The CARBA-MAP study: national mapping of carbapenemases in Spain (2014-2018).

Introduction: Infections caused by carbapenem-resistant Enterobacterales (CRE) and carbapenem-resistant Pseudomonas aeruginosa, including isolates producing acquired carbapenemases, constitute a prevalent health problem worldwide. The primary objective of this study was to determine the distribution of the different carbapenemases among carbapenemase-producing Enterobacterales (CPE, specifically Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae complex, and Klebsiella aerogenes) and carbapenemase-producing P. aeruginosa (CPPA) in Spain from January 2014 to December 2018. Methods: A national, retrospective, cross-sectional multicenter study was performed. The study included the first isolate per patient and year obtained from clinical samples and obtained for diagnosis of infection in hospitalized patients. A structured questionnaire was completed by the participating centers using the REDCap platform, and results were analyzed using IBM SPSS Statistics 29.0.0. Results: A total of 2,704 carbapenemase-producing microorganisms were included, for which the type of carbapenemase was determined in 2692 cases: 2280 CPE (84.7%) and 412 CPPA (15.3%), most often using molecular methods and immunochromatographic assays. Globally, the most frequent types of carbapenemase in Enterobacterales and P. aeruginosa were OXA-48-like, alone or in combination with other enzymes (1,523 cases, 66.8%) and VIM (365 cases, 88.6%), respectively. Among Enterobacterales, carbapenemase-producing K. pneumoniae was reported in 1821 cases (79.9%), followed by E. cloacae complex in 334 cases (14.6%). In Enterobacterales, KPC is mainly present in the South and South-East regions of Spain and OXA-48-like in the rest of the country. Regarding P. aeruginosa, VIM is widely distributed all over the country. Globally, an increasing percentage of OXA-48-like enzymes was observed from 2014 to 2017. KPC enzymes were more frequent in 2017-2018 compared to 2014-2016. Discussion: Data from this study help to understand the situation and evolution of the main species of CPE and CPPA in Spain, with practical implications for control and optimal treatment of infections caused by these multi-drug resistant organisms.

Case report: BA.1 subvariant showing a BA.2-like pattern using a variant-specific PCR assay due to a single point mutation downstream the spike 69/70 deletion.

BACKGROUND: SARS-CoV-2 variant tracking is key to the genomic surveillance of the COVID-19 pandemic. While next-generation sequencing (NGS) is commonly used for variant determination, it is expensive and time-consuming. Variant-specific PCR (vsPCR) is a faster, cheaper method that detects specific mutations that are considered variant-defining. These tests usually rely on specific amplification when a mutation is present or a specific melting temperature peak after amplification. CASE PRESENTATION: A discrepant result between vsPCR and NGS was found in seventeen SARS-CoV-2 samples from Galicia, Spain. A cluster of BA.1 Omicron SARS-CoV-2 variant showed a BA.2-like melting temperature pattern due to a point mutation (C21772T) downstream the deletion of the spike amino acids 69/70. As the 69/70 deletion is widely used for differentiation between BA.1 and BA.2 by vsPCR, C21772T can cause BA.1 samples to be misinterpreted as BA.2. Over a thousand BA.1 sequences in the EpiCoV database contain this mutation. CONCLUSIONS: To our knowledge, this is the first case of a point mutation causing a vsPCR algorithm to misclassify BA.1 samples as BA.2. This is an example of how mutations in the probe target area of vsPCR tests based on melting curve analysis can lead to variant misclassification. NGS confirmation of vsPCR results is relevant for the accuracy of the epidemiological surveillance. In order to overcome the possible impact of novel mutations, diagnostic tools must be constantly updated.

Clínica diferencial en niños infectados por SARS-CoV-2, trazabilidad de contactos y rentabilidad de pruebas diagnósticas: estudio observacional transversal / Differential clinic in children infected by SARS-CoV-2, traceability of contacts and cost-effectiveness of diagnostic tests: Cross-sectional observational study

INTRODUCCIÓN: Ante la posible coexistencia de la infección por el virus SARS-CoV-2 con otras infecciones estacionales, se pretende identificar síntomas diferenciales. Se ha estudiado el papel de los niños en el contagio intrafamiliar y la sensibilidad de la reacción en cadena de la polimerasa con transcriptasa inversa (RT-PCR) en un área con baja transmisión comunitaria. MATERIAL Y MÉTODOS: Estudio observacional transversal. Pacientes entre 0-15 años estudiados por técnica RT-PCR por sospecha clínica de infección por virus SARS-CoV-2 en los meses de marzo-mayo del 2020. Encuesta sobre síntomas y contactos. Determinación de anticuerpos anti-SARS-CoV-2 al menos 21 días después del test RT- PCR. RESULTADOS: Se incluyó a 126 pacientes, 33 con infección confirmada y edad media 8,4 años (IC del 95%, 6,8-10,0) superior a los no infectados. La fiebre fue el síntoma más común y con mayor sensibilidad. Las diferencias encontradas fueron una mayor frecuencia de anosmia (p = 0,029) y cefalea (p = 0,009) entre los niños infectados con una especificidad del 96,7 y el 81,5% respectivamente. No hubo diferencias en la duración de los síntomas. Un 81,8% de los infectados fue probablemente contagiado en el núcleo familiar, en un 85,2% por un progenitor que trabajaba fuera del hogar. La sensibilidad de RT-PCR fue 70,9% y su valor predictivo negativo 91,1%. CONCLUSIONES: El cuadro clínico es inespecífico y los síntomas más específicos difíciles de detectar en niños más pequeños. Los niños tuvieron un papel reducido en la transmisión intrafamiliar. La sensibilidad de la RT-PCR podría estar relacionada con una menor contagiosidad infantil tras una semana de infección

INTRODUCTION: Given the possible coexistence of infection by the SARS-CoV-2 with other seasonal infections, the aim is to identify differential symptoms. I know has studied the role of children in intrafamily contagion and the sensitivity of reverse transcriptase polymerase chain reaction (RT-PCR) in an area with low community transmission. MATERIAL AND METHODS: Cross-sectional observational study. Patients between 0-15 years studied by RT-PCR technique due to clinical suspicion of infection by SARS-CoV-2 virus in the months of March-May 2020. Survey on symptoms and contacts. Determination of Anti-SARS-CoV-2 antibodies at least 21 days after the RT-PCR test. RESULTS: 126 patients were included, 33 with confirmed infection and age mean 8.4 years (95% CI 6.8-10,5) higher than not infected. Fever was the most common and with greater sensitivity. The differences found were a greater frequency of anosmia (P = 0.029) and headache (P = .009) among children infected with a specificity of 96.7% and 81.5% respectively. There were no differences in the duration of the symptoms. 81.8% of those infected were probably infected in the nucleus 85.2% by a parent who worked outside the home. The sensitivity of RT-PCR was 70.9% and its negative predictive value 91.1%. CONCLUSIONS: The clinical picture is nonspecific and the symptoms more specific difficult to detect in younger children. Children had a reduced role in the intrafamily transmission. The sensitivity of RT-PCR could be related to a less contagiousness in children after one week of infection

Differential clinic in children infected by SARS-CoV-2, traceability of contacts and cost-effectiveness of diagnostic tests: Cross-sectional observational study.

INTRODUCTION: Given the possible coexistence of infection by the SARS-CoV-2 with other seasonal infections, the aim is to identify differential symptoms. The role of children in intrafamily contagion and the sensitivity of reverse transcriptase polymerase chain reaction (RT-PCR) in an area with low community transmission has been studied. MATERIAL AND METHODS: Cross-sectional observational study. Patients between 0-15 years studied by RT-PCR technique due to clinical suspicion of infection by SARS-CoV-2 virus in the months of March-May 2020. Survey on symptoms and contacts. Determination of Anti-SARS-CoV-2 antibodies at least 21 days after the RT-PCR test. RESULTS: 126 patients were included, 33 with confirmed infection and age mean 8.4 years (95% CI 6.8-10.5) higher than not infected. Fever was the most common and with greater sensitivity. The differences found were a greater frequency of anosmia (P = .029) and headache (P = .009) among children infected with a specificity of 96.7% and 81.5% respectively. There were no differences in the duration of the symptoms. 81.8% of those infected were probably infected in the nucleus 85.2% by a parent who worked outside the home. The sensitivity of RT-PCR was 70.9% and its negative predictive value 91.1%. CONCLUSIONS: The clinical picture is nonspecific and the symptoms more specific difficult to detect in younger children. Children had a reduced role in the intrafamily transmission. The sensitivity of RT-PCR could be related to a less contagiousness in children after one week of infection.

INTRODUCCIÓN: Ante la posible coexistencia de la infección por el virus SARS-CoV-2 con otras infecciones estacionales, se pretende identificar síntomas diferenciales. Se ha estudiado el papel de los niños en el contagio intrafamiliar y la sensibilidad de la reacción en cadena de la polimerasa con transcriptasa inversa (RT-PCR) en un área con baja transmisión comunitaria. MATERIAL Y MÉTODOS: Estudio observacional transversal. Pacientes entre 0-15 años estudiados por técnica RT-PCR por sospecha clínica de infección por virus SARS-CoV-2 en los meses de marzo-mayo del 2020. Encuesta sobre síntomas y contactos. Determinación de anticuerpos anti-SARS-CoV-2 al menos 21 días después del test RT- PCR. RESULTADOS: Se incluyó a 126 pacientes, 33 con infección confirmada y edad media 8,4 años (IC del 95%, 6,8-10,0) superior a los no infectados. La fiebre fue el síntoma más común y con mayor sensibilidad. Las diferencias encontradas fueron una mayor frecuencia de anosmia (p = 0,029) y cefalea (p = 0,009) entre los niños infectados con una especificidad del 96,7 y el 81,5% respectivamente. No hubo diferencias en la duración de los síntomas.Un 81,8% de los infectados fue probablemente contagiado en el núcleo familiar, en un 85,2% por un progenitor que trabajaba fuera del hogar. La sensibilidad de RT-PCR fue 70,9% y su valor predictivo negativo 91,1%. CONCLUSIONES: El cuadro clínico es inespecífico y los síntomas más específicos difíciles de detectar en niños más pequeños. Los niños tuvieron un papel reducido en la transmisión intrafamiliar. La sensibilidad de la RT-PCR podría estar relacionada con una menor contagiosidad infantil tras una semana de infección.

[Differential clinic in children infected by SARS-CoV-2, traceability of contacts and cost-effectiveness of diagnostic tests: Cross-sectional observational study]. / Clínica diferencial en niños infectados por SARS-CoV-2, trazabilidad de contactos y rentabilidad de pruebas diagnósticas: estudio observacional transversal.

INTRODUCTION: Given the possible coexistence of infection by the SARS-CoV-2 with other seasonal infections, the aim is to identify differential symptoms. There has been studied the role of children in intrafamily contagion and the sensitivity of reverse transcriptase polymerase chain reaction (RT-PCR) in an area with low community transmission. MATERIAL AND METHODS: Cross-sectional observational study. Patients between 0-15 years studied by RT-PCR technique due to clinical suspicion of infection by SARS-CoV-2 virus in the months of March-May 2020. Survey on symptoms and contacts. Determination of Anti-SARS-CoV-2 antibodies at least 21 days after the RT-PCR test. RESULTS: 126 patients were included, 33 with confirmed infection and mean age 8.4 years (95% CI 6.8-10,5), age higher than not infected. Fever was the most common symptom and with greater sensitivity. The differences found were a greater frequency of anosmia (P=0.029) and headache (P=.009) among children infected with a specificity of 96.7% and 81.5% respectively. There were no differences in the duration of the symptoms. 81.8% of those infected were probably infected in the family nucleus, 85.2% by a parent who worked outside the home. The sensitivity of RT-PCR was 70.9% and its negative predictive value 91.1%. CONCLUSIONS: The clinical picture is nonspecific and the more specific symptoms difficult to detect in younger children. Children had a reduced role in the intrafamily transmission. The sensitivity of RT-PCR could be related to a less contagiousness in children after one week of infection.

Pooling for SARS-CoV-2 control in care institutions.

BACKGROUND: Workers and residents in Care Homes are considered at special risk for the acquisition of SARS-CoV-2 infection, due to the infectivity and high mortality rate in the case of residents, compared to other containment areas. The role of presymptomatic people in transmission has been shown to be important and the early detection of these people is critical for the control of new outbreaks. Pooling strategies have proven to preserve SARS-CoV-2 testing resources. The aims of the present study, based in our local experience, were (a) to describe SARS-CoV-2 prevalence in institutionalized people in Galicia (Spain) during the Coronavirus pandemic and (b) to evaluate the expected performance of a pooling strategy using RT-PCR for the next rounds of screening of institutionalized people. METHODS: A total of 25,386 Nasopharyngeal swab samples from the total of the residents and workers at Care Homes in Galicia (March to May 2020) were individually tested using RT-PCR. Prevalence and quantification cycle (Cq) value distribution of positives was calculated. Besides, 26 pools of 20 samples and 14 pools of 5 samples were tested using RT-PCR as well (1 positive/pool). Pooling proof of concept was performed in two populations with 1.7 and 2% prevalence. RESULTS: Distribution of SARS-CoV-2 infection at Care Homes was uneven (0-60%). As the virus circulation global rate was low in our area (3.32%), the number of people at risk of acquiring the infection continues to be very high. In this work, we have successfully demonstrated that pooling of different groups of samples at low prevalence clusters, can be done with a small average delay on Cq values (5 and 2.85 cycles for pools of 20 and 5 samples, respectively). CONCLUSIONS: A new screening system with guaranteed protection is required for small clusters, previously covered with individual testing. Our proposal for Care Homes, once prevalence zero is achieved, would include successive rounds of testing using a pooling solution for transmission control preserving testing resources. Scale-up of this method may be of utility to confront larger clusters to avoid the viral circulation and keeping them operative.

Molecular epidemiology of an enterovirus A71 outbreak associated with severe neurological disease, Spain, 2016.

IntroductionEnterovirus A71 (EV-A71) is an emerging pathogen that causes a wide range of disorders including severe neurological manifestations. In the past 20 years, this virus has been associated with large outbreaks of hand, foot and mouth disease with neurological complications in the Asia-Pacific region, while in Europe mainly sporadic cases have been reported. In spring 2016, however, an EV-A71 outbreak associated with severe neurological cases was reported in Catalonia and spread further to other Spanish regions.AimOur objective was to investigate the epidemiology and clinical characteristics of the outbreak.MethodsWe carried out a retrospective study which included 233 EV-A71-positive samples collected during 2016 from hospitalised patients. We analysed the clinical manifestations associated with EV-A71 infections and performed phylogenetic analyses of the 3'-VP1 and 3Dpol regions from all Spanish strains and a set of EV-A71 from other countries.ResultsMost EV-A71 infections were reported in children (mean age: 2.6 years) and the highest incidence was between May and July 2016 (83%). Most isolates (218/233) were classified as subgenogroup C1 and 217 of them were grouped in one cluster phylogenetically related to a new recombinant variant strain associated with severe neurological diseases in Germany and France in 2015 and 2016. Moreover, we found a clear association of EV-A71-C1 infection with severe neurological disorders, brainstem encephalitis being the most commonly reported.ConclusionAn emerging recombinant variant of EV-A71-C1 was responsible for the large outbreak in 2016 in Spain that was associated with many severe neurological cases.

Molecular epidemiology of enterovirus and parechovirus infections according to patient age over a 4-year period in Spain.

The epidemiology and clinical association of enterovirus (EV) and parechovirus (HPeV) infections, as well as the type-distribution-according-to-age, were determined during a 4-year study period in Spain. During 2010-2013, a total of 21,832 clinical samples were screened for EV and the detection frequency was 6.5% (1,430). Of the total EV-negative samples, only 1,873 samples from 2011 to 2013 were available for HPeV testing. HPeV was detected in 42 (2%) of them. Positive samples were genotyped using PCR and sequencing. EV infections occurred in all age groups of patients: neonates (17%), children 28 days to 2 years (29%), children 2-14 years (40%), and adults (14%). Thirty-four different EV types were identified. HPeV infections were detected exclusively in infants <8 m (70% neonates, P < 0.05). All but one HPeV were HPeV-3. Differences in type frequency detection were found according to age and clinical manifestation. Coxsackievirus (CV)-B4 (61%), CV-B5 (83%), and HPeV-3 (64%) were more frequent in neonates than in older patients (P < 0.05). Echovirus (E)-3 (60%), E-18 (47%), E-25 (62%), CV-A6 (61%), CV-A16 (72%), and EV-71 (75%) were mainly detected in children 28 days to 2 years (P < 0.05), whereas, E-6 (79%), E-20 (88%), and E-30 (85%) were predominant in children >2 years and adults (P < 0.05). Clinically, meningitis was associated with EV (P < 0.01) whereas, encephalitis was more frequent in HPeV-infected patients. CV-B types were associated with myocarditis (90%; P < 0.05) and EV species A with hand-foot-mouth-disease/atypical exanthema (88%; P < 0.05). J. Med. Virol. 89:435-442, 2017. © 2016 Wiley Periodicals, Inc.