at sampling sites and generated a regional pressure index (RPI) that accounted for potential effects of multiple human pressures. For the MMICS development, we first defined reference sites (least disturbed) and secondly quantified differences of fish metrics between reference and impaired sites. We used a Generalised Linear Model (GLM) to describe metric responses to natural environmental differences in least disturbed conditions. By including impaired sites, the residual distributions of these models described the response range of each metric to human pressures, independently of natural environmental influence. Finally, seven fish metrics showed the best ability to discriminate between impaired and reference sites. The multi-metric fish index performed well in discriminating human pressure classes, giving a significant negative linear response to a gradient of the RPI. These methods can be used for further development of a standardised monitoring tool to assess the ecological status and trends in biological condition for streams of the whole country, considering its complex and diverse geology and climate.

Predicting presence and absence of trout (Salmo trutta) in Iran.

Species distribution modelling, as a central issue in freshwater ecology, is an important tool for conservation and management of aquatic ecosystems. The brown trout (Salmo trutta) is a sensitive species which reacts to habitat changes induced by human impacts. Therefore, the identification of suitable habitats is essential. This study explores the potential distribution of brown trout by a species distribution modelling approach for Iran. Furthermore, modelling results are compared to the distribution described in the literature. Areas outside the currently known distribution which may offer potential habitats for brown trout are identified. The species distribution modelling was based on five different modelling techniques: Generalised Linear Model, Generalised Additive Model, Generalised Boosting Model, Classification Tree Analysis and Random Forests, which are finally summarised in an ensemble forecasting approach. We considered four environmental descriptors at the local scale (slope, bankfull width, wetted width, and elevation) and three climatic parameters (mean air temperature, range of air temperature and annual precipitation) which were extracted on three different spatial extents (1/5/10 km). The performance of all models was excellent (≥0.8) according to the TSS (True Skill Statistic) criterion. Slope, mean and range of air temperature were the most important variables in predicting brown trout occurrence. Presented results deepen the knowledge about distribution patterns of brown trout in Iran. Moreover, this study gives a basic background for the future development of assessment methods for riverine ecosystems in Iran.

Retrospective Genotyping of Enteroviruses Using a Diagnostic Nanopore Sequencing Workflow.

Enteroviruses are among the most common viruses pathogenic to humans. They are associated with various forms of disease, ranging from mild respiratory illness to severe neurological diseases. In recent years, an increasing number of isolated cases of children developing meningitis or encephalitis as a result of enterovirus infection have been reported, as well as discrete enterovirus D68 outbreaks in North America in 2014 and 2016. We developed an assay to rapidly genotype enteroviruses by sequencing a region within the VP1 gene using nanopore Flongles. We retrospectively analyzed enterovirus-/rhinovirus-positive clinical samples from the Zurich, Switzerland area mainly collected during two seasons in 2019/2020 and 2021/2022. Respiratory, cerebrospinal fluid, and stool samples were analyzed. Whole-genome sequencing was performed on samples with ambiguous genotyping results and enterovirus D68-positive samples. Out of 255 isolates, a total of 95 different genotypes were found. A difference in the prevalence of enterovirus and rhinovirus infections was observed for both sample type and age group. In particular, children aged 0-4 years showed a higher frequency of enterovirus infections. Comparing the respiratory seasons, a higher prevalence was found, especially for enterovirus A and rhinovirus A after the SARS-CoV-2 pandemic. The enterovirus genotyping workflow provides a rapid diagnostic tool for individual analysis and continuous enterovirus surveillance.

Exploring viral aetiology in upper respiratory tract infections: insights from metagenomic next-generation sequencing in Swiss outpatients before and during the SARS-CoV-2 pandemic.

AIMS OF THE STUDY: Upper respiratory tract infections are among the most common reasons for primary care consultations. They are diagnosed predominantly based on clinical assessment. Here, we investigated the benefit of viral metagenomic next-generation sequencing (mNGS) in an outpatient setting. METHODS: This prospective cross-sectional study included immunocompetent patients with acute upper respiratory tract infections. General practitioners collected pharyngeal swabs and demographic and clinical data. Specimens were analysed using viral mNGS and conventional tests. RESULTS: Two hundred seventy-seven patients were recruited by 21 general practitioners between 10/2019 and 12/2020, of which 91% had a suspected viral aetiology. For 138 patients (49.8%), mNGS identified one or more respiratory viruses. The mNGS showed a high overall agreement with conventional routine diagnostic tests. Rhinoviruses were the most frequently detected respiratory viruses (20.2% of patients). Viral mNGS reflected the influenza wave in early 2020 and the SARS-CoV-2 pandemic outbreak in Switzerland in March 2020. Notably, rhinoviruses continued to circulate despite non-pharmaceutical hygiene measures. CONCLUSIONS: Viral mNGS allowed the initial diagnosis to be retrospectively re-evaluated. Assuming reduced turnaround times, mNGS has the potential to directly guide the treatment of upper respiratory tract infections. On an epidemiological level, our study highlights the utility of mNGS in respiratory infection surveillance, allowing early detection of epidemics and providing information crucial for prevention.

Multicenter benchmarking of short and long read wet lab protocols for clinical viral metagenomics.

Metagenomics is gradually being implemented for diagnosing infectious diseases. However, in-depth protocol comparisons for viral detection have been limited to individual sets of experimental workflows and laboratories. In this study, we present a benchmark of metagenomics protocols used in clinical diagnostic laboratories initiated by the European Society for Clinical Virology (ESCV) Network on NGS (ENNGS). A mock viral reference panel was designed to mimic low biomass clinical specimens. The panel was used to assess the performance of twelve metagenomic wet lab protocols currently in use in the diagnostic laboratories of participating ENNGS member institutions. Both Illumina and Nanopore, shotgun and targeted capture probe protocols were included. Performance metrics sensitivity, specificity, and quantitative potential were assessed using a central bioinformatics pipeline. Overall, viral pathogens with loads down to 104 copies/ml (corresponding to CT values of 31 in our PCR assays) were detected by all the evaluated metagenomic wet lab protocols. In contrast, lower abundant mixed viruses of CT values of 35 and higher were detected only by a minority of the protocols. Considering the reference panel as the gold standard, optimal thresholds to define a positive result were determined per protocol, based on the horizontal genome coverage. Implementing these thresholds, sensitivity and specificity of the protocols ranged from 67 to 100 % and 87 to 100 %, respectively. A variety of metagenomic protocols are currently in use in clinical diagnostic laboratories. Detection of low abundant viral pathogens and mixed infections remains a challenge, implying the need for standardization of metagenomic analysis for use in clinical settings.

Pressure-specific and multiple pressure response of fish assemblages in European running waters.

We classified homogenous river types across Europe and searched for fish metrics qualified to show responses to specific pressures (hydromorphological pressures or water quality pressures) vs. multiple pressures in these river types. We analysed fish taxa lists from 3105 sites in 16 ecoregions and 14 countries. Sites were pre-classified for 15 selected pressures to separate unimpacted from impacted sites. Hierarchical cluster analysis was used to split unimpacted sites into four homogenous river types based on species composition and geographical location. Classification trees were employed to predict associated river types for impacted sites with four environmental variables. We defined a set of 129 candidate fish metrics to select the best reacting metrics for each river type. The candidate metrics represented tolerances/intolerances of species associated with six metric types: habitat, migration, water quality sensitivity, reproduction, trophic level and biodiversity. The results showed that 17 uncorrelated metrics reacted to pressures in the four river types. Metrics responded specifically to water quality pressures and hydromorphological pressures in three river types and to multiple pressures in all river types. Four metrics associated with water quality sensitivity showed a significant reaction in up to three river types, whereas 13 metrics were specific to individual river types. Our results contribute to the better understanding of fish assemblage response to human pressures at a pan-European scale. The results are especially important for European river management and restoration, as it is necessary to uncover underlying processes and effects of human pressures on aquatic communities.

Rapid and sensitive single-sample viral metagenomics using Nanopore Flongle sequencing.

The ability of viral metagenomic Next-Generation Sequencing (mNGS) to unbiasedly detect nucleic acids in a clinical sample is a powerful tool for advanced diagnosis of viral infections. When clinical symptoms do not provide a clear differential diagnosis, extensive laboratory testing with virus-specific PCR and serology can be replaced by a single viral mNGS analysis. However, widespread diagnostic use of viral mNGS is thus far limited by long sample-to-result times, as most protocols rely on Illumina sequencing, which provides high and accurate sequencing output but is time-consuming and expensive. Here, we describe the development of an mNGS protocol based on the more cost-effective Nanopore Flongle sequencing with decreased turnaround time and lower, yet sufficient sequencing output to provide sensitive virus detection. Sample preparation (6 h) and sequencing (2 h) times are substantially reduced compared to Illumina mNGS and allow detection of DNA/RNA viruses at low input (up to 33-38 cycle threshold of specific qPCR). Although Flongles yield lower sequencing output, direct comparison with Illumina mNGS on diverse clinical samples showed similar results. Collectively, the novel Nanopore mNGS approach is specifically tailored for use in clinical diagnostics and provides a rapid and cost-effective mNGS strategy for individual testing of severe cases.

Cross-continental evaluation of landscape-scale drivers and their impacts to fluvial fishes: Understanding frequency and severity to improve fish conservation in Europe and the United States.

Fluvial fishes are threatened globally from intensive human landscape stressors degrading aquatic ecosystems. However, impacts vary regionally, as stressors and natural environmental factors differ between ecoregions and continents. To date, a comparison of fish responses to landscape stressors over continents is lacking, limiting understanding of consistency of impacts and hampering efficiencies in conserving fishes over large regions. This study addresses these shortcomings through a novel, integrative assessment of fluvial fishes throughout Europe and the conterminous United States. Using large-scale datasets, including information on fish assemblages from more than 30,000 locations on both continents, we identified threshold responses of fishes summarized by functional traits to landscape stressors including agriculture, pasture, urban area, road crossings, and human population density. After summarizing stressors by catchment unit (local and network) and constraining analyses by stream size (creeks vs. rivers), we analyzed stressor frequency (number of significant thresholds) and stressor severity (value of identified thresholds) within ecoregions across Europe and the United States. We document hundreds of responses of fish metrics to multi-scale stressors in ecoregions across two continents, providing rich findings to aid in understanding and comparing threats to fishes across the study regions. Collectively, we found that lithophilic species and, as expected, intolerant species are most sensitive to stressors in both continents, while migratory and rheophilic species are similarly strongly affected in the United States. Also, urban land use and human population density were most frequently associated with declines in fish assemblages, underscoring the pervasiveness of these stressors in both continents. This study offers an unprecedented comparison of landscape stressor effects on fluvial fishes in a consistent and comparable manner, supporting conservation of freshwater habitats in both continents and worldwide.

Assessing immunogenicity barriers of the HIV-1 envelope trimer.

Understanding the balance between epitope shielding and accessibility on HIV-1 envelope (Env) trimers is essential to guide immunogen selection for broadly neutralizing antibody (bnAb) based vaccines. To investigate the antigenic space of Env immunogens, we created a strategy based on synthetic, high diversity, Designed Ankyrin Repeat Protein (DARPin) libraries. We show that DARPin Antigenicity Analysis (DANA), a purely in vitro screening tool, has the capability to extrapolate relevant information of antigenic properties of Env immunogens. DANA screens of stabilized, soluble Env trimers revealed that stronger trimer stabilization led to the selection of highly mutated DARPins with length variations and framework mutations mirroring observations made for bnAbs. By mimicking heterotypic prime-boost immunization regimens, DANA may be used to select immunogen combinations that favor the selection of trimer-reactive binders. This positions DANA as a versatile strategy for distilling fundamental antigenic features of immunogens, complementary to preclinical immunogenicity testing.

Swiss public health measures associated with reduced SARS-CoV-2 transmission using genome data.

Genome sequences from evolving infectious pathogens allow quantification of case introductions and local transmission dynamics. We sequenced 11,357 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomes from Switzerland in 2020-the sixth largest effort globally. Using a representative subset of these data, we estimated viral introductions to Switzerland and their persistence over the course of 2020. We contrasted these estimates with simple null models representing the absence of certain public health measures. We show that Switzerland's border closures decoupled case introductions from incidence in neighboring countries. Under a simple model, we estimate an 86 to 98% reduction in introductions during Switzerland's strictest border closures. Furthermore, the Swiss 2020 partial lockdown roughly halved the time for sampled introductions to die out. Last, we quantified local transmission dynamics once introductions into Switzerland occurred using a phylodynamic model. We found that transmission slowed 35 to 63% upon outbreak detection in summer 2020 but not in fall. This finding may indicate successful contact tracing over summer before overburdening in fall. The study highlights the added value of genome sequencing data for understanding transmission dynamics.

Secondary attack rates from asymptomatic and symptomatic influenza virus shedders in hospitals: Results from the TransFLUas influenza transmission study.

OBJECTIVE: Nosocomial transmission of influenza is a major concern for infection control. We aimed to dissect transmission dynamics of influenza, including asymptomatic transmission events, in acute care. DESIGN: Prospective surveillance study during 2 influenza seasons. SETTING: Tertiary-care hospital. PARTICIPANTS: Volunteer sample of inpatients on medical wards and healthcare workers (HCWs). METHODS: Participants provided daily illness diaries and nasal swabs for influenza A and B detection and whole-genome sequencing for phylogenetic analyses. Contacts between study participants were tracked. Secondary influenza attack rates were calculated based on spatial and temporal proximity and phylogenetic evidence for transmission. RESULTS: In total, 152 HCWs and 542 inpatients were included; 16 HCWs (10.5%) and 19 inpatients (3.5%) tested positive for influenza on 109 study days. Study participants had symptoms of disease on most of the days they tested positive for influenza (83.1% and 91.9% for HCWs and inpatients, respectively). Also, 11(15.5%) of 71 influenza-positive swabs among HCWs and 3 (7.9%) of 38 influenza-positive swabs among inpatients were collected on days without symptoms; 2 (12.5%) of 16 HCWs and 2 (10.5%) of 19 inpatients remained fully asymptomatic. The secondary attack rate was low: we recorded 1 transmission event over 159 contact days (0.6%) that originated from a symptomatic case. No transmission event occurred in 61 monitored days of contacts with asymptomatic influenza-positive individuals. CONCLUSIONS: Influenza in acute care is common, and individuals regularly shed influenza virus without harboring symptoms. Nevertheless, both symptomatic and asymptomatic transmission events proved rare. We suggest that healthcare-associated influenza prevention strategies that are based on preseason vaccination and barrier precautions for symptomatic individuals seem to be effective.

Antibodies from convalescent plasma promote SARS-CoV-2 clearance in individuals with and without endogenous antibody response.

BACKGROUNDNeutralizing antibodies are considered a key correlate of protection by current SARS-CoV-2 vaccines. The manner in which human infections respond to therapeutic SARS-CoV-2 antibodies, including convalescent plasma therapy, remains to be fully elucidated.METHODSWe conducted a proof-of-principle study of convalescent plasma therapy based on a phase I trial in 30 hospitalized COVID-19 patients with a median interval between onset of symptoms and first transfusion of 9 days (IQR, 7-11.8 days). Comprehensive longitudinal monitoring of the virological, serological, and disease status of recipients allowed deciphering of parameters on which plasma therapy efficacy depends.RESULTSIn this trial, convalescent plasma therapy was safe as evidenced by the absence of transfusion-related adverse events and low mortality (3.3%). Treatment with highly neutralizing plasma was significantly associated with faster virus clearance, as demonstrated by Kaplan-Meier analysis (P = 0.034) and confirmed in a parametric survival model including viral load and comorbidity (adjusted hazard ratio, 3.0; 95% CI, 1.1-8.1; P = 0.026). The onset of endogenous neutralization affected viral clearance, but even after adjustment for their pretransfusion endogenous neutralization status, recipients benefitted from plasma therapy with high neutralizing antibodies (hazard ratio, 3.5; 95% CI, 1.1-11; P = 0.034).CONCLUSIONOur data demonstrate a clear impact of exogenous antibody therapy on the rapid clearance of viremia before and after onset of the endogenous neutralizing response, and point beyond antibody-based interventions to critical laboratory parameters for improved evaluation of current and future SARS-CoV-2 therapies.TRIAL REGISTRATIONClinicalTrials.gov NCT04869072.FUNDINGThis study was funded via an Innovation Pool project by the University Hospital Zurich; the Swiss Red Cross Glückskette Corona Funding; Pandemiefonds of the UZH Foundation; and the Clinical Research Priority Program "Comprehensive Genomic Pathogen Detection" of the University of Zurich.