SARS-CoV-2 Infection During the First and Second Pandemic Waves in Spain: the ENE-COVID Study.

Objectives. To describe participant characteristics associated with severe acute respiratory syndrome coronavirus 2 infection in Spain's first 2 COVID-19 waves per the Spanish National Seroepidemiological Survey of SARS-CoV-2 Infection (ENE-COVID). Methods. A representative cohort of the noninstitutionalized Spanish population, selected through stratified 2-stage sampling, answered a questionnaire and received point-of-care testing April to June 2020 (first wave: n = 68 287); previously seronegative participants repeated the questionnaire and test November 2020 (second wave: n = 44 451). We estimated seropositivity by wave and participant characteristics, accounting for sampling weights, nonresponse, and design effects. Results. We found that 6.0% (95% confidence interval [CI] = 5.7%, 6.4%) of Spain's population was infected by June and 3.8% (95% CI = 3.5%, 4.1%) more by November 2020. Both genders were equally affected. Seroprevalence decreased with age in adults 20 years and older in the second wave; socioeconomic differences increased. Health care workers were affected at 11.1% (95% CI = 9.0%, 13.6%) and 6.1% (95% CI = 4.4%, 8.5%) in the first and second waves, respectively. Living with an infected person increased infection risk to 22.1% (95% CI = 18.9%, 25.6%) in the first and 35.0% (95% CI = 30.8%, 39.4%) in the second wave. Conclusions. ENE-COVID characterized the first 2 pandemic waves, when information from surveillance systems was incomplete. (Am J Public Health. 2023;113(5):533-544. https://doi.org/10.2105/AJPH.2023.307233).

Design and Implementation of a Nationwide Population-Based Longitudinal Survey of SARS-CoV-2 Infection in Spain: The ENE-COVID Study.

Data System. The Spanish National Seroepidemiological Survey of SARS-CoV-2 (or ENE-COVID; SARS-CoV-2 [severe acute respiratory syndrome coronavirus 2] is the causative agent of COVID-19) was funded by the Spanish Ministry of Health, the Instituto de Salud Carlos III, and the Spanish National Health System. Data Collection/Processing. A stratified 2-stage probability sampling was used to select a representative cohort of the noninstitutionalized population of Spain. ENE-COVID collected longitudinal data from epidemiological questionnaires and 2 SARS-CoV-2 IgG antibody tests. From April 27 to June 22, 2020, 68 287 participants (77.0% of contacted persons) received a point-of-care test and 61 095 (68.9%) also underwent a laboratory immunoassay. A second follow-up phase was conducted between November 16 and 30, 2020. Data Analysis/Dissemination. Analyses use weights to adjust for oversampling and nonresponse and account for design effects of stratification and clustering. ENE-COVID data for research purposes will be available upon request from the official study Web page. Public Health Implications. ENE-COVID, a nationwide population-based study, allowed monitoring seroprevalence of antibodies against SARS-CoV-2 at the national and regional levels, providing accurate figures by gender, age (from babies to nonagenarians), and selected risk factors; characterizing symptomatic and asymptomatic infections; and estimating the infection fatality risk during the first pandemic wave. (Am J Public Health. 2023;113(5):525-532. https://doi.org/10.2105/AJPH.2022.307167).

An interregional measles outbreak in Spain with nosocomial transmission, November 2017 to July 2018.

Given sustained high vaccination coverage and enhanced surveillance for measles, Spain has been free of endemic measles transmission since 2014, achieving elimination certification from the World Health Organization in 2017. In November 2017, measles was introduced through an imported case travelling to the Valencian Community, causing an interregional outbreak. Here, we describe the outbreak using data reported to the national epidemiological surveillance network. The outbreak involved 154 cases (67 males, 87 females) notified in four regions; 148 were laboratory-confirmed and six epidemiologically linked. Most cases were adults aged 30-39 (n = 62, 40.3%) years. Sixty-two cases were hospitalised (40.3%) and 35 presented complications (22.7%). Two thirds of the cases (n = 102) were unvaccinated including 11 infants (≤ 1 year) not yet eligible for vaccination. The main route of transmission was nosocomial; at least six healthcare facilities and 41 healthcare workers and support personnel were affected. Sequencing of the viral nucleoprotein C-terminus (N450) identified genotype B3, belonging to the circulating MVs/Dublin.IRL/8.16-variant. Control measures were implemented, and the outbreak was contained in July 2018. The outbreak highlighted that raising awareness about measles and improving the vaccination coverage in under-vaccinated subgroups and personnel of healthcare facilities are key measures for prevention of future outbreaks.

Prevalence of SARS-CoV-2 in Spain (ENE-COVID): a nationwide, population-based seroepidemiological study.

BACKGROUND: Spain is one of the European countries most affected by the COVID-19 pandemic. Serological surveys are a valuable tool to assess the extent of the epidemic, given the existence of asymptomatic cases and little access to diagnostic tests. This nationwide population-based study aims to estimate the seroprevalence of SARS-CoV-2 infection in Spain at national and regional level. METHODS: 35 883 households were selected from municipal rolls using two-stage random sampling stratified by province and municipality size, with all residents invited to participate. From April 27 to May 11, 2020, 61 075 participants (75·1% of all contacted individuals within selected households) answered a questionnaire on history of symptoms compatible with COVID-19 and risk factors, received a point-of-care antibody test, and, if agreed, donated a blood sample for additional testing with a chemiluminescent microparticle immunoassay. Prevalences of IgG antibodies were adjusted using sampling weights and post-stratification to allow for differences in non-response rates based on age group, sex, and census-tract income. Using results for both tests, we calculated a seroprevalence range maximising either specificity (positive for both tests) or sensitivity (positive for either test). FINDINGS: Seroprevalence was 5·0% (95% CI 4·7-5·4) by the point-of-care test and 4·6% (4·3-5·0) by immunoassay, with a specificity-sensitivity range of 3·7% (3·3-4·0; both tests positive) to 6·2% (5·8-6·6; either test positive), with no differences by sex and lower seroprevalence in children younger than 10 years (<3·1% by the point-of-care test). There was substantial geographical variability, with higher prevalence around Madrid (>10%) and lower in coastal areas (<3%). Seroprevalence among 195 participants with positive PCR more than 14 days before the study visit ranged from 87·6% (81·1-92·1; both tests positive) to 91·8% (86·3-95·3; either test positive). In 7273 individuals with anosmia or at least three symptoms, seroprevalence ranged from 15·3% (13·8-16·8) to 19·3% (17·7-21·0). Around a third of seropositive participants were asymptomatic, ranging from 21·9% (19·1-24·9) to 35·8% (33·1-38·5). Only 19·5% (16·3-23·2) of symptomatic participants who were seropositive by both the point-of-care test and immunoassay reported a previous PCR test. INTERPRETATION: The majority of the Spanish population is seronegative to SARS-CoV-2 infection, even in hotspot areas. Most PCR-confirmed cases have detectable antibodies, but a substantial proportion of people with symptoms compatible with COVID-19 did not have a PCR test and at least a third of infections determined by serology were asymptomatic. These results emphasise the need for maintaining public health measures to avoid a new epidemic wave. FUNDING: Spanish Ministry of Health, Institute of Health Carlos III, and Spanish National Health System.

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.

Predominance of CXCR4 tropism in HIV-1 CRF14_BG strains from newly diagnosed infections.

OBJECTIVES: R5-tropic viruses are associated with HIV-1 transmission and predominate during the early stages of infection. X4-tropic populations have been detected in ~50% of patients with late-stage disease infected with subtype B viruses. In this study, we compared the frequency of X4 tropism in individuals infected with HIV-1 CRF14_BG viruses, which have a V3 loop of subtype B, with a control group of individuals infected with subtype B viruses. METHODS: Sixty-three individuals infected with HIV-1 CRF14_BG (n = 31) or subtype B (n = 32) were studied. Similar proportions of newly diagnosed and chronically infected individuals were included in the subtype B and CRF14_BG groups. V3 sequences were obtained and coreceptor tropism was predicted using the Geno2pheno[coreceptor] algorithm. V3 net charge and 11/25 rules were also used for coreceptor prediction. RESULTS: Overall, X4 tropism was more frequent among individuals infected with CRF14_BG viruses (87.1%) than subtype B viruses (34.3%), a difference that was statistically highly significant (P = 0.00001). Importantly, the frequencies among newly diagnosed individuals were 90% and 13.3%, respectively (P = 0.0007). Characteristic amino acids in the V3 loop (T13, M14, V19 and W20) were identified at higher frequencies in CRF14_BG viruses (54%) than subtype B viruses (0%; P < 0.000001). CONCLUSIONS: CRF14_BG is the genetic form with the highest proportion of X4-tropic viruses reported to date in newly diagnosed and chronic infections. This suggests high pathogenicity for CRF14_BG viruses, potentially leading to rapid disease progression. CCR5 antagonists will be ineffective in most CRF14_BG-infected patients, even at early stages of infection.

Improvement of HIV-1 coreceptor tropism prediction by employing selected nucleotide positions of the env gene in a Bayesian network classifier.

OBJECTIVES: This study aimed to develop a genotypic method to predict HIV-1 coreceptor usage by employing the nucleotide sequence of the env gene in a tree-augmented naive Bayes (TAN) classifier, and to evaluate its accuracy in prediction compared with other available tools. METHODS: A wrapper data-mining strategy interleaved with a TAN algorithm was employed to evaluate the predictor value of every single-nucleotide position throughout the HIV-1 env gene. Based on these results, different nucleotide positions were selected to develop a TAN classifier, which was employed to predict the coreceptor tropism of all the full-length env gene sequences with information on coreceptor tropism currently available at the Los Alamos HIV Sequence Database. RESULTS: Employing 26 nucleotide positions in the TAN classifier, an accuracy of 95.6%, a specificity (identification of CCR5-tropic viruses) of 99.4% and a sensitivity (identification of CXCR4/dual-tropic viruses) of 80.5% were achieved for the in silico cross-validation. Compared with the phenotypic determination of coreceptor usage, the TAN algorithm achieved more accurate predictions than WebPSSM and Geno2pheno [coreceptor] (P<0.05). CONCLUSIONS: The use of the methodology presented in this work constitutes a robust strategy to identify genetic patterns throughout the HIV-1 env gene differently present in CCR5-tropic and CXCR4/dual-tropic viruses. Moreover, the TAN classifier can be used as a genotypic tool to predict the coreceptor usage of HIV-1 isolates reaching more accurate predictions than with other widely used genotypic tools. The use of this algorithm could improve the correct prescribing of CCR5 antagonist drugs to HIV-1-infected patients.

Identification of Besnoitia besnoiti proteins that showed differences in abundance between tachyzoite and bradyzoite stages by difference gel electrophoresis.

Bovine besnoitiosis is a chronic and debilitating disease, caused by the apicomplexan parasite Besnoitia besnoiti. Infection of cattle by B. besnoiti is governed by the tachyzoite stage, which is related to acute infection, and the bradyzoite stage gathered into macroscopic cysts located in subcutaneous tissue in the skin, mucosal membranes and sclera conjunctiva and related to persistence and chronic infection. However, the entire life cycle of this parasite and the molecular mechanisms underlying tachyzoite-to-bradyzoite conversion remain unknown. In this context, a different antigenic pattern has been observed between tachyzoite and bradyzoite extracts. Thus, to identify stage-specific proteins, a difference gel electrophoresis (DIGE) approach was used on tachyzoite and bradyzoite extracts followed by mass spectrometry (MS) analysis. A total of 130 and 132 spots were differentially expressed in bradyzoites and tachyzoites, respectively (average ratio ± 1.5, P<0.05 in t-test). Furthermore, 25 differentially expressed spots were selected and analysed by MALDI-TOF/MS. As a result, 5 up-regulated bradyzoite proteins (GAPDH, ENO1, LDH, SOD and RNA polymerase) and 5 up-regulated tachyzoite proteins (ENO2; LDH; ATP synthase; HSP70 and PDI) were identified. The present results set the basis for the identification of new proteins as drug targets. Moreover, the role of these proteins in tachyzoite-to-bradyzoite conversion and the role of the host cell environment should be a subject of further research.

Utility of MF-non coding region for measles molecular surveillance during post-elimination phase, Spain, 2017-2020.

Background: In countries entering the post-elimination phase for measles, the study of variants by sequencing of 450 nucleotides of the N gene (N450) does not always allow the tracing of chains of transmission. Indeed, between 2017 and 2020, most measles virus sequences belonged to either the MVs/Dublin.IRL/8.16 (B3-Dublin) or the MVs/Gir Somnath.IND/42.16 (D8-Gir Somnath) variants. We evaluated the additional use of a non-coding region (MF-NCR) as a tool to enhance resolution and infer case origin, chains of transmission and characterize outbreaks. Methods: We obtained 115 high-quality MF-NCR sequences from strains collected from Spanish patients infected with either B3-Dublin or D8-Gir Somnath variants between 2017 and 2020, performed epidemiological, phylogenetic and phylodynamic analyses and applied a mathematical model to determine relatedness among identified clades. Results: Applying this model allowed us to identify phylogenetic clades potentially derived from concomitant importations of the virus rather than single chain of transmission, inferred based on only N450 and epidemiology data. In a third outbreak, we found two related clades that corresponded to two chains of transmission. Discussion: Our results show the ability of the proposed method to improve identification of simultaneous importations in the same region which could trigger enhanced contact tracing. Moreover, the identification of further transmission chains indicates that the size of import-related outbreaks was smaller than previously found, supporting the interpretation that endemic measles transmission was absent in Spain between 2017 and 2020. We suggest considering the use of the MF-NCR region in conjunction with the study of N450 variants in future WHO recommendations for measles surveillance.

Comparison of circulation patterns of mumps virus in the Netherlands and Spain (2015-2020).

Background: Mumps is a viral infection mainly characterized by inflammation of the parotid glands. Despite of vaccination programs, infections among fully vaccinated populations were reported. The World Health Organization (WHO) recommends molecular surveillance of mumps based on sequencing of the small hydrophobic (SH) gene. The use of hypervariable non-coding regions (NCR) as additional molecular markers was proposed in multiple studies. Circulation of mumps virus (MuV) genotypes and variants in different European countries were described in the literature. From 2010 to 2020, mumps outbreaks caused by genotype G were described. However, this issue has not been analyzed from a wider geographical perspective. In the present study, sequence data from MuV detected in Spain and in The Netherlands during a period of 5 years (2015- March 2020) were analyzed to gain insights in the spatiotemporal spread of MuV at a larger geographical scale than in previous local studies. Methods: A total of 1,121 SH and 262 NCR between the Matrix and Fusion protein genes (MF-NCR) sequences from both countries were included in this study. Analysis of SH revealed 106 different haplotypes (set of identical sequences). Results: Of them, seven showing extensive circulation were considered variants. All seven were detected in both countries in coincident temporal periods. A single MF-NCR haplotype was detected in 156 sequences (59.3% of total), and was shared by five of the seven SH variants, as well as three minor MF-NCR haplotypes. All SH variants and MF-NCR haplotypes shared by both countries were detected first in Spain. Discussion: Our results suggest a transmission way from south to north Europe. The higher incidence rate of mumps in Spain in spite of similar immunization coverage in both countries, could be associated with higher risk of MuV exportation. In conclusion, the present study provided novel insights into the circulation of MuV variants and haplotypes beyond the borders of single countries. In fact, the use of MF-NCR molecular tool allowed to reveal MuV transmission flows between The Netherlands and Spain. Similar studies including other (European) countries are needed to provide a broader view of the data presented in this study.

Investigating Local Patterns of Mumps Virus Circulation, Using a Combination of Molecular Tools.

Mumps is a vaccine-preventable disease caused by the mumps virus (MuV). However, MuV has re-emerged in many countries with high vaccine coverage. The World Health Organization (WHO) recommends molecular surveillance based on sequencing of the small hydrophobic (SH) gene. Additionally, the combined use of SH and non-coding regions (NCR) has been described in different studies, proving to be a useful complement marker to discriminate general patterns of circulation at national and international levels. The aim of this work is to test local-level usefulness of the combination of SH and MF-NCR sequencing in tracing hidden transmission clusters and chains during the last epidemic wave (2015-2020) in Spain. A database with 903 cases from the Autonomous Community of Madrid was generated by the integration of microbiological and epidemiological data. Of these, 453 representative cases were genotyped. Eight different SH variants and thirty-four SH haplotypes were detected. Local MuV circulation showed the same temporal pattern previously described at a national level. Only two of the thirteen previously identified outbreaks were caused by more than one variant/haplotype. Geographical representation of SH variants allowed the identification of several previously undetected clusters, which were analysed phylogenetically by the combination of SH and MF-NCR, in a total of 90 cases. MF-NCR was not able to improve the discrimination of geographical clusters based on SH sequencing, showing limited resolution for outbreak investigations.

Evaluation of genotypic tropism prediction tests compared with in vitro co-receptor usage in HIV-1 primary isolates of diverse subtypes.

OBJECTIVES: To evaluate the sensitivity and specificity of genotypic methods for predicting the co-receptor usage of subtypes B and non-B HIV-1 primary isolates, using as gold standard the infectivity of each primary isolate in GHOST cells stably expressing HIV-1 co-receptors. METHODS: Primary isolates were obtained by co-culturing either patient's peripheral blood mononuclear cells (PBMCs) or ultracentrifuged plasma with donor-activated PBMCs. In vitro co-receptor usage was determined by infecting GHOST cells. Tropism prediction, based on V3 sequences, was determined with simple rules and bioinformatic tools (Geno2pheno[coreceptor] and WebPSSM). RESULTS: This study includes 102 HIV-1 primary isolates; 23 (22.5%) subtype B and 79 (77.5%) non-B genetic forms. V3 sequences were classified into six subtypes (A-G), although 32 (31.4%) were circulating recombinant forms and 21 (20.6%) were unique recombinant forms. Sixty-nine isolates were R5, 27 R5X4 and 6 X4. The highest levels of sensitivity and specificity for the detection of X4 strains among V3 sequences, between 91% and 100%, were obtained by using PSSM(x4r5), PSSM(si/nsi) and the 11/25 rule for sequences of subtypes A, B and G, but not for subtype F. Establishing the recommended cut-off for clinical settings of a 10% false positive rate for Geno2pheno, we obtained 93% specificity and 97% sensitivity. CONCLUSIONS: Comparing genotypic assays for HIV-1 co-receptor use with a cell-culture phenotypic assay could provide more reliable results of sensitivity and specificity for the detection of X4 strains than comparing them with recombinant assays, considered as gold standard. In general, except for subtype F isolates, there is a good correlation for tropism prediction.

Mind your Ps: A probabilistic model to aid the interpretation of molecular epidemiology data.

BACKGROUND: Assessing relatedness of pathogen sequences in clinical samples is a core goal in molecular epidemiology. Tools for Bayesian analysis of phylogeny, such as the BEAST software package, have been typically used in the analysis of sequence/time data in public health. However, they are computationally-, time-, and knowledge-intensive, demanding resources that many laboratories do not have available or cannot allocate frequently. METHODS: To evaluate a faster and simpler alternative method to support the routine interpretation of sequence data for epidemiology, we obtained sequences for two regions in the measles virus genome, N-450 and MF-NCR, from patient samples of genotypes B3, D4 and D8 taken between 2011 and 2017 in the UK and Romania. A mathematical model incorporating time, possible shared ancestry and the Poisson distribution describing the number of expected substitutions at a given time point was developed to exclude epidemiological relatedness between pairs of sequences. The model was validated against the commonly used Bayesian phylogenetic method using an independent dataset collected in 2017-19. FINDINGS: We demonstrate that our model, using time and sequence information to predict whether two samples may be related within a given time frame, minimises the risk of erroneous exclusion of relatedness. An easy-to-use implementation in the form of a guide and spreadsheet is provided for convenient application. INTERPRETATION: The proposed model only requires a previously calculated substitution rate for the locus and pathogen of interest. It allows for an informed but quick decision on the likelihood of relatedness between two samples within a time frame, without the need for phylogenetic reconstruction, thus facilitating rapid epidemiological interpretation of sequence data. FUNDING: This work was funded by the United Kingdom Health Security Agency (UKHSA). The World Health Organization European Regional Office funded Aurora Fernández-García and Mihaela Lazar training visits to UKHSA.

Measles outbreak in the sanitary area of Guadalajara (Spain): difficulty in microbiological diagnosis in the era of its elimination.

INTRODUCTION: In Spain, like in other countries where endemic measles has been eliminated, there is a need for available diagnostic tolos for confirming any cases in order to prevent and control its transmission. We describe the different microbiological tests used for the diagnosis of measles during an outbreak that occurred in 2019 in the province of Guadalajara (Spain). METHODS: Serological and molecular tests were performed at the Microbiology laboratory of the Guadalajara University Hospital and at the National Center for Microbiology of the Carlos III Health Institute (Majadahonda, Spain). Patient data were obtained from the surveillance system. RESULTS: A total of 43 patients had a laboratory diagnosis of measles: 29 cases by PCR (pharyngeal exudate or urine) and positive specific IgM, 11 cases by PCR, and 3 cases only by a positive IgM. Genotype D8 was identified in 35 confirmed cases and genotype A in two that were discarded as post-vaccination cases. PCR was positive in the acute sera of 11 out of 14 patients with a negative IgM. Eleven confirmed cases had recieved one or two vaccine doses. Twelve adult patients were hospitalizated, all of them with a diagnostic of hepatitis. CONCLUSIONS: The combination of molecular tests and the presence of specific IgM is necessary for a correct diagnosis of measles and also to classify patients with a breakthrough infection or vaccine failures (primary or secondary). Genotyping is essential for the correct classification of the patients in the context of a measles elimination program.

Evolution of antibodies against SARS-CoV-2 over seven months: Experience of the nationwide seroprevalence ENE-COVID study in Spain.

BACKGROUND: The main aims of this study were to analyze trends of SARS-CoV-2 anti-nucleocapsid IgG throughout the four rounds of the seroepidemiologic study ENE-COVID, and compare the fourth-round results of two immunoassays detecting anti-nucleocapsid and anti-RBD IgG. METHODS: ENE-COVID was developed in 2020 (two phases). Phase one included three rounds carried out in April 27-May 11, May 18-June 1, and June 8-June 22. Phase two included a fourth round in the same cohort (November 16-29). A chemiluminescent microparticle immunoassay was offered to participants in the first three rounds (Abbott; anti-nucleocapsid IgG). In the fourth round, we offered this test and a chemiluminescence immunoassay (Beckman; anti-RBD IgG) to i) a randomly selected sub-cohort, ii) participants who were IgG-positive in any of the three first rounds; and iii) participants who were IgG-positive in the fourth round by point-of-care immunochromatography. RESULTS: 10,153 individuals (82.2% of people invited) participated in the fourth round. Of them, 2595 (35.1% of participants with results in the four rounds) were positive for anti-nucleocapsid IgG in at least one round. Anti-nucleocapsid IgG became undetectable in 43.3% of participants with positive first-round results. In fourth round, anti-nucleocapsid and anti-RBD IgG were detected in 5.5% (321/5827) and 5.4% (315/5827) participants of the randomly selected sub-cohort, and in 26.6% (867/3261) and 25.9% (846/3261) participants with at least one previous positive result, respectively. CONCLUSIONS: The IgG response is heterogeneous and conditioned by infection severity. A proportion of SARS-CoV-2 infected population may have negative serologic results in the post-infection months.

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).

Mumps virus outbreak related to a water pipe (narghile) shared smoking.

INTRODUCTION: This study describes a mumps outbreak among a group of young people who shared a same narghile to smoking. Saliva and blood samples were obtained from 3 cases for RT-PCR and serology respectively. METHODS: The notification of a mumps case started an epidemiological investigation. Information of other 6 additional symptomatic persons who had gathered with the case in a discotheque where they smoking in a same narghile was achieved. RT-PCR positive samples were genotyped by sequencing. RESULTS: The 7 patients resided in 3 different municipalities, and they do not have get together for more than a month until the meeting in the discotheque. Four cases were confirmed by RT-PCR and/or IgM determinations. The genomic investigation showed identical nucleic sequences. CONCLUSIONS: This outbreak is consequence of the common use of a narghile to smoking. The public usage of these water pipes should be regulated.

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.

Measles outbreak in the sanitary area of Guadalajara (Spain): Difficulty in microbiological diagnosis in the era of its elimination. / Brote de sarampión en el área sanitaria de Guadalajara (España): dificultad en el diagnóstico microbiológico en la era de su eliminación.

INTRODUCTION: In Spain, like in other countries where endemic measles has been eliminated, there is a need for available diagnostic tools for confirming any cases in order to prevent and control its transmission. We describe the different microbiological tests used for the diagnosis of measles during an outbreak that occurred in 2019 in the province of Guadalajara (Spain). METHODS: Serological and molecular tests were performed at the Microbiology laboratory of the Guadalajara University Hospital and at the National Center for Microbiology of the Carlos III Health Institute (Majadahonda, Spain). Patient data were obtained from the surveillance system. RESULTS: A total of 43 patients had a laboratory diagnosis of measles: 29 cases by PCR (pharyngeal exudate or urine) and positive specific IgM, 11 cases by PCR, and 3 cases only by a positive IgM. Genotype D8 was identified in 35 confirmed cases and genotype A in 2 that were discarded as post-vaccination cases. PCR was positive in the acute sera of 11 out of 14 patients with a negative IgM. Eleven confirmed cases had recieved one or 2 vaccine doses. Twelve adult patients were hospitalizated, all of them with a diagnostic of hepatitis. CONCLUSIONS: The combination of molecular tests and the presence of specific IgG and IgM are necessary for a correct diagnosis of measles and also to classify patients with a breakthrough infection or vaccine failures (primary or secondary). Genotyping is essential for the correct classification of the patients in the context of a measles elimination program.

ENE-COVID nationwide serosurvey served to characterize asymptomatic infections and to develop a symptom-based risk score to predict COVID-19.

OBJECTIVES: To characterize asymptomatic SARS-CoV-2 infections and develop a symptom-based risk score useful in primary healthcare. STUDY DESIGN AND SETTING: Sixty-one thousand ninty-two community-dwelling participants in a nationwide population-based serosurvey completed a questionnaire on COVID-19 symptoms and received an immunoassay for SARS-CoV-2 IgG antibodies between April 27 and June 22, 2020. Standardized prevalence ratios for asymptomatic infection were estimated across participant characteristics. We constructed a symptom-based risk score and evaluated its ability to predict SARS-CoV-2 infection. RESULTS: Of all, 28.7% of infections were asymptomatic (95% CI 26.1-31.4%). Standardized asymptomatic prevalence ratios were 1.19 (1.02-1.40) for men vs. women, 1.82 (1.33-2.50) and 1.45 (0.96-2.18) for individuals <20 and ≥80 years vs. those aged 40-59, 1.27 (1.03-1.55) for smokers vs. nonsmokers, and 1.91 (1.59-2.29) for individuals without vs. with case contact. In symptomatic population, a symptom-based score (weights: severe tiredness = 1; absence of sore throat = 1; fever = 2; anosmia/ageusia = 5) reached standardized seroprevalence ratio of 8.71 (7.37-10.3), discrimination index of 0.79 (0.77-0.81), and sensitivity and specificity of 71.4% (68.1-74.4%) and 74.2% (73.1-75.2%) for a score ≥3. CONCLUSION: The presence of anosmia/ageusia, fever with severe tiredness, or fever without sore throat should serve to suspect COVID-19 in areas with active viral circulation. The proportion of asymptomatics in children and adolescents challenges infection control.