Publications by authors named "Chantal B E M Reusken"

59 Publications

Towards a sensitive and accurate interpretation of molecular testing for SARS-CoV-2: a rapid review of 264 studies.

Euro Surveill 2021 03;26(10)

Global Outbreak Alert and Response Network (GOARN), Geneva, Switzerland.

BackgroundSensitive molecular diagnostics and correct test interpretation are crucial for accurate COVID-19 diagnosis and thereby essential for good clinical practice. Furthermore, they are a key factor in outbreak control where active case finding in combination with isolation and contact tracing are crucial.AimWith the objective to inform the public health and laboratory responses to the pandemic, we reviewed current published knowledge on the kinetics of SARS-CoV-2 infection as assessed by RNA molecular detection in a wide range of clinical samples.MethodsWe performed an extensive search on studies published between 1 December 2019 and 15 May 2020, reporting on molecular detection and/or isolation of SARS-CoV-2 in any human laboratory specimen.ResultsWe compiled a dataset of 264 studies including 32,515 COVID-19 cases, and additionally aggregated data points (n = 2,777) from sampling of 217 adults with known infection timeline. We summarised data on SARS-CoV-2 detection in the respiratory and gastrointestinal tract, blood, oral fluid, tears, cerebrospinal fluid, peritoneal fluid, semen, vaginal fluid; where provided, we also summarised specific observations on SARS-CoV-2 detection in pregnancy, infancy, children, adolescents and immunocompromised individuals.ConclusionOptimal SARS-CoV-2 molecular testing relies on choosing the most appropriate sample type, collected with adequate sampling technique, and with the infection timeline in mind. We outlined knowledge gaps and directions for future well-documented systematic studies.
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http://dx.doi.org/10.2807/1560-7917.ES.2021.26.10.2001134DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7953531PMC
March 2021

Possible host-adaptation of SARS-CoV-2 due to improved ACE2 receptor binding in mink.

Virus Evol 2021 Jan 4;7(1):veaa094. Epub 2021 Jan 4.

Department of Medical Microbiology & Infection Prevention, Amsterdam University Medical Centers, location AMC, Meibergdreef 9, 1105AZ, Amsterdam, The Netherlands.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections on mink farms are increasingly observed in several countries, leading to the massive culling of animals on affected farms. Recent studies showed multiple (anthropo)zoonotic transmission events between humans and mink on these farms. Mink-derived SARS-CoV-2 sequences from The Netherlands and Denmark contain multiple substitutions in the S protein receptor binding domain (RBD). Molecular modeling showed that these substitutions increase the mean binding energy, suggestive of potential adaptation of the SARS-CoV-2 S protein to the mink angiotensin-converting enzyme 2 (ACE2) receptor. These substitutions could possibly also impact human ACE2 binding affinity as well as humoral immune responses directed to the RBD region of the SARS-CoV-2 S protein in humans. We wish to highlight these observations to raise awareness and urge for the continued surveillance of mink (and other animal)-related infections.
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http://dx.doi.org/10.1093/ve/veaa094DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7798643PMC
January 2021

Development of a Comparative European Orthohantavirus Microneutralization Assay With Multi- Species Validation and Evaluation in a Human Diagnostic Cohort.

Front Cell Infect Microbiol 2020 22;10:580478. Epub 2020 Dec 22.

Centre for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, Netherlands.

Orthohantaviruses (family , order ) can cause two serious syndromes in humans: hemorrhagic fever with renal syndrome (HFRS), associated with the Old World orthohantaviruses, and hantavirus cardiopulmonary syndrome (HCPS), associated with orthohantaviruses in the Americas. In Europe, four different orthohantaviruses (DOBV, PUUV, SEOV, and TULV) are associated with human disease. As disease severity and zoonotic source differ between orthohantavirus species, conclusive determination of the infecting species by either RT-PCR or comparative virus neutralization test (VNT) is of importance. Currently, the focus reduction neutralization test (FRNT) is considered the 'Gold Standard' for orthohantavirus VNTs, however this test is laborious and time-consuming. Consequently, more high-throughput alternatives are needed. In this study, we developed a comparative orthohantavirus microneutralization test (MNT) including all four human pathogenic orthohantavirus species circulating in Europe. The assay was validated using RT-PCR-confirmed rodent (n=17) and human sera (n=17), DOBV-suspected human sera (n=3) and cohorts of orthohantavirus-negative rodent (n=3) and human sera (n=85). 16/17 RT-PCR-confirmed rodent sera and 18/20 of the RT-PCR-confirmed and DOBV-suspected human sera were serotyped successfully, while for the remaining rodent (n=1) and human sera (n=2) no neutralizing titers could be detected. All negative control sera tested negative in the MNT. The assay was subsequently evaluated using a clinical cohort of 50 orthohantavirus patients. Orthohantavirus infection was confirmed in all 50 patients, and 47/50 (94%) sera were serotyped successfully, confirming PUUV as the major cause of orthohantavirus infections in Netherlands. Notably, two previously unrecognized SEOV cases from 2013 were diagnosed using the MNT, underlining the added value of the MNT in a diagnostic setting. In conclusion, we demonstrate the successful development and clinical implementation of a comparative European orthohantavirus MNT to determine the infecting virus species in European HFRS patients. Identification of the causative species is needed for an adequate Public Health response and can support individual patient care. For many labs, the implementation of orthohantavirus neutralization tests has not been a straightforward procedure. This issue will be addressed by the rollout of the comparative MNT to multiple European laboratories to support patient diagnostics, surveillance and Public Health responses.
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http://dx.doi.org/10.3389/fcimb.2020.580478DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7783042PMC
December 2020

Variable Sensitivity of SARS-CoV-2 Molecular Detection in European Expert Laboratories: External Quality Assessment, June and July 2020.

J Clin Microbiol 2021 02 18;59(3). Epub 2021 Feb 18.

Centre for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, the Netherlands

During the ongoing coronavirus disease 2019 (COVID-19) outbreak, robust detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a key element for clinical management and to interrupt transmission chains. We organized an external quality assessment (EQA) of molecular detection of SARS-CoV-2 for European expert laboratories. An EQA panel composed of 12 samples, containing either SARS-CoV-2 at different concentrations to evaluate sensitivity or other respiratory viruses to evaluate specificity of SARS-CoV-2 testing, was distributed to 68 laboratories in 35 countries. Specificity samples included seasonal human coronaviruses hCoV-229E, hCoV-NL63, and hCoV-OC43, as well as Middle East respiratory syndrome coronavirus (MERS-CoV), SARS-CoV, and human influenza viruses A and B. Sensitivity results differed among laboratories, particularly for low-concentration SARS-CoV-2 samples. Results indicated that performance was mostly independent of the selection of specific extraction or PCR methods.
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http://dx.doi.org/10.1128/JCM.02676-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8106723PMC
February 2021

Low SARS-CoV-2 seroprevalence in blood donors in the early COVID-19 epidemic in the Netherlands.

Nat Commun 2020 11 12;11(1):5744. Epub 2020 Nov 12.

Department of Blood-borne Infections, Sanquin Research, Amsterdam, The Netherlands.

The world is combating an ongoing COVID-19 pandemic with health-care systems, society and economies impacted in an unprecedented way. It is unclear how many people have contracted the causative coronavirus (SARS-CoV-2) unknowingly and are asymptomatic. Therefore, reported COVID-19 cases do not reflect the true scale of outbreak. Here we present the prevalence and distribution of antibodies to SARS-CoV-2 in a healthy adult population of the Netherlands, which is a highly affected country, using a high-performance immunoassay. Our results indicate that one month into the outbreak (i) the seroprevalence in the Netherlands was 2.7% with substantial regional variation, (ii) the hardest-hit areas showed a seroprevalence of up to 9.5%, (iii) the seroprevalence was sex-independent throughout age groups (18-72 years), and (iv) antibodies were significantly more often present in younger people (18-30 years). Our study provides vital information on the extent of exposure to SARS-CoV-2 in a country where social distancing is in place.
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http://dx.doi.org/10.1038/s41467-020-19481-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7665189PMC
November 2020

Spatial risk analysis for the introduction and circulation of six arboviruses in the Netherlands.

Parasit Vectors 2020 Sep 10;13(1):464. Epub 2020 Sep 10.

Wildlife Ecology & Conservation Group, Wageningen University & Research, Wageningen, The Netherlands.

Background: Arboviruses are a growing public health concern in Europe, with both endemic and exotic arboviruses expected to spread further into novel areas in the next decades. Predicting where future outbreaks will occur is a major challenge, particularly for regions where these arboviruses are not endemic. Spatial modelling of ecological risk factors for arbovirus circulation can help identify areas of potential emergence. Moreover, combining hazard maps of different arboviruses may facilitate a cost-efficient, targeted multiplex-surveillance strategy in areas where virus transmission is most likely. Here, we developed predictive hazard maps for the introduction and/or establishment of six arboviruses that were previously prioritized for the Netherlands: West Nile virus, Japanese encephalitis virus, Rift Valley fever virus, tick-borne encephalitis virus, louping-ill virus and Crimean-Congo haemorrhagic fever virus.

Methods: Our spatial model included ecological risk factors that were identified as relevant for these arboviruses by an earlier systematic review, including abiotic conditions, vector abundance, and host availability. We used geographic information system (GIS)-based tools and geostatistical analyses to model spatially continuous datasets on these risk factors to identify regions in the Netherlands with suitable ecological conditions for arbovirus introduction and establishment.

Results: The resulting hazard maps show that there is spatial clustering of areas with either a relatively low or relatively high environmental suitability for arbovirus circulation. Moreover, there was some overlap in high-hazard areas for virus introduction and/or establishment, particularly in the southern part of the country.

Conclusions: The similarities in environmental suitability for some of the arboviruses provide opportunities for targeted sampling of vectors and/or sentinel hosts in these potential hotspots of emergence, thereby increasing the efficient use of limited resources for surveillance.
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http://dx.doi.org/10.1186/s13071-020-04339-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7488554PMC
September 2020

Orthohantavirus Pathogenesis and Cell Tropism.

Front Cell Infect Microbiol 2020 4;10:399. Epub 2020 Aug 4.

Department of Viroscience, Erasmus University Medical Center, Rotterdam, Netherlands.

Orthohantaviruses are zoonotic viruses that are naturally maintained by persistent infection in specific reservoir species. Although these viruses mainly circulate among rodents worldwide, spill-over infection to humans occurs. Orthohantavirus infection in humans can result in two distinct clinical outcomes: hemorrhagic fever with renal syndrome (HFRS) and hantavirus cardiopulmonary syndrome (HCPS). While both syndromes develop following respiratory transmission and are associated with multi-organ failure and high mortality rates, little is known about the mechanisms that result in these distinct clinical outcomes. Therefore, it is important to identify which cell types and tissues play a role in the differential development of pathogenesis in humans. Here, we review current knowledge on cell tropism and its role in pathogenesis during orthohantavirus infection in humans and reservoir rodents. Orthohantaviruses predominantly infect microvascular endothelial cells (ECs) of a variety of organs (lungs, heart, kidney, liver, and spleen) in humans. However, in this review we demonstrate that other cell types (e.g., macrophages, dendritic cells, and tubular epithelium) are infected as well and may play a role in the early steps in pathogenesis. A key driver for pathogenesis is increased vascular permeability, which can be direct effect of viral infection in ECs or result of an imbalanced immune response in an attempt to clear the virus. Future studies should focus on the role of identifying how infection of organ-specific endothelial cells as well as other cell types contribute to pathogenesis.
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http://dx.doi.org/10.3389/fcimb.2020.00399DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7438779PMC
August 2020

Response to letter of concern by Oladimeji and Pickford of PrimerDesign.

J Clin Virol 2020 08 29;129:104526. Epub 2020 Jun 29.

National Institute for Public Health and the Environment (RIVM), Center for Infectious Disease Control (CIb), Bilthoven, the Netherlands. Electronic address:

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http://dx.doi.org/10.1016/j.jcv.2020.104526DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7323679PMC
August 2020

Comparison of seven commercial RT-PCR diagnostic kits for COVID-19.

J Clin Virol 2020 07 8;128:104412. Epub 2020 May 8.

National Institute for Public Health and the Environment, Center for Infectious Disease Control, A. van Leeuwenhoeklaan 9, 3721 MA, Bilthoven, the Netherlands. Electronic address:

The final months of 2019 witnessed the emergence of a novel coronavirus in the human population. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has since spread across the globe and is posing a major burden on society. Measures taken to reduce its spread critically depend on timely and accurate identification of virus-infected individuals by the most sensitive and specific method available, i.e. real-time reverse transcriptase PCR (RT-PCR). Many commercial kits have recently become available, but their performance has not yet been independently assessed. The aim of this study was to compare basic analytical and clinical performance of selected RT-PCR kits from seven different manufacturers (Altona Diagnostics, BGI, CerTest Biotec, KH Medical, PrimerDesign, R-Biopharm AG, and Seegene). We used serial dilutions of viral RNA to establish PCR efficiency and estimate the 95 % limit of detection (LOD95). Furthermore, we ran a panel of SARS-CoV-2-positive clinical samples (n = 13) for a preliminary evaluation of clinical sensitivity. Finally, we used clinical samples positive for non-coronavirus respiratory viral infections (n = 6) and a panel of RNA from related human coronaviruses to evaluate assay specificity. PCR efficiency was ≥96 % for all assays and the estimated LOD95 varied within a 6-fold range. Using clinical samples, we observed some variations in detection rate between kits. Importantly, none of the assays showed cross-reactivity with other respiratory (corona)viruses, except as expected for the SARS-CoV-1 E-gene. We conclude that all RT-PCR kits assessed in this study may be used for routine diagnostics of COVID-19 in patients by experienced molecular diagnostic laboratories.
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http://dx.doi.org/10.1016/j.jcv.2020.104412DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7206434PMC
July 2020

The invasive Asian bush mosquito Aedes japonicus found in the Netherlands can experimentally transmit Zika virus and Usutu virus.

PLoS Negl Trop Dis 2020 04 13;14(4):e0008217. Epub 2020 Apr 13.

Laboratory of Virology, Wageningen University & Research, Wageningen, the Netherlands.

Background: The Asian bush mosquito Aedes japonicus is invading Europe and was first discovered in Lelystad, the Netherlands in 2013, where it has established a permanent population. In this study, we investigated the vector competence of Ae. japonicus from the Netherlands for the emerging Zika virus (ZIKV) and zoonotic Usutu virus (USUV). ZIKV causes severe congenital microcephaly and Guillain-Barré syndrome in humans. USUV is closely related to West Nile virus, has recently spread throughout Europe and is causing mass mortality of birds. USUV infection in humans can result in clinical manifestations ranging from mild disease to severe neurological impairments.

Methodology/principal Findings: In our study, field-collected Ae. japonicus females received an infectious blood meal with ZIKV or USUV by droplet feeding. After 14 days at 28°C, 3% of the ZIKV-blood fed mosquitoes and 13% of the USUV-blood fed mosquitoes showed virus-positive saliva, indicating that Ae. japonicus can transmit both viruses. To investigate the effect of the mosquito midgut barrier on virus transmission, female mosquitoes were intrathoracically injected with ZIKV or USUV. Of the injected mosquitoes, 96% (ZIKV) and 88% (USUV) showed virus-positive saliva after 14 days at 28°C. This indicates that ZIKV and USUV can efficiently replicate in Ae. japonicus but that a strong midgut barrier is normally restricting virus dissemination. Small RNA deep sequencing of orally infected mosquitoes confirmed active replication of ZIKV and USUV, as demonstrated by potent small interfering RNA responses against both viruses. Additionally, de novo small RNA assembly revealed the presence of a novel narnavirus in Ae. japonicus.

Conclusions/significance: Given that Ae. japonicus can experimentally transmit arthropod-borne viruses (arboviruses) like ZIKV and USUV and is currently expanding its territories, we should consider this mosquito as a potential vector for arboviral diseases in Europe.
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http://dx.doi.org/10.1371/journal.pntd.0008217DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7153878PMC
April 2020

Severe Acute Respiratory Syndrome Coronavirus 2-Specific Antibody Responses in Coronavirus Disease Patients.

Emerg Infect Dis 2020 Jul 21;26(7):1478-1488. Epub 2020 Jun 21.

A new coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has recently emerged to cause a human pandemic. Although molecular diagnostic tests were rapidly developed, serologic assays are still lacking, yet urgently needed. Validated serologic assays are needed for contact tracing, identifying the viral reservoir, and epidemiologic studies. We developed serologic assays for detection of SARS-CoV-2 neutralizing, spike protein-specific, and nucleocapsid-specific antibodies. Using serum samples from patients with PCR-confirmed SARS-CoV-2 infections, other coronaviruses, or other respiratory pathogenic infections, we validated and tested various antigens in different in-house and commercial ELISAs. We demonstrated that most PCR-confirmed SARS-CoV-2-infected persons seroconverted by 2 weeks after disease onset. We found that commercial S1 IgG or IgA ELISAs were of lower specificity, and sensitivity varied between the 2 assays; the IgA ELISA showed higher sensitivity. Overall, the validated assays described can be instrumental for detection of SARS-CoV-2-specific antibodies for diagnostic, seroepidemiologic, and vaccine evaluation studies.
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http://dx.doi.org/10.3201/eid2607.200841DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7323511PMC
July 2020

Serologic Detection of Middle East Respiratory Syndrome Coronavirus Functional Antibodies.

Emerg Infect Dis 2020 May 17;26(5):1024-1027. Epub 2020 May 17.

We developed and validated 2 species-independent protein-based assays to detect Middle East respiratory syndrome coronavirus functional antibodies that can block virus receptor-binding or sialic acid-attachment. Antibody levels measured in both assays correlated strongly with virus-neutralizing antibody titers, proving their use for serologic confirmatory diagnosis of Middle East respiratory syndrome.
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http://dx.doi.org/10.3201/eid2605.190921DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7181916PMC
May 2020

Laboratory readiness and response for novel coronavirus (2019-nCoV) in expert laboratories in 30 EU/EEA countries, January 2020.

Euro Surveill 2020 02 11;25(6). Epub 2020 Feb 11.

The participating members of EVD-LabNet and ERLI-Net are acknowledged at the end of the article.

Timely detection of novel coronavirus (2019-nCoV) infection cases is crucial to interrupt the spread of this virus. We assessed the required expertise and capacity for molecular detection of 2019-nCoV in specialised laboratories in 30 European Union/European Economic Area (EU/EEA) countries. Thirty-eight laboratories in 24 EU/EEA countries had diagnostic tests available by 29 January 2020. A coverage of all EU/EEA countries was expected by mid-February. Availability of primers/probes, positive controls and personnel were main implementation barriers.
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http://dx.doi.org/10.2807/1560-7917.ES.2020.25.6.2000082DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7029448PMC
February 2020

Faeces as a novel material to estimate lyssavirus prevalence in bat populations.

Zoonoses Public Health 2020 03 8;67(2):198-202. Epub 2019 Dec 8.

Department of Viroscience, Erasmus University Medical Centre, Rotterdam, The Netherlands.

Rabies is caused by infection with a lyssavirus. Bat rabies is of concern for both public health and bat conservation. The current method for lyssavirus prevalence studies in bat populations is by oral swabbing, which is invasive for the bats, dangerous for handlers, time-consuming and expensive. In many situations, such sampling is not feasible, and hence, our understanding of epidemiology of bat rabies is limited. Faeces are usually easy to collect from bat colonies without disturbing the bats and thus could be a practical and feasible material for lyssavirus prevalence studies. To further explore this idea, we performed virological analysis on faecal pellets and oral swabs of seven serotine bats (Eptesicus serotinus) that were positive for European bat 1 lyssavirus in the brain. We also performed immunohistochemical and virological analyses on digestive tract samples of these bats to determine potential sources of lyssavirus in the faeces. We found that lyssavirus detection by RT-qPCR was nearly as sensitive in faecal pellets (6/7 bats positive, 86%) as in oral swabs (7/7 bats positive, 100%). The likely source of lyssavirus in the faeces was virus excreted into the oral cavity from the salivary glands (5/6 bats positive by immunohistochemistry and RT-qPCR) or tongue (3/4 bats positive by immunohistochemistry) and swallowed with saliva. Virus could not be isolated from any of the seven faecal pellets, suggesting the lyssavirus detected in faeces is not infectious. Lyssavirus detection in the majority of faecal pellets of infected bats shows that this novel material should be further explored for lyssavirus prevalence studies in bats.
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http://dx.doi.org/10.1111/zph.12672DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7027462PMC
March 2020

Geographical Variability Affects CCHFV Detection by RT-PCR: A Tool for In-Silico Evaluation of Molecular Assays.

Viruses 2019 10 16;11(10). Epub 2019 Oct 16.

National Institute for Infectious Diseases (INMI) "L. Spallanzani" IRCCS, WHO Collaborating Center for clinical care, diagnosis, response and training on Highly Infectious Diseases, 00149 Rome, Italy.

The Crimean-Congo hemorrhagic fever virus (CCHFV) is considered to be a major emerging infectious threat, according to the WHO R&D blueprint. A wide range of CCHFV molecular assays have been developed, employing varied primer/probe combinations. The high genetic variability of CCHFV often hampers the efficacy of available molecular tests and can affect their diagnostic potential. Recently, increasing numbers of complete CCHFV genomic sequences have become available, allowing a better appreciation of the genomic evolution of this virus. We summarized the current knowledge on molecular methods and developed a new bioinformatics tool to evaluate the existing assays for CCHFV detection, with a special focus on strains circulating in different geographical areas. Twenty-two molecular methods and 181 sequences of CCHFV were collected, respectively, from PubMed and GenBank databases. Up to 28 mismatches between primers and probes of each assay and CCHFV strains were detected through in-silico PCR analysis. Combinations of up to three molecular methods markedly decreased the number of mismatches within most geographic areas. These results supported the good practice of CCHFV detection of performing more than one assay, aimed for different sequence targets. The choice of the most appropriate tests must take into account patient's travel history and geographic distribution of the different CCHFV strains.
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http://dx.doi.org/10.3390/v11100953DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6833031PMC
October 2019

Sensitive and Specific Detection of Low-Level Antibody Responses in Mild Middle East Respiratory Syndrome Coronavirus Infections.

Emerg Infect Dis 2019 10 17;25(10):1868-1877. Epub 2019 Oct 17.

Middle East respiratory syndrome coronavirus (MERS-CoV) infections in humans can cause asymptomatic to fatal lower respiratory lung disease. Despite posing a probable risk for virus transmission, asymptomatic to mild infections can go unnoticed; a lack of seroconversion among some PCR-confirmed cases has been reported. We found that a MERS-CoV spike S1 protein-based ELISA, routinely used in surveillance studies, showed low sensitivity in detecting infections among PCR-confirmed patients with mild clinical symptoms and cross-reactivity of human coronavirus OC43-positive serum samples. Using in-house S1 ELISA and protein microarray, we demonstrate that most PCR-confirmed MERS-CoV case-patients with mild infections seroconverted; nonetheless, some of these samples did not have detectable levels of virus-neutralizing antibodies. The use of a sensitive and specific serologic S1-based assay can be instrumental in the accurate estimation of MERS-CoV prevalence.
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http://dx.doi.org/10.3201/eid2510.190051DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6759241PMC
October 2019

Distribution of zoonotic variegated squirrel bornavirus 1 in naturally infected variegated and Prevost's squirrels.

Sci Rep 2019 08 6;9(1):11402. Epub 2019 Aug 6.

Institute of Veterinary Pathology, Justus Liebig University, Giessen, Germany.

Recently, the zoonotic capacity of the newly discovered variegated squirrel bornavirus 1 (VSBV-1) was confirmed in humans with a lethal encephalitis. Transmission to humans occurred by variegated and Prevost's squirrels as presumed reservoir hosts but possible ways of virus shedding and the route of infection still need to be elucidated. Thus, the tissue distribution of VSBV-1 antigen and RNA was investigated in detail via immunohistochemistry (IHC) in six variegated and eight Prevost's squirrels and by in situ hybridisation (ISH) in one Prevost's squirrel, respectively. VSBV-1 antigen and RNA positive cells were most numerous in the nervous system and were also found in nearly all tissues and different cell types indicating a broad organ and cell tropism of VSBV-1. Presence of VSBV-1 in several organs might indicate potential virus shedding via various routes and implies the risk of intra- and interspecies transmission, respectively.
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http://dx.doi.org/10.1038/s41598-019-47767-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6684602PMC
August 2019

Usutu virus infection in Dutch blood donors.

Transfusion 2019 09 4;59(9):2931-2937. Epub 2019 Jul 4.

National Screening Laboratory, Sanquin Blood Supply Foundation, Amsterdam, the Netherlands.

Background: The screening of Dutch blood donations for West Nile virus (WNV) may be imminent, as WNV emerges in nearby countries and more donors travel to WNV-affected regions. Since 2016 the related, mosquito-borne Usutu virus (USUV) causes seasonal mortality in Dutch birds. To what extent will human USUV infections affect Dutch WNV donor screening?

Study Design And Methods: From April through September 2018, plasma samples from blood donations in blackbird-rich regions were stored. When increased bird mortality was reported in August, samples from July, August, and September were tested for USUV-RNA in pools of eight, using a home-brew combined WNV/USUV-PCR assay. Reactive pools were deconstructed. Original plasma units and samples of previous and follow-up donations of reactive donors were tested for USUV- and WNV-RNA, and for antibody responses.

Results: The number of USUV RNA-positive, WNV RNA-negative donations was 0 of 2688 donations in July, 6 of 4416 in August (1:736), and 1 of 4936 in September. The seven infected donors tested negative for USUV-RNA in preceding and follow-up donations. For 6 donors, seroconversion for USUV-antibodies was demonstrated. All index donations tested positive in a commonly used PCR-assay for WNV donor screening. Three exposed recipients did not show signs of infection. Screening a random subset of 1092 donations from September for USUV-IgG antibodies showed that 22 donors tested reactive; for three donors retrospective testing identified an USUV PCR-positive pre-seroconversion donation.

Conclusion: Seasonal USUV infection in Dutch blood donors is common. Cross-reactivity in molecular assays for WNV-screening occurs, but can be resolved using USUV- and WNV-specific PCR-primers and sequencing of viral RNA.
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http://dx.doi.org/10.1111/trf.15444DOI Listing
September 2019

Risk factors associated with sustained circulation of six zoonotic arboviruses: a systematic review for selection of surveillance sites in non-endemic areas.

Parasit Vectors 2019 May 27;12(1):265. Epub 2019 May 27.

Department of Viroscience, WHO CC for arbovirus and viral hemorrhagic fever reference and research, Erasmus University Medical Centre, Rotterdam, The Netherlands.

Arboviruses represent a significant burden to public health and local economies due to their ability to cause unpredictable and widespread epidemics. To maximize early detection of arbovirus emergence in non-endemic areas, surveillance efforts should target areas where circulation is most likely. However, identifying such hotspots of potential emergence is a major challenge. The ecological conditions leading to arbovirus outbreaks are shaped by complex interactions between the virus, its vertebrate hosts, arthropod vector, and abiotic environment that are often poorly understood. Here, we systematically review the ecological risk factors associated with the circulation of six arboviruses that are of considerable concern to northwestern Europe. These include three mosquito-borne viruses (Japanese encephalitis virus, West Nile virus, Rift Valley fever virus) and three tick-borne viruses (Crimean-Congo hemorrhagic fever virus, tick-borne encephalitis virus, and louping-ill virus). We consider both intrinsic (e.g. vector and reservoir host competence) and extrinsic (e.g. temperature, precipitation, host densities, land use) risk factors, identify current knowledge gaps, and discuss future directions. Our systematic review provides baseline information for the identification of regions and habitats that have suitable ecological conditions for endemic circulation, and therefore may be used to target early warning surveillance programs aimed at detecting multi-virus and/or arbovirus emergence.
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http://dx.doi.org/10.1186/s13071-019-3515-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6537422PMC
May 2019

Whole-Blood Testing for Diagnosis of Acute Zika Virus Infections in Routine Diagnostic Setting.

Emerg Infect Dis 2019 07 17;25(7):1394-1396. Epub 2019 Jul 17.

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.
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http://dx.doi.org/10.3201/eid2507.182000DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6590769PMC
July 2019

[Yellow fever in Brazil: the importance of vaccination].

Ned Tijdschr Geneeskd 2018 09 24;162. Epub 2018 Sep 24.

Havenpolikliniek Rotterdam, Diagnostisch Centrum Tropenziekten.

Background: Since 2016 outbreaks of yellow fever are reported in Brazil. This is a risk to unvaccinated travellers in that area.

Case Description: In early January, an unvaccinated traveller returning from São Paulo attended our outpatient clinic complaining of symptoms later diagnosed as yellow fever. The disease manifested itself as fever, lower back pain, nausea and highly elevated liver enzymes. A yellow fever infection has multiple stages. The first stage is acute infection which merges into the second stage which is when improvement occurs. Either improvement continues or transfers into a third stage which is characterized by multi-organ failure. In this particular patient, stage three did not occur.

Conclusion: The goal of this case report is to show that vaccination against yellow fever is the most important preventive measure when travelling to an area where the yellow fever virus is in circulation. Yellow fever should not be forgotten in the differential diagnosis of a traveller with fever.
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September 2018

Need for additional capacity and improved capability for molecular detection of yellow fever virus in European Expert Laboratories: External Quality Assessment, March 2018.

Euro Surveill 2018 07;23(28)

Department of Viroscience, World Health Organization (WHO) Collaborating Centre for Arbovirus and Haemorrhagic Fever Reference and Research, Erasmus MC, Rotterdam, The Netherlands.

An external quality assessment of yellow fever virus (YFV) molecular detection in European laboratories was organised in rapid response to an increase in human cases in Brazil in 2018 with risk of import to Europe. Detection of YFV was assessed among 32 laboratories in 23/31 European Union (EU) and European Economic Area (EEA) countries and two laboratories in one non-EU/EEA country. Adequate capabilities were lacking in 10/23 countries; five did not participate as they lacked implemented assays.
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http://dx.doi.org/10.2807/1560-7917.ES.2018.23.28.1800341DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6152149PMC
July 2018

Diagnosis of Zika Virus Infection by Peptide Array and Enzyme-Linked Immunosorbent Assay.

mBio 2018 03 6;9(2). Epub 2018 Mar 6.

Center for Infection and Immunity, Mailman School of Public Health, Columbia University, New York, New York, USA

Zika virus (ZIKV) is implicated in fetal stillbirth, microcephaly, intracranial calcifications, and ocular anomalies following vertical transmission from infected mothers. In adults, infection may trigger autoimmune inflammatory polyneuropathy. Transmission most commonly follows the bite of infected mosquitoes but may also occur through sexual intercourse or receipt of blood products. Definitive diagnosis through detection of viral RNA is possible in serum or plasma within 10 days of disease onset, in whole blood within 3 weeks of onset, and in semen for up to 3 months. Serological diagnosis is nonetheless critical because few patients have access to molecular diagnostics during the acute phase of infection and infection may be associated with only mild or inapparent disease that does not prompt molecular testing. Serological diagnosis is confounded by cross-reactivity of immune sera with other flaviviruses endemic in the areas where ZIKV has recently emerged. Accordingly, we built a high-density microarray comprising nonredundant 12-mer peptides that tile, with one-residue overlap, the proteomes of Zika, dengue, yellow fever, West Nile, Ilheus, Oropouche, and chikungunya viruses. Serological analysis enabled discovery of a ZIKV NS2B 20-residue peptide that had high sensitivity (96.0%) and specificity (95.9%) versus natural infection with or vaccination against dengue, chikungunya, yellow fever, West Nile, tick-borne encephalitis, or Japanese encephalitis virus in a microarray assay and an enzyme-linked immunosorbent assay (ELISA) of early-convalescent-phase sera (2 to 3 weeks after onset of symptomatic infection). The emergence of Zika virus (ZIKV) as a teratogen is a profound challenge to global public health. Molecular diagnosis of infection is straightforward during the 3-week period when patients are viremic. However, serological diagnosis thereafter of historical exposure has been confounded by cross-reactivity. Using high-density peptide arrays that tile the proteomes of a selection of flaviviruses to identify a ZIKV-specific peptide, we established two assays that enable sensitive and specific diagnosis of exposure to ZIKV. These assays may be useful in guiding clinical management of mothers at risk for potential exposure to ZIKV and enable insights into the epidemiology of ZIKV infections.
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http://dx.doi.org/10.1128/mBio.00095-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5844993PMC
March 2018

BCG Vaccination Protects against Experimental Viral Infection in Humans through the Induction of Cytokines Associated with Trained Immunity.

Cell Host Microbe 2018 Jan;23(1):89-100.e5

Department of Internal Medicine and Radboud Center for Infectious Diseases, Radboud University Medical Center, Nijmegen, the Netherlands; Department for Genomics & Immunoregulation, Life and Medical Sciences Institute (LIMES), University of Bonn, 53115 Bonn, Germany. Electronic address:

The tuberculosis vaccine bacillus Calmette-Guérin (BCG) has heterologous beneficial effects against non-related infections. The basis of these effects has been poorly explored in humans. In a randomized placebo-controlled human challenge study, we found that BCG vaccination induced genome-wide epigenetic reprograming of monocytes and protected against experimental infection with an attenuated yellow fever virus vaccine strain. Epigenetic reprogramming was accompanied by functional changes indicative of trained immunity. Reduction of viremia was highly correlated with the upregulation of IL-1β, a heterologous cytokine associated with the induction of trained immunity, but not with the specific IFNγ response. The importance of IL-1β for the induction of trained immunity was validated through genetic, epigenetic, and immunological studies. In conclusion, BCG induces epigenetic reprogramming in human monocytes in vivo, followed by functional reprogramming and protection against non-related viral infections, with a key role for IL-1β as a mediator of trained immunity responses.
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http://dx.doi.org/10.1016/j.chom.2017.12.010DOI Listing
January 2018

Ebola Virus Inactivation by Detergents Is Annulled in Serum.

J Infect Dis 2017 10;216(7):859-866

Department of Viroscience.

Background: Treatment of blood samples from hemorrhagic fever virus (HFV)-infected patients with 0.1% detergents has been recommended for virus inactivation and subsequent safe laboratory testing. However, data on virus inactivation by this procedure are lacking. Here we show the effect of this procedure on diagnostic test results and infectious Ebola virus (EBOV) titers.

Methods: Serum and whole-blood samples were treated with 0.1% or 1% sodium dodecyl sulfate (SDS) or 0.1% Triton X-100 and assayed for clinical chemistry and malaria antigen detection. Infectious EBOV titers were determined in SDS-treated plasma and whole blood from EBOV-infected nonhuman primates (NHPs). Infectious titers of EBOV or herpes simplex virus type 1 (HSV-1) in detergents-treated cell culture medium containing various serum concentrations were determined.

Results: Laboratory test results were not affected by 0.1% detergent treatment of blood samples, in contrast with 1% SDS treatment. However, 0.1% detergent treatment did not inactivate EBOV in blood samples from infected NHPs. Experiments with cell culture medium showed that virus inactivation by detergents is annulled at physiological serum concentrations.

Conclusions: Treatment of blood samples with 0.1% SDS or Triton X-100 does not inactivate EBOV. Inactivation protocols for HFV should be validated with serum and whole blood.
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http://dx.doi.org/10.1093/infdis/jix401DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5853228PMC
October 2017

Cell-line dependent antiviral activity of sofosbuvir against Zika virus.

Antiviral Res 2017 Oct 11;146:161-163. Epub 2017 Sep 11.

Department of Viroscience, Unit Clinical Virology, Erasmus MC, Rotterdam, The Netherlands. Electronic address:

The recent epidemic of Zika virus (ZIKV) in the Americas and its association with fetal and neurological complications has shown the need to develop a treatment. Repurposing of drugs that are already FDA approved or in clinical development may shorten drug development timelines in case of emerging viral diseases like ZIKV. Initial studies have shown conflicting results when testing sofosbuvir developed for treatment of infections with another Flaviviridae virus, hepatitis C virus. We hypothesized that the conflicting results could be explained by differences in intracellular processing of the compound. We assessed the antiviral activity of sofosbuvir and mericitabine against ZIKV using Vero, A549, and Huh7 cells and measured the level of the active sofosbuvir metabolite by mass spectrometry. Mericitabine did not show activity, while sofosbuvir inhibited ZIKV with an IC of ∼4 μM, but only in Huh7 cells. This correlated with differences in intracellular concentration of the active triphosphate metabolite of sofosbuvir, GS-461203 or 007-TP, which was 11-342 times higher in Huh7 cells compared to Vero and A549 cells. These results show that a careful selection of cell system for repurposing trials of prodrugs is needed for evaluation of antiviral activity. Furthermore, the intracellular levels of 007-TP in tissues and cell types that support ZIKV replication in vivo should be determined to further investigate the potential of sofosbuvir as anti-ZIKV compound.
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http://dx.doi.org/10.1016/j.antiviral.2017.09.004DOI Listing
October 2017

Urban Chikungunya in the Middle East and North Africa: A systematic review.

PLoS Negl Trop Dis 2017 Jun 26;11(6):e0005707. Epub 2017 Jun 26.

Department of Healthcare Policy and Research, Weill Cornell Medicine, Cornell University, New York, New York, United States of America.

Background: The epidemiology of Chikungunya virus (CHIKV) in the Middle East and North Africa (MENA) is not well characterized despite increasing recognition of its expanding infection and disease burden in recent years.

Methodology / Principal Findings: Following Cochrane Collaboration guidelines and reporting our findings following PRISMA guidelines, we systematically reviewed records describing the human prevalence and incidence, CHIKV prevalence/infection rates in vectors, outbreaks, and reported cases for CHIKV across the MENA region. We identified 29 human seroprevalence measures, one human incidence study, one study reporting CHIKV infection rates in Aedes, and nine outbreaks and case reports/series reported in the MENA from 1970-2015. Overall, anti-CHIKV antibody or reports of autochthonous transmission were identified from 10 of 23 countries in the MENA region (Djibouti, Egypt, Iraq, Iran, Kuwait, Pakistan, Saudi Arabia, Somalia, Sudan, and Yemen), with seroprevalence measures among general populations (median 1.0%, range 0-43%) and acute febrile illness populations (median 9.8%, range 0-30%). Sudan reported the highest number of studies (n = 11) and the highest seroprevalence among general populations (median 12%, range 0-43%) and undifferentiated acute febrile illness populations (median 18%, range 10-23%). CHIKV outbreaks were reported from Djibouti, Pakistan, Sudan, and Yemen.

Conclusions / Significance: Seroprevalence studies and outbreak reports suggest endemic transmission of urban cycle CHIKV in at least the Red Sea region and Pakistan. However, indications of seroprevalence despite a low quantity of CHIKV epidemiologic research from the region suggests that CHIKV transmission is currently underrecognized.
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http://dx.doi.org/10.1371/journal.pntd.0005707DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5501693PMC
June 2017