Fatal Case of Crimean-Congo Hemorrhagic Fever Caused by Reassortant Virus, Spain, 2018.

In August 2018, a fatal autochthonous case of Crimean-Congo hemorrhagic fever was confirmed in western Spain. The complete sequence of the viral genome revealed circulation of a new virus because the genotype differs from that of the virus responsible for another case in 2016. Practitioners should be alert to possible new cases.

Hantavirus Pulmonary Syndrome in Traveler Returning from Nepal to Spain.

Most human hantavirus infections occur in Asia, but some cases have been described in Europe in travelers returning from Asia. We describe a case of hantavirus pulmonary syndrome in a previously healthy traveler occurring shortly after he returned to Spain from Nepal. Serologic tests suggested a Puumala virus-like infection.

Comparison of commercial methods of immunoblot, ELISA, and chemiluminescent immunoassay for detecting type-specific herpes simplex viruses-1 and -2 IgG.

BACKGROUND: Serology for type-specific herpes simplex virus (HSV) is based on the use of the respective glycoprotein G (gG). METHODS: Chemiluminescent immunoassay (CLIA; BIO-FLASH® , Biokit, Spain), ELISA (HerpeSelect® , Focus, USA), and immunoblot (IB; Virotech, Germany) for detecting HSV-1- and HSV-2-specific IgG were compared using 384 serum samples received for HSV serology. The samples were classified as positive or negative according to a consensus criterion. RESULTS: For HSV-1, 262 samples were positive and 118 were negative (four samples were unclassifiable). IB showed agreement, sensitivity, and specificity values of 98.68%, 98.47% and 99.15%, respectively. The corresponding figures for CLIA and ELISA were 98.95%, 99.24% and 98.31%, and 98.16%, 99.62% and 94.92%, respectively. For HSV-2, 106 samples were positive and 278 were negative. Agreement, sensitivity, and specificity of IB were 99.48%, 98.11%, and 100%, respectively. The corresponding figures for CLIA and ELISA were 99.48%, 99.06% and 99.64%, and 98.18%, 99.06% and 97.84%, respectively. CONCLUSION: The three methods showed excellent and equivalent performance characteristics for the detection of type-specific IgG to HSV-1 and HSV-2.

Genomic non-coding regions reveal hidden patterns of mumps virus circulation in Spain, 2005 to 2015.

BackgroundSince mumps vaccination was introduced in 1981 in Spain, the incidence of the disease has dropped significantly. However, cyclic epidemic waves and outbreaks still occur, despite high vaccination coverage. The World Health Organization (WHO) recommends genotyping to trace the pattern of mumps virus (MuV) circulation. Genotype H was predominant in Spain, but was replaced in 2005 by genotype G which has subsequently remained dominant. Of the small hydrophobic protein gene sequences, 78% are identical and belong to the MuVi/ Sheffield.GBR.1.05/[G]-variant. Aim: Our study aimed to investigate whether the circulation of MuV strains in Spain was continuous after the emergence of genotype G in 2005. Method: We obtained 46 samples from Spanish patients infected with MuVi/Sheffield.GBR.1.05/[G] during two epidemic waves and analysed them using new molecular markers based on genomic non-coding regions (NCRs) that discriminate subvariants of this virus strain. Results: Phylogenetic analyses of the nucleoprotein-phosphoprotein and matrix protein-fusion protein NCR indicated strain replacement after a drop in incidence in 2009, which had not been detectable by SH sequencing. Clustering of sequences from patients epidemiologically linked in the same outbreak suggests a potential use for these NCRs in outbreak characterisation. Conclusion: We suggest to consider their use in conjunction with the SH gene in the future WHO recommendations for MuV epidemiological surveillance.

Zika Virus Screening among Spanish Team Members After 2016 Rio de Janeiro, Brazil, Olympic Games.

We evaluated the risk for the Spanish Olympic Team acquiring Zika virus in Rio de Janeiro, Brazil, during 2016. We recruited 117 team members, and all tested negative for Zika virus. Lack of cases in this cohort supports the minimum risk estimates made before the Games.

Rapid diagnosis in serology. / Diagnóstico rápido en serología.

Serological diagnosis of acute phase infections implies the detection of IgM specific response, an effective marker of primary infection, but with less clinical significance in reactivations or reinfections. The aim of this article is to provide an updated view of the rapid diagnosis in serology by detecting the IgM isotype and reviewing its applications and limitations. Point-of-care (PoC) tests are analyzed. PoC tests are used in geographical areas where traditional tests are not available, as well as in other circumstances where their use brings the diagnosis directly to the target population. Likewise, their use reduces the response time between taking the sample and the diagnosis, making it easier to make clinical decisions. PoC assays have proven cost-effective, especially in preventing vertical transmission of syphilis and HIV infection.

Chikungunya virus infections among travellers returning to Spain, 2008 to 2014.

Since the first documented autochthonous transmission of chikungunya virus in the Caribbean island of Saint Martin in 2013, the infection has been reported within the Caribbean region as well as North, Central and South America. The risk of autochthonous transmission of chikungunya virus becoming established in Spain may be elevated due to the large numbers of travellers returning to Spain from countries affected by the 2013 epidemic in the Caribbean and South America, as well as the existence of the Aedes albopictus vector in certain parts of Spain. We retrospectively analysed the laboratory diagnostic database of the National Centre for Microbiology, Institute of Health Carlos III (CNM-ISCIII) from 2008 to 2014. During the study period, 264 confirmed cases, of 1,371 suspected cases, were diagnosed at the CNM-ISCIII. In 2014 alone, there were 234 confirmed cases. The highest number of confirmed cases were reported from the Dominican Republic (n = 136), Venezuela (n = 30) and Haiti (n = 11). Six cases were viraemic in areas of Spain where the vector is present. This report highlights the need for integrated active case and vector surveillance in Spain and other parts of Europe where chikungunya virus may be introduced by returning travellers.

First case of imported Zika virus infection in Spain.

We report a case of Zika virus (ZIKV) infection in a patient with diarrhea, fever, synovitis, non-purulent conjunctivitis, and with discreet retro-orbital pain, after returning from Colombia in January 2016. The patient referred several mosquito bites. Presence of ZIKV was detected by PCR (polymerase chain reaction) in plasma. Rapid microbiological diagnosis of ZIKV infection is needed in European countries with circulation of its vector, in order to avoid autochthonous circulation. The recent association of ZIKV infection with abortion and microcephaly, and a Guillain-Barré syndrome highlights the need for laboratory differentiation of ZIKV from other virus infection. Women with potential risk for Zika virus infection who are pregnant or planning to become pregnant must mention that fact during prenatal visits in order to be evaluated and properly monitored.

International external quality control assessment for the serological diagnosis of dengue infections.

BACKGROUND: Dengue is endemic to the tropics and subtropics, and the most frequent of arthropod-borne viral diseases. Reliable diagnosis of dengue infection is important not only in clinical care but also in disease surveillance, the control of outbreaks, and the development of new vaccines. The diagnosis of dengue infection is usually established by a variety of commercial or in-house serological protocols. The European Network for the Diagnostics of Imported Viral Diseases (ENIVD) recognized the need to survey the accuracy of dengue serological diagnostics in current use, and organized an external quality assurance (EQA) study of dengue serological practice in diagnostic laboratories. METHODS: A 15-sample panel, consisting of sera reactive against dengue plus specificity and negative controls, was sent to 48 laboratories for serological testing. The results returned by the participating laboratories were anonymized, scored, and subjected to comparison and statistical analysis. RESULTS: Ten laboratories rated all samples correctly with regard to IgM, and only three achieved the full score for IgG detection. The main handicaps in assay performance were suboptimal sensitivity of in-house IgM detection protocols by comparison with better-performing commercial ELISA tests, and the presence of IgG cross-reactivity with heterologous flaviviruses. Differences of detail in the methodology of dengue IgG antibody detection appear to underlie the disparities in accuracy observed between laboratories. CONCLUSION: This EQA study demonstrates that there is room for many laboratories to improve sensitivity in the detection of anti-dengue virus IgM antibodies, against the benchmark set by commercial antibody capture ELISA tests. The EQA shows also that cross-reactivity is a continuing issue, and IgG detection protocols must be optimized to increase their specificity.

Application of a commercial immunoblot to define EBV IgG seroprofiles.

BACKGROUND: Immunoblot (IB) techniques using different Epstein-Barr virus (EBV) antigens have been applied for detecting specific antibodies, making possible to obtain EBV seroprofiles in a single determination. The aim of this study was to evaluate a commercial IB for the detection of EBV-specific IgG (Euroimmun, Lübeck, Germany). METHODS: A total of 117 samples classified as EBV primary recent infections (n = 70), past infections (n = 29), or not infected (n = 18) have been used. The samples were characterized by immunofluorescence, by testing EBV capsid antigens IgM and IgG (using indirect approaches) and EBV nuclear antigen (by anticomplement technique; Meridian Bioscience Inc.). RESULTS: Using the cut-off value as defined by the IB manufacturer, the concordance, relative sensitivity, and relative specificity were 85.5 (100/117), 94.3% (66/70), and 72.3% (34/47), respectively. If a corrected cut-off value was considered to classify the samples, the corresponding corrected figures were 89.7, 88.6, and 91.5%, respectively. CONCLUSION: Being a useful serological diagnostic tool, IB for testing EBV IgG seems to be an adequate approach to define EBV seroprofiles. However, efforts to better define the cut-off value should be made in order to improve the performance of the assay in evaluation.

[Performance of the VIDAS automated immunoassay for the determination of Epstein-Barr virus serological status]. / Evaluación del sistema VIDAS para estudio de marcadores serológicos de infección por el virus Epstein Barr.

Enzyme linked fluorescent assays (VIDAS EBV VCA IgM, VIDAS EBV VCA/EA IgG and VIDAS EBV EBNA IgG (Biomérieux, France) were evaluated to determine markers for infection of Epstein Barr virus, as well as to establish antibody profiles, compared with immunofluorescence assays as reference. The assays evaluated showed good values for sensitivity, specificity and agreement, making them useful for their application in clinical laboratories.

Seroprevalence of antibodies to the influenza A (H1N1) virus among healthcare workers prior to the 2009 pandemic peak.

OBJECTIVE: Our aim was to study the proportion of healthcare workers with a positive serology for Influenza A(H1N1)2009 without having flu, in a Spanish hospital at the beginning of the pandemic. METHODS: A survey study carried out during August 2009 (before the peak of the pandemic in Spain) in the Hospital Costa del Sol, a second level hospital with almost 300 beds in the South of Spain. The participants were workers in the following hospital units: Emergencies, Medical Area (Internal Medicine, Chest Diseases), Surgical Area (General Surgery and Anaesthesia) of any professional category. A study was made of the proportion of healthcare workers in our hospital with positive serology for the new influenza A (H1N1)2009 virus, as determined by the haemagglutination inhibition technique (≥1/40). The subjects completed a health status questionnaire, and provided a blood sample for serology testing. RESULTS: A total of 239 workers participated, of whom 25.1% had positive serology. The hospital area in which most individuals had positive serology was the Emergency Department (36.6%), while the professional category in which most individuals with a positive serology worked was that of the orderlies (41.7%). CONCLUSION: Around 25% of healthcare workers in our hospital had positive serology before the peak of the pandemic, none of them had received vaccine for Influenza A (H1N1) 2009 or had been diagnosed of influenza previously.

Comparative evaluation of assays for IgM detection of rubella and measles infections.

BACKGROUND: Serological diagnosis of infections due to measles and rubella viruses is done by IgM detection. The aim of this study was to comparatively evaluate commercial systems for detecting IgM against both viruses, including those of ELISA, in indirect and capture formats, chemiluminescence and electrochemiluminescence. METHODS: Seven (for rubella) and six (for measles) assays were studied. One hundred and sixty two samples were included in the study (from 90 rubella and 72 measles cases), and all were analyzed in all the assays. RESULTS: The ranges of sensitivity, specificity and agreement for rubella were 94.8-100%, 52.4-100% and 75.5-98.1%, respectively. The corresponding ranges for measles assays were 87.0-100%, 53.3-100%, and 73.0-99.4%. CONCLUSION: The best-performing assays were chemiluminescence (for measles and rubella IgM), and electrochemiluminescence (for rubella IgM).

Myocarditis caused by human parainfluenza virus in an immunocompetent child initially associated with 2009 influenza A (H1N1) virus.

The association between respiratory viruses and myocarditis has hardly ever been described. We report a case of acute myocarditis in an immunocompetent child associated with the presence of parainfluenza virus type 3 infection, in a context of recent influenza illness, confirmed by molecular and serological studies.

Measles in Vaccinated People: Epidemiology and Challenges in Surveillance and Diagnosis in the Post-Elimination Phase. Spain, 2014-2020.

The MMR vaccination program was introduced in Spain in 1981. Consistently high vaccination coverage has led to Spain being declared free of endemic measles transmission since 2014. A few imported and import-related cases were reported during the post-elimination phase (2014 to 2020), with very low incidence: three cases per million of inhabitants a year, 70% in adults. In the post-elimination phase an increasing proportion of measles appeared in two-dose vaccinated individuals (up to 14%), posing a challenge to surveillance and laboratory investigations. Severity and clinical presentation were milder among the vaccinated. The IgM response varied and the viral load decreased, making the virus more difficult to detect. A valid set of samples (serum, urine and throat swab) is strongly recommended for accurate case classification. One third of measles in fully vaccinated people was contracted in healthcare settings, mainly in doctors and nurses, consistent with the important role of high intensity exposure in measles breakthrough cases. Surveillance protocols and laboratory algorithms should be adapted in advanced elimination settings. Reinforcing the immunity of people working in high exposure environments, such as healthcare settings, and implementing additional infection control measures, such as masking and social distancing, are becoming crucial for the global aim of measles eradication.

Phlebovirus-associated diseases transmitted by phlebotominae in Spain: Are we at risk?

The genera Phlebovirus transmitted by Diptera belonging to the Psychodidae family are a cause of self-limited febrile syndrome in the Mediterranean basin in summer and autumn. Toscana virus can also cause meningitis and meningoencephalitis. In Spain, Toscana, Granada, Naples, Sicily, Arbia and Arrabida-like viruses have been detected. The almost widespread distribution of Phlebotomus genus vectors, and especially Phlebotomus perniciosus, in which several of these viruses have been detected, makes it very likely that there will be regular human infections in our country, with this risk considered moderate for Toscana virus and low for the other ones, in areas with the highest vector activity. Most of the infections would be undiagnosed, while only Toscana virus would have a greater impact due to the potential severity of the illness.

Zika virus infection in pregnant travellers and impact on childhood neurodevelopment in the first two years of life: A prospective observational study.

BACKGROUND: The emergence of Zika virus (ZIKV) represents a threat with consequences on maternal and children's health. We aimed to assess the clinical and epidemiological characteristics of pregnant women returning from ZIKV affected areas, and the effects of maternal ZIKV infection on birth outcomes and children's health. METHODS: This was a hospital-based prospective observational study conducted at the Hospital Clínic of Barcelona and Hospital Sant Joan de Déu, Barcelona, Spain, from January 2016 to February 2020. RESULTS: One hundred and ninety-five pregnant women who had travelled to ZIKV affected areas during pregnancy were recruited. Four women (2.1%) had a confirmed ZIKV infection, 40 women (20.5%) a probable infection, and 151 (77.4%) were negative for ZIKV. Among the ZIKV confirmed cases, a pregnant woman suffered a miscarriage, highly plausible to be associated with ZIKV infection. Brain cysts and microcalcifications were detected in 7% of fetuses or infants from women with confirmed or probable ZIKV infection. Neurodevelopmental delay in the language function was found in 33.3% out of the 21 children evaluated. CONCLUSIONS: These findings contribute to the understanding of ZIKV prevalence estimates, and the impact of maternal ZIKV infection on pregnancy outcomes and children's health. Results highlight the importance of long-term surveillance in pregnant travellers and their children.

[Recommendations for implementing the UNE-EN-ISO 15189 quality standard in the clinical microbiology laboratory: bacteriology and serology]. / Recomendaciones para la implantación de la normativa de calidad UNE-EN-ISO 15189 en el laboratorio de microbiología clínica: bacteriología y serología.

The UNE-EN-ISO 15189:2007 standard specifies the management and technical requirements that clinical microbiology laboratories must meet to achieve optimal quality when performing microbiological analyses. With implementation of this standard, a laboratory can receive the accreditation and formal recognition of an authorized body, certifying that it is apt for performing an assay or group of assays. In Spain, laboratories that apply these standards can be accredited by the Entidad Nacional de Acreditación (ENAC, Spanish accreditation body). The purpose of this review is to familiarize clinical microbiology laboratory specialists with the UNE-EN-ISO 15189:2007 standard through a practical approach focussed on bacteriology and serology studies. We briefly define the scope and specify the requisites required for managing the quality of the procedures and processes involved in performing tests on human specimens, for document control, and for management of instruments and equipment, personnel, information systems, supply systems, and external services. Lastly, evaluation approaches are indicated to achieve continuing improvement of the processes carried out and the services the laboratory provides.

Evaluation of the LIAISON XL Zika Capture IgM II for the Diagnosis of Zika Virus Infections.

The aim of this study is to evaluate the performance characteristics of the LIAISON XL Zika Capture IgM II. For this purpose we tested 128 samples obtained from recent infections caused by the Zika (ZIKV; 74 samples), dengue (DENV; 10 samples), chikungunya (CHIK V; 11 samples), rubella (RUBV; 10 samples) and measles (MeV; 10 samples) viruses, as well as human parvovirus B19 (HPVB19; 13 samples). The results of the assay under evaluation are compared with those obtained from an indirect immunofluorescence (IIF) assay, and the discrepancies are resolved by considering other laboratory results (PCR and a plaque-reduction neutralization test). The LIAISON showed excellent sensitivity (100%). The specificity (91.25%) was hampered by some false-positive results in recent dengue virus, chikungunya virus, measles virus and human parvovirus B19 infections. The method evaluated is adequate, but the low specificity makes it necessary to consider the clinical and epidemiological contexts of patients, as well as other laboratory results.

Imported Human West Nile Virus Lineage 2 Infection in Spain: Neurological and Gastrointestinal Complications.

We report the first human case of West Nile virus (WNV) lineage 2 infection imported to Spain by a traveler returning from Romania. Serum, cerebrospinal fluid and urine samples were analyzed and West Nile virus infection was identified by PCR and serological tests. The patient developed fever, diarrhea and neurological symptoms, accompanied by mild pancreatitis, described previously in very few cases as a complication of WNV infection and by alithiasic cholecystitis. Viral RNA was detected in urine until 30 days after the onset of symptoms and neutralizing antibodies were detected at very low titers. The phylogenetic analysis in a fragment of the NS5 gene of the virus showed a homology with sequences from WNV lineage 2 belonging to the monophyletic Central/Southern European group.