Plasma C24:0- and C26:0-lysophosphatidylcholines are reliable biomarkers for the diagnosis of peroxisomal ß-oxidation disorders.

The gold-standard diagnostic test for peroxisomal disorders (PDs) is plasma concentration analysis of very long-chain fatty acids (VLCFAs). However, this method's time-consuming nature and limitations in cases which present normal VLCFA levels necessitates alternative approaches. The analysis of C26:0-lysophosphatydylcholine (C26:0-LPC) in dried blood spot samples by tandem-mass spectrometry (MS/MS) has successfully been implemented in certain newborn screening programs to diagnose X-linked adrenoleukodystrophy (ALD). However, the diagnostic potential of very long-chain LPCs concentrations in plasma remains poorly understood. This study sought to evaluate the diagnostic performance of C26:0-LPC and other very long-chain LPCs, comparing them to VLCFA analysis in plasma. The study, which included 330 individuals affected by a peroxisomal ß-oxidation deficiency and 407 control individuals, revealed that C26:0- and C24:0-LPC concentrations demonstrated the highest diagnostic accuracy (98.8% and 98.4%, respectively), outperforming VLCFA when C26:0/C22:0 and C24:0/C22:0 ratios were combined (98.1%). Combining C24:0- and C26:0-LPC gave the highest sensitivity (99.7%), with ALD females exhibiting notably higher sensitivity compared with the VLCFA ratio combination (98.7% vs. 93.5%, respectively). In contrast, C22:0-LPC exhibited suboptimal performance, primarily due to its low sensitivity (75%), but we identified a potential use to help distinguish between ALD and Zellweger spectrum disorders. In summary, MS/MS analysis of plasma C24:0- and C26:0-LPC concentrations represents a rapid and straightforward approach to diagnose PDs, demonstrating superior diagnostic accuracy, particularly in ALD females, compared with conventional VLCFA biomarkers. We strongly recommend integrating very-long chain LPC plasma analysis in the diagnostic evaluation of individuals suspected of having a PD.

Analysis of a second-tier test panel in dried blood spot samples using liquid chromatography-tandem mass spectrometry in Catalonia's newborn screening programme.

OBJECTIVES: Acylcarnitine and amino acid analyses of dried blood spot (DBS) samples using tandem mass spectrometry in newborn screening (NBS) programmes can generate false positive (FP) results. Therefore, implementation of second-tier tests (2TTs) using DBS samples has become increasingly important to avoid FPs. The most widely used 2TT metabolites include methylmalonic acid, 3-hydroxypropionic acid, methylcitric acid, and homocysteine. METHODS: We simultaneously measured 46 underivatised metabolites, including organic acids, acylglycine and acylcarnitine isomers, homocysteine, and orotic acid, in DBS samples using tandem mass spectrometry. To validate this method, we analysed samples from 147 healthy newborns, 160 patients with genetic disorders diagnosed via NBS, 20 patients with acquired vitamin B12 deficiency, 10 newborns receiving antibiotic treatment, and nine external quality control samples. RESULTS: The validation study revealed that 31 metabolites showed good analytical performance. Furthermore, this method detected key metabolites for all diseases associated with increased levels of the following acylcarnitines: C3, C4, C5, C4DC/C5OH, and C5DC. The sensitivity of this method to detect all diseases was 100 %, and the specificity was 74-99 %, except for glutaric aciduria type 1. This method can also be used to diagnose mitochondrial fatty acid ß-oxidation disorders (FAODs) and urea cycle defects (UCDs). CONCLUSIONS: We have described a 2TT panel of 31 metabolites in DBS samples based on an easy and rapid method without derivatisation. Its implementation allowed us to distinguish between different organic acidurias, some FAODs, and UCDs. This new strategy has increased the efficiency of our NBS programme by reducing FP and false negative results, second sample requests, and the time required for diagnosis.

Foetal Haemoglobin as a Marker of Bone Marrow Suppression Secondary to Anti-Kell Alloimmunisation.

Anti-Kell alloimmunisation is a potentially severe minor blood group type incompatibility, not only as a cause of haemolytic disease of the foetus and newborn, but also due to the destruction of red blood cells (RBC) and mature form in the bone marrow with the subsequent hyporegenerative anaemia. In severe cases and when the foetus shows signs of anaemia, an intrauterine transfusion (IUT) may be necessary. When repeated, this treatment can suppress erythropoiesis and worsen the anaemia. We report the case of a newborn who required four IUTs plus an additional RBC transfusion at one month of life due to late onset anaemia. The identification of an adult haemoglobin profile with a complete absence of foetal haemoglobin in the patient's newborn screening samples at 2 and 10 days of life warned us of a possible late anaemia. The newborn was successfully treated with transfusion, oral supplements and subcutaneous erythropoietin. A blood sample taken at 4 months of life showed the expected haemoglobin profile for that age with a foetal haemoglobin of 17.7%. This case illustrates the importance of a close follow-up of these patients, as well as the usefulness of the haemoglobin profile screening as a tool for anaemia assessment.

Inter-rater reliability assessment for the new-born screening quality assurance.

Introduction: To ensure the quality of the new-born screening (NBS), our laboratory reviewed the analytical procedure to detect subjective steps that may represent a risk to the patient. Two subjective activities were identified in the extra-analytical phases: the classification of dried blood spots (DBS) according to their quality and the assignment of haemoglobin patterns. To keep these activities under control, inter-rater studies were implemented. This study aimed to evaluate the inter-rater reliability and the effectiveness of the measures taken to improve the agreement between observers, to assure NBS results' quality. Materials and methods: Dried blood spots specimens were used for the inter-rater studies. Ten studies were performed to assess DBS quality classification, and four to assess the assignment of haemoglobin patterns. Krippendorff's alpha test was used to estimate inter-rater reliability. Causes were investigated when alpha values were below 0.80. Results: For both activities, the reliability obtained in the first studies was inadequate. After investigation, we detected that the criterion to classify a DBS as scant was not consolidated, and also a lack of consensus on whether or not to report Bart's haemoglobin depending on its percentage. Alpha estimates became higher once the training was reinforced and a consensus about the appropriate criteria to be applied was reached. Conclusion: Inter-rater reliability assessment helped us to ensure the quality of subjective activities that could add variability to NBS results. Furthermore, the evolution of the alpha value over time allowed us to verify the effectiveness of the measures adopted.

Early Diagnosis and Treatment of Purine Nucleoside Phosphorylase (PNP) Deficiency through TREC-Based Newborn Screening.

Purine nucleoside phosphorylase (PNP) deficiency is a rare inherited disorder, resulting in severe combined immunodeficiency. To date, PNP deficiency has been detected in newborn screening only through the use of liquid chromatography tandem mass spectrometry. We report the first case in which PNP deficiency was detected by TREC analysis.

Newborn Screening for SCID. Experience in Spain (Catalonia).

Newborn screening (NBS) for severe combined immunodeficiency (SCID) started in Catalonia in January-2017, being the first Spanish and European region to universally include this testing. In Spain, a pilot study with 5000 samples was carried out in Seville in 2014; also, a research project with about 35,000 newborns will be carried out in 2021-2022 in the NBS laboratory of Eastern Andalusia. At present, the inclusion of SCID is being evaluated in Spain. The results obtained in the first three and a half years of experience in Catalonia are presented here. All babies born between January-2017 and June-2020 were screened through TREC-quantification in DBS with the Enlite Neonatal TREC-kit from PerkinElmer. A total of 222,857 newborns were screened, of which 48 tested positive. During the study period, three patients were diagnosed with SCID: an incidence of 1 in 74,187 newborns; 17 patients had clinically significant T-cell lymphopenia (non-SCID) with an incidence of 1 in 13,109 newborns who also benefited from the NBS program. The results obtained provide further evidence of the benefits of early diagnosis and curative treatment to justify the inclusion of this disease in NBS programs. A national NBS program is needed, also to define the exact SCID incidence in Spain.

[Beginnings, evolution and current situation of the Newborn Screening Programs in Spain.] / Inicio, evolución y situación actual de los Programas de Cribado Neonatal en España.

Newborn Screening Programs (NSP) in Spain were born in the city of Granada in 1968. Till the 1980s, they were developed around the so-called "National Plan for Preventing Subnormality", covering up to 30% of the Spanish newborns. From 1982, when the health system management was transferred to the different autonomous regions, the NSP began to expand, and the bases to transform them into an organized and multidisciplinary activity, integrated and coordinated from the National Health System were settled. Despite this expansion, it is not until the 1990s when their coverage reaches almost 100% newborns in Spain. NSP grew up asymmetrically across the different autonomous regions. In 2005 and 2006 the scientific societies SEQC (Spanish Society of Clinical Chemistry) and AECNE (Spanish Society of Newborn Screening), coordinated by the Health Promotion Area of the General Directorate of Public Health, gathered together the necessary information to elaborate a report on the NSP in Spain addressed to the Interterritorial Council of the National Health System. In July 2013, that Council approved the seven diseases that should be part of each region newborn screening panel, being the first step towards the NSP harmonization in Spain. Currently, the NSP include between 8 and 29 diseases in their panels, thus more still more efforts are needed in order to achieve a higher uniformity.

Los Programas de Cribado Neonatal (PCN) nacen en España en Granada en el año 1968. Posteriormente, y hasta los años 80, se fueron desarrollando en torno al llamado "Plan Nacional de Prevención de la Subnormalidad" con una cobertura cercana al 30% de los recién nacidos españoles. A partir de 1982, con el inicio de la gestión de la sanidad a las comunidades autónomas (CCAA), los PCN se expandieron y se comenzaron a sentar las bases para que éstos se convirtieran en una actividad organizada y multidisciplinar, integrados y coordinados desde el Sistema de Salud. A pesar de dicha expansión no es hasta el inicio de la década de los 90 cuando se consigue una cobertura próxima al 100% de los RN en España. Los PCN fueron creciendo de forma muy asimétrica en las diferentes CCAA y en los años 2005 y 2006 las Sociedades Científicas SEQC (Sociedad Española de Química Clínica) y AECNE (Asociación Española de Cribado Neonatal), con la coordinación del Área de Promoción de la Salud de la Dirección General de Salud Pública, recopilaron la información y elaboraron un informe, sobre los PCN en España para el Consejo Interterritorial del sistema Nacional de Salud (CISNS). En julio de 2013 este Consejo aprobó las siete enfermedades que debían formar parte del panel de detección de los PCN territoriales, primer paso hacia la armonización de estos programas. Actualmente, los PCN incluyen entre 8 y 29 enfermedades por lo que es necesario seguir trabajando para conseguir una mayor uniformidad.

[Analysis of the implementation of a unified transportation system of the neonatal screening samples in Catalonia.] / Análisis de la implementación de un sistema de transporte unificado de las muestras de cribado neonatal en Cataluña.

The Neonatal Screening Program in Catalonia from its inception fifty years ago until today, has enabled the early diagnosis and treatment of more than 2,000 newborns. In the last decade, the Program has undergone various extensions regarding its panel of diseases and has improved its evaluation with the inclusion of quality indicators in all its stages. One of the pending subjects of the screening program has been the improvement of the quality indexes related to the sample's arrival time to the laboratory after their extraction. The extension of the territory, the dispersion of numerous maternal centers, as well as the diversity and heterogeneity of the sample transport systems, have been an obstacle to quality compliance of these indexes. With the aim of reducing the period of samples arrival to the laboratory and continue to move towards meeting the standards established by the Ministry of Health, in 2020 a unified sample transport system has been implemented for the entire Catalan territory. The times obtained during the first months with the new system, have shown a notable improvement in the results, achieving a reduction of 50% of the days between the extraction of the sample and its arrival at the laboratory.

El Programa de Cribado Neonatal (PCN) de Cataluña ha permitido el diagnóstico y tratamiento precoz de más de 2.000 recién nacidos desde su inicio hace cincuenta años hasta la actualidad. En la última década, el PCN ha experimentado diversas ampliaciones en cuanto a su panel de enfermedades y ha mejorado su evaluación con la inclusión de indicadores de calidad en todas sus etapas. Una de las asignaturas pendientes del programa de cribado ha sido la mejora de los indicadores relativos al tiempo de llegada de las muestras al laboratorio desde su extracción. La extensión territorial, la dispersión de los sesenta y seis centros maternales, así como la diversidad y heterogeneidad de los sistemas de transporte de muestras, han supuesto un obstáculo para el cumplimiento de la calidad de este indicador. Con el objetivo de reducir el período de llegada de las muestras al laboratorio y seguir avanzando en el cumplimiento de los estándares establecidos por el Ministerio de Sanidad, en 2020 se ha implementado un sistema de transporte de muestras unificado para todo el territorio catalán. Los tiempos obtenidos durante los primeros meses con el nuevo sistema muestran una mejora notable de los resultados, consiguiendo una reducción del 50% de los días transcurridos desde la extracción de la muestra hasta su llegada al laboratorio.

[Response of the Catalonia neonatal screening laboratory to the pandemic of SARS-CoV-2.] / Respuesta del Laboratorio de Cribado Neonatal de Cataluña ante la pandemia por SARS-CoV-2.

Faced with the prospect of a collapsed health system due to the COVID-19 pandemic, the professionals involved in the Neonatal Screening Programme (NSP) of Catalonia had to adapt to this situation in a flexible, forceful and efficient manner. The most important goals were to prevent the risk of infection in the professionals, in families and their newborns, as well as to ensure the same effectiveness for the early detection of the diseases included in our programme. To this end, the laboratory was reorganised by dividing the staff into groups and the spaces were redistributed. It was also necessary to modify several protocols and circuits, especially for the management of early discharges from maternity centres, and for the collection of the necessary second samples (from newborns with inconclusive results or for low quality samples). In general, a 36% reduction in the time of arrival of these second samples at the laboratory was achieved with respect to the previous circuit. In the specific case of cystic fibrosis detection, the implementation of a new strategy meant a 100% reduction in the request for second samples and a 70% reduction in the age of diagnosis of the newborn. After evaluating these changes, it can be concluded that in the face of the pandemic, the NSP of Catalonia showed determined leadership, aligning all its professionals, ensuring the continuity of the activity in the programme and generating new opportunities. The new processes and circuits implemented have been definitively consolidated, improving the efficiency of the programme.

Ante la crisis de un sistema sanitario colapsado debido a la pandemia por la COVID-19, los profesionales implicados en el Programa de Cribado Neonatal (PCN) de Cataluña nos tuvimos que adaptar a dicha situación de forma ágil, contundente y eficiente. Los objetivos prioritarios fueron prevenir el riesgo de contagio tanto en los profesionales sanitarios como en las familias y sus recién nacidos, así como asegurar la misma eficacia para la detección precoz de las enfermedades incluidas en el PCN. Para ello, se reorganizó el laboratorio dividiendo en grupos al personal y se redistribuyeron los espacios. También fue necesario modificar varios protocolos y circuitos, en especial para la gestión de las altas precoces de los centros maternales y para la toma de las segundas muestras necesarias (de recién nacidos que presentaron resultados dudosos o por muestra inválida). En general, se consiguió una reducción del 36% del tiempo de llegada de estas segundas muestras al laboratorio respecto al circuito anterior. Para la detección de la fibrosis quística, la implementación de una nueva estrategia supuso una reducción del 100% en la solicitud de segundas muestras y del 70% en la edad de diagnóstico del recién nacido. Tras la evaluación de estos cambios, se puede concluir que ante la pandemia el PCN de Cataluña mostró un liderazgo decidido, alineando a todos sus profesionales, asegurando la continuidad de la actividad en el programa y generando nuevas oportunidades. Los nuevos procesos y circuitos de trabajo implantados han quedado definitivamente consolidados, mejorando la eficiencia del programa.

[Impact of the inclusion of second-tier tests in the newborn screening program of Catalonia and in other international programs.] / Impacto de la inclusión de pruebas de segundo nivel en el programa de cribado neonatal de Cataluña y en otros programas internacionales.

OBJECTIVE: Newborn screening programmes (NBSP) have experienced a qualitative breakthrough due to the implementation of tandem mass spectrometry. However, the tests used give rise to false positives (FP) generating an excessive request for second samples with the consequent anxiety of the families. In order to avoid this problem several programmes have developed second-tier tests (2TT). METHODS: This article presents our experience in the implementation of 2TT in the NBSP of Catalonia, as well as in other international programmes. RESULTS: From 2004 to the present, 2TT tests have been developed for more than 30 diseases. The use of 2TT helps to decrease the FP rate and increase the positive predictive value (PPV). In the NBSP of Catalonia, the implementation of 2TT for the detection of methylmalonic and propionic acidemias, homocystinurias, maple syrup disease and citrulinaemia, has managed to increase the PPV to 95% and decrease the PF rate to less than 0.01%. In cystic fibrosis, the application of 2TT slightly increases PPV but with a significant decrease in the request for second samples and in the number of cases referred to clinical units. CONCLUSIONS: The introduction of 2TT in the NBSP allows to reduce considerably the FP, decreases the number of requested samples, as well as both anxiety and stress of the families, at the same time that the hospital costs are reduced and the PPV is increased, improving notably the efficiency of the NBSP.

OBJETIVO: Los programas de cribado neonatal (PCN) han experimentado un gran avance cualitativo debido a la implementación de la espectrometría de masas en tándem. Sin embargo, las pruebas utilizadas dan lugar a falsos positivos (FP) generando una excesiva solicitud de segundas muestras con la consiguiente ansiedad de las familias. Con el fin de evitar este problema diversos programas han desarrollado pruebas de segundo nivel (2TT). METODOS: En este artículo se presenta nuestra experiencia en la implementación de 2TT en el PCN de Cataluña, así como en otros programas internacionales. RESULTADOS: Desde el año 2004 hasta la actualidad se han desarrollado pruebas de 2TT para más de 30 enfermedades. La utilización de 2TT ayuda a disminuir la tasa de FP y aumentar el valor predictivo positivo (VPP). En el PCN de Cataluña, la implementación de 2TT para la detección de acidemias metilmalónicas y propiónica, homocistinurias, jarabe de arce y citrulinemia, ha conseguido aumentar el VPP a un 95% y disminuir la tasa de FP a menos del 0,01%. En la fibrosis quística la aplicación de 2TT aumenta ligeramente el VPP pero con disminución significativa de la solicitud de segundas muestras y de los casos referidos a las unidades clínicas. CONCLUSIONES: La introducción de los 2TT en los PCN permite reducir considerablemente los FP, disminuye el número de muestras solicitadas, así como la ansiedad y el estrés de las familias, a la vez que se reducen los costes hospitalarios y se aumenta el VPP, mejorando notablemente la eficiencia de los PCN.

[First universal newborn screening program for severe combined immunodeficiency in Europe. Three-years' experience in Catalonia.] / Primer programa europeo de cribado neonatal para la inmunodeficiencia combinada grave: experiencia de tres años en Cataluña.

Severe combined immunodeficiency (SCID), the most severe form of T-cell immunodeficiency, can be screened at birth by quantifying T-cell receptor excision circles (TREC) in dried blood spot (DBS) samples. Early detection of this condition speeds up the establishment of appropriate treatment and increases the patient's life expectancy. Newborn screening for SCID started in January 2017 in Catalonia, the first Spanish and European region to universally include this testing. The results obtained in the first three years and a half of experience (January 2017 - June 2020) are shown here, using EnLite Neonatal TREC kit (Perkin Elmer) with 20 copies/µL as TREC detection cutoff. Of 222,857 newborns screened, 48 tested positive: three patients were diagnosed with SCID (incidence 1:74,285); 17 patients had clinically significant T-cell lymphopenia (non-SCID) with an incidence of 1 in 13,109 newborns; twenty two patients were considered false-positive cases because of an initially normal lymphocyte count with normalization of TREC between 3 and 6 months of life; one case had transient lymphopenia due to an initially low lymphocyte count with recovery in the following months; and five patients are still under study. The results obtained provide further evidence of the benefits of including this disease in newborn screening programs. Even longer follow-up could be necessary to define the exact incidence of SCID in Catalonia.

La inmunodeficiencia combinada grave (IDCG) es la forma más grave de inmunodeficiencia primaria, que afecta sobre todo a los linfocitos T, y puede ser detectada al nacer mediante la cuantificación de los círculos de escisión del receptor de linfocitos T (TREC) en una muestra de sangre impregnada en papel (DBS). La detección precoz de esta enfermedad permite establecer de forma temprana un tratamiento adecuado en el paciente, permitiendo así su curación. El cribado neonatal de IDCG comenzó en Cataluña en enero de 2017, siendo la primera región española y europea en incluirla oficial y universalmente en su programa. En el presente trabajo se presentan los resultados obtenidos durante los tres primeros años y medio de experiencia (enero 2017 ­ junio 2020) empleando el kit EnLite Neonatal TREC (Perkin Elmer), con un cutoff de detección de TREC de 20 copias/µL. De 222.857 recién nacidos analizados, cuarenta y ocho fueron detecciones positivas: tres casos de IDCG (incidencia de 1:74.285); diecisiete casos de linfopenia T no IDCG (incidencia de 1:13.109); veintidós casos falsos positivos (recuento de linfocitos inicialmente normal, con normalización de TREC entre los tres y seis meses de vida); un caso con linfopenia transitoria (con un recuento de linfocitos inicialmente bajo, que se normaliza en los meses siguientes); y cinco pacientes se encuentran todavía en estudio. Los resultados obtenidos aportan evidencias de los beneficios que supone incluir esta enfermedad en los programas de cribado neonatal. Podría ser necesario un seguimiento todavía más prolongado para acabar de definir la incidencia exacta de IDCG en Cataluña.

[50 years of the Neonatal Screening Program in Catalonia.] / 50 años del Programa de Cribado Neonatal en Cataluña.

The Catalonian Newborn Screening Program (CNSP) began in 1969, in Barcelona. It was promoted by Dr. Juan Sabater Tobella and supported by Barcelona Provincial Council and Juan March Foundation. That is how the Institute of Clinical Biochemistry was born, whose aims were diagnosis, research and teaching, along with the spirit of contributing to the prevention of mental retardation. The CNSP began with the detection of phenylketonuria (PKU), and, in 1982, the Program was expanded with the inclusion of congenital hypothyroidism detection. Towards 1990, the Program covered almost 100% of all newborns (NB) in Catalonia. In 1999, the CNSP was expanded with the incorporation of cystic fibrosis. It took fourteen years, until 2013, to make the largest expansion so far, with the incorporation of 19 metabolic diseases to the screening panel. The detection of sickle cell disease began in 2015 and in 2017 the detection of severe combined immunodeficiency was included. Currently, the CNSP includes 24 diseases in its main panel. Since 1969, 2,787,807 NBs have been screened, of whom 1,724 have been diagnosed with any of these diseases, and 252 of other disorders by differential diagnosis with those included in the main panel. The global prevalence is 1: 1,617 NBs affected by any of the diseases included in the CNSP and 1: 1,140 NBs if incidental findings diagnosed through the CNSP are included.

El Programa de Cribado Neonatal de Cataluña (PCNC) se inició en el año 1969, en Barcelona, impulsado por el Dr. Juan Sabater Tobella y apoyado por la Diputación de Barcelona y la Fundación Juan March. Así nació el Instituto de Bioquímica Clínica para acometer funciones asistenciales, de investigación y docencia, con el espíritu de contribuir a la prevención del retraso mental. El PCNC se inició con la detección de la fenilcetonuria (PKU) y en el año 1982 se amplió con la detección del hipotiroidismo congénito. Hacia el año 1990 la cobertura territorial llegó casi al 100% de todos los recién nacidos en Cataluña. En 1999 se amplió el PCNC con la incorporación de la fibrosis quística y tras catorce años, en 2013, se realizó la ampliación más numerosa hasta ahora, con la incorporación de la detección de 19 enfermedades metabólicas hereditarias. En el año 2015 comenzó la detección de la enfermedad de células falciformes y en el 2017 la detección de la inmunodeficiencia combinada grave. Actualmente, el PCNC incluye la detección de 24 enfermedades. Desde su inicio en el año 1969, se han cribado 2.787.807 recién nacidos, de los cuales 1.724 han sido diagnosticados de alguna de las 24 enfermedades que componen nuestro panel principal y 252 por diagnóstico diferencial de las primeras. En total la prevalencia global es de 1:1.617 RN afectos de alguna de las enfermedades incluidas en el PCNC y de 1:1.140 RN si se incluyen los hallazgos incidentales encontrados.

50 años del Programa de Cribado Neonatal en Cataluña / 50 years of the Neonatal Screening Program in Catalonia

El Programa de Cribado Neonatal de Cataluña (PCNC) se inició en el año 1969, en Barcelona, impulsado por el Dr. Juan Sabater Tobella y apoyado por la Diputación de Barcelona y la Fundación Juan March. Así nació el Instituto de Bioquímica Clínica para acometer funciones asistenciales, de investigación y docencia, con el espíritu de contribuir a la prevención del retraso mental. El PCNC se inició con la detección de la fenilcetonuria (PKU) y en el año 1982 se amplió con la detección del hipotiroidismo congénito. Hacia el año 1990 la cobertura territorial llegó casi al 100% de todos los recién nacidos en Cataluña. En 1999 se amplió el PCNC con la incorporación de la fibrosis quística y tras catorce años, en 2013, se realizó la ampliación más numerosa hasta ahora, con la incorporación de la detección de 19 enfermedades metabólicas hereditarias. En el año 2015 comenzó la detección de la enfermedad de células falciformes y en el 2017 la detección de la inmunodeficiencia combinada grave. Actualmente, el PCNC incluye la detección de 24 enfermedades. Desde su inicio en el año 1969, se han cribado 2.787.807 recién nacidos, de los cuales 1.724 han sido diagnosticados de alguna de las 24 enfermedades que componen nuestro panel principal y 252 por diagnóstico diferencial de las primeras. En total la prevalencia global es de 1:1.617 RN afectos de alguna de las enfermedades incluidas en el PCNC y de 1:1.140 RN si se incluyen los hallazgos incidentales encontrados

The Catalonian Newborn Screening Program (CNSP) began in 1969, in Barcelona. It was promoted by Dr. Juan Sabater Tobella and supported by Barcelona Provincial Council and Juan March Foundation. That is how the Institute of Clinical Biochemistry was born, whose aims were diagnosis, research and teaching, along with the spirit of contributing to the prevention of mental retardation. The CNSP began with the detection of phenylketonuria (PKU), and, in 1982, the Program was expanded with the inclusion of congenital hypothyroidism detection. Towards 1990, the Program covered almost 100% of all newborns (NB) in Catalonia. In 1999, the CNSP was expanded with the incorporation of cystic fibrosis. It took fourteen years, until 2013, to make the largest expansion so far, with the incorporation of 19 metabolic diseases to the screening panel. The detection of sickle cell disease began in 2015 and in 2017 the detection of severe combined immunodeficiency was included. Currently, the CNSP includes 24 diseases in its main panel. Since 1969, 2,787,807 NBs have been screened, of whom 1,724 have been diagnosed with any of these diseases, and 252 of other disorders by differential diagnosis with those included in the main panel. The global prevalence is 1: 1,617 NBs affected by any of the diseases included in the CNSP and 1: 1,140 NBs if incidental findings diagnosed through the CNSP are included

Análisis de la implementación de un sistema de transporte unificado de las muestras de cribado neonatal en Cataluña / Analysis of the implementation of a unified transportation system of the neonatal screening samples in Catalonia

El Programa de Cribado Neonatal (PCN) de Cataluña ha permitido el diagnóstico y tratamiento precoz de más de 2.000 recién nacidos desde su inicio hace cincuenta años hasta la actualidad. En la última década, el PCN ha experimentado diversas ampliaciones en cuanto a su panel de enfermedades y ha mejorado su evaluación con la inclusión de indicadores de calidad en todas sus etapas. Una de las asignaturas pendientes del programa de cribado ha sido la mejora de los indicadores relativos al tiempo de llegada de las muestras al laboratorio desde su extracción. La extensión territorial, la dispersión de los sesenta y seis centros maternales, así como la diversidad y heterogeneidad de los sistemas de transporte de muestras, han supuesto un obstáculo para el cumplimiento de la calidad de este indicador. Con el objetivo de reducir el período de llegada de las muestras al laboratorio y seguir avanzando en el cumplimiento de los estándares establecidos por el Ministerio de Sanidad, en 2020 se ha implementado un sistema de transporte de muestras unificado para todo el territorio catalán. Los tiempos obtenidos durante los primeros meses con el nuevo sistema muestran una mejora notable de los resultados, consiguiendo una reducción del 50% de los días transcurridos desde la extracción de la muestra hasta su llegada al laboratorio

The Neonatal Screening Program in Catalonia from its inception fifty years ago until today, has enabled the early diagnosis and treatment of more than 2,000 newborns. In the last decade, the Program has undergone various extensions regarding its panel of diseases and has improved its evaluation with the inclusion of quality indicators in all its stages. One of the pending subjects of the screening program has been the improvement of the quality indexes related to the sample's arrival time to the laboratory after their extraction. The extension of the territory, the dispersion of numerous maternal centers, as well as the diversity and heterogeneity of the sample transport systems, have been an obstacle to quality compliance of these indexes. With the aim of reducing the period of samples arrival to the laboratory and continue to move towards meeting the standards established by the Ministry of Health, in 2020 a unified sample transport system has been implemented for the entire Catalan territory. The times obtained during the first months with the new system, have shown a notable improvement in the results, achieving a reduction of 50% of the days between the extraction of the sample and its arrival at the laboratory

Impacto de la inclusión de pruebas de segundo nivel en el programa de cribado neonatal de Cataluña y en otros programas internacionales / Impact of the inclusion of second-tier tests in the newborn screening program of Catalonia and in other international programs

OBJETIVO: Los programas de cribado neonatal (PCN) han experimentado un gran avance cualitativo debido a la implementación de la espectrometría de masas en tándem. Sin embargo, las pruebas utilizadas dan lugar a falsos positivos (FP) generando una excesiva solicitud de segundas muestras con la consiguiente ansiedad de las familias. Con el fin de evitar este problema diversos programas han desarrollado pruebas de segundo nivel (2TT). MÉTODOS: En este artículo se presenta nuestra experiencia en la implementación de 2TT en el PCN de Cataluña, así como en otros programas internacionales. RESULTADOS: Desde el año 2004 hasta la actualidad se han desarrollado pruebas de 2TT para más de 30 enfermedades. La utilización de 2TT ayuda a disminuir la tasa de FP y aumentar el valor predictivo positivo (VPP). En el PCN de Cataluña, la implementación de 2TT para la detección de acidemias metilmalónicas y propiónica, homocistinurias, jarabe de arce y citrulinemia, ha conseguido aumentar el VPP a un 95% y disminuir la tasa de FP a menos del 0,01%. En la fibrosis quística la aplicación de 2TT aumenta ligeramente el VPP pero con disminución significativa de la solicitud de segundas muestras y de los casos referidos a las unidades clínicas. CONCLUSIONES: La introducción de los 2TT en los PCN permite reducir considerablemente los FP, disminuye el número de muestras solicitadas, así como la ansiedad y el estrés de las familias, a la vez que se reducen los costes hospitalarios y se aumenta el VPP, mejorando notablemente la eficiencia de los PCN

OBJECTIVE: Newborn screening programmes (NBSP) have experienced a qualitative breakthrough due to the implementation of tandem mass spectrometry. However, the tests used give rise to false positives (FP) generating an excessive request for second samples with the consequent anxiety of the families. In order to avoid this problem several programmes have developed second-tier tests (2TT). METHODS: This article presents our experience in the implementation of 2TT in the NBSP of Catalonia, as well as in other international programmes. RESULTS: From 2004 to the present, 2TT tests have been developed for more than 30 diseases. The use of 2TT helps to decrease the FP rate and increase the positive predictive value (PPV). In the NBSP of Catalonia, the implementation of 2TT for the detection of methylmalonic and propionic acidemias, homocystinurias, maple syrup disease and citrulinaemia, has managed to increase the PPV to 95% and decrease the PF rate to less than 0.01%. In cystic fibrosis, the application of 2TT slightly increases PPV but with a significant decrease in the request for second samples and in the number of cases referred to clinical units. CONCLUSIONS: The introduction of 2TT in the NBSP allows to reduce considerably the FP, decreases the number of requested samples, as well as both anxiety and stress of the families, at the same time that the hospital costs are reduced and the PPV is increased, improving notably the efficiency of the NBSP

Primer programa europeo de cribado neonatal para la inmunodeficiencia combinada grave: experiencia de tres años en Cataluña / First universal newborn screening program for severe combined immunodeficiency in Europe. Three-years' experience in Catalonia

La inmunodeficiencia combinada grave (IDCG) es la forma más grave de inmunodeficiencia primaria, que afecta sobre todo a los linfocitos T, y puede ser detectada al nacer mediante la cuantificación de los círculos de escisión del receptor de linfocitos T (TREC) en una muestra de sangre impregnada en papel (DBS). La detección precoz de esta enfermedad permite establecer de forma temprana un tratamiento adecuado en el paciente, permitiendo así su curación. El cribado neonatal de IDCG comenzó en Cataluña en enero de 2017, siendo la primera región española y europea en incluirla oficial y universalmente en su programa. En el presente trabajo se presentan los resultados obtenidos durante los tres primeros años y medio de experiencia (enero 2017 - junio 2020) empleando el kit EnLite Neonatal TREC (Perkin Elmer), con un cutoff de detección de TREC de 20 copias/miL. De 222.857 recién nacidos analizados, cuarenta y ocho fueron detecciones positivas: tres casos de IDCG (incidencia de 1:74.285); diecisiete casos de linfopenia T no IDCG (incidencia de 1:13.109); veintidós casos falsos positivos (recuento de linfocitos inicialmente normal, con normalización de TREC entre los tres y seis meses de vida); un caso con linfopenia transitoria (con un recuento de linfocitos inicialmente bajo, que se normaliza en los meses siguientes); y cinco pacientes se encuentran todavía en estudio. Los resultados obtenidos aportan evidencias de los beneficios que supone incluir esta enfermedad en los programas de cribado neonatal. Podría ser necesario un seguimiento todavía más prolongado para acabar de definir la incidencia exacta de IDCG en Cataluña

Severe combined immunodeficiency (SCID), the most severe form of T-cell immunodeficiency, can be screened at birth by quantifying T-cell receptor excision circles (TREC) in dried blood spot (DBS) samples. Early detection of this condition speeds up the establishment of appropriate treatment and increases the patient's life expectancy. Newborn screening for SCID started in January 2017 in Catalonia, the first Spanish and European region to universally include this testing. The results obtained in the first three years and a half of experience (January 2017 - June 2020) are shown here, using EnLite Neonatal TREC kit (Perkin Elmer) with 20 copies/miL as TREC detection cutoff. Of 222,857 newborns screened, 48 tested positive: three patients were diagnosed with SCID (incidence 1:74,285); 17 patients had clinically significant T-cell lymphopenia (non-SCID) with an incidence of 1 in 13,109 newborns; twenty two patients were considered false-positive cases because of an initially normal lymphocyte count with normalization of TREC between 3 and 6 months of life; one case had transient lymphopenia due to an initially low lymphocyte count with recovery in the following months; and five patients are still under study. The results obtained provide further evidence of the benefits of including this disease in newborn screening programs. Even longer follow-up could be necessary to define the exact incidence of SCID in Catalonia

Respuesta del laboratorio de cribado neonatal de Cataluña ante la pandemia por SARS-CoV-2 / Response of the Catalonia neonatal screening laboratory to the pandemic of SARS-CoV-2

Ante la crisis de un sistema sanitario colapsado debido a la pandemia por la COVID-19, los profesionales implicados en el Programa de Cribado Neonatal (PCN) de Cataluña nos tuvimos que adaptar a dicha situación de forma ágil, contundente y eficiente. Los objetivos prioritarios fueron prevenir el riesgo de contagio tanto en los profesionales sanitarios como en las familias y sus recién nacidos, así como asegurar la misma eficacia para la detección precoz de las enfermedades incluidas en el PCN. Para ello, se reorganizó el laboratorio dividiendo en grupos al personal y se redistribuyeron los espacios. También fue necesario modificar varios protocolos y circuitos, en especial para la gestión de las altas precoces de los centros maternales y para la toma de las segundas muestras necesarias (de recién nacidos que presentaron resultados dudosos o por muestra inválida). En general, se consiguió una reducción del 36% del tiempo de llegada de estas segundas muestras al laboratorio respecto al circuito anterior. Para la detección de la fibrosis quística, la implementación de una nueva estrategia supuso una reducción del 100% en la solicitud de segundas muestras y del 70% en la edad de diagnóstico del recién nacido. Tras la evaluación de estos cambios, se puede concluir que ante la pandemia el PCN de Cataluña mostró un liderazgo decidido, alineando a todos sus profesionales, asegurando la continuidad de la actividad en el programa y generando nuevas oportunidades. Los nuevos procesos y circuitos de trabajo implantados han quedado definitivamente consolidados, mejorando la eficiencia del programa

Faced with the prospect of a collapsed health system due to the COVID-19 pandemic, the professionals involved in the Neonatal Screening Programme (NSP) of Catalonia had to adapt to this situation in a flexible, forceful and efficient manner. The most important goals were to prevent the risk of infection in the professionals, in families and their newborns, as well as to ensure the same effectiveness for the early detection of the diseases included in our programme. To this end, the laboratory was reorganised by dividing the staff into groups and the spaces were redistributed. It was also necessary to modify several protocols and circuits, especially for the management of early discharges from maternity centres, and for the collection of the necessary second samples (from newborns with inconclusive results or for low quality samples). In general, a 36% reduction in the time of arrival of these second samples at the laboratory was achieved with respect to the previous circuit. In the specific case of cystic fibrosis detection, the implementation of a new strategy meant a 100% reduction in the request for second samples and a 70% reduction in the age of diagnosis of the newborn. After evaluating these changes, it can be concluded that in the face of the pandemic, the NSP of Catalonia showed determined leadership, aligning all its professionals, ensuring the continuity of the activity in the programme and generating new opportunities. The new processes and circuits implemented have been definitively consolidated, improving the efficiency of the programme

First Universal Newborn Screening Program for Severe Combined Immunodeficiency in Europe. Two-Years' Experience in Catalonia (Spain).

Severe combined immunodeficiency (SCID), the most severe form of T-cell immunodeficiency, can be screened at birth by quantifying T-cell receptor excision circles (TRECs) in dried blood spot (DBS) samples. Early detection of this condition speeds up the establishment of appropriate treatment and increases the patient's life expectancy. Newborn screening for SCID started in January 2017 in Catalonia, the first Spanish and European region to universally include this testing. The results obtained in the first 2 years of experience are evaluated here. All babies born between January 2017 and December 2018 were screened. TREC quantification in DBS (1.5 mm diameter) was performed with the Enlite Neonatal TREC kit from PerkinElmer (Turku, Finland). In 2018, the retest cutoff in the detection algorithm was updated based on the experience gained in the first year, and changed from 34 to 24 copies/µL. This decreased the retest rate from 3.34 to 1.4% (global retest rate, 2.4%), with a requested second sample rate of 0.23% and a positive detection rate of 0.02%. Lymphocyte phenotype (T, B, NK populations), expression of CD45RA/RO isoforms, percentage and intensity of TCR αß and TCR γδ, presence of HLA-DR+ T lymphocytes, and in vitro lymphocyte proliferation were studied in all patients by flow cytometry. Of 130,903 newborns screened, 30 tested positive, 15 of which were male. During the study period, one patient was diagnosed with SCID: incidence, 1 in 130,903 births in Catalonia. Thirteen patients had clinically significant T-cell lymphopenia (non-SCID) with an incidence of 1 in 10,069 newborns (43% of positive detections). Nine patients were considered false-positive cases because of an initially normal lymphocyte count with normalization of TRECs between 3 and 6 months of life, four infants had transient lymphopenia due to an initially low lymphocyte count with recovery in the following months, and three patients are still under study. The results obtained provide further evidence of the benefits of including this disease in newborn screening programs. Longer follow-up is needed to define the exact incidence of SCID in Catalonia.

Identification of 22q11.2 deletion syndrome via newborn screening for severe combined immunodeficiency. Two years' experience in Catalonia (Spain).

BACKGROUND: The current scenario of newborn screening is changing as DNA studies are being included in the programs of several countries. Severe combined immunodeficiency (SCID) disorders can be detected using quantitative PCR assays to measure T-cell receptor excision circles (TRECs), a byproduct of correct T-cell development. However, in addition to SCID, other T-cell-deficient phenotypes such as 22q11.2 deletion syndrome 22q11.2 duplication syndrome, CHARGE syndrome, and trisomy 21 are detected. METHODS: We present our experience with the detection of 22q11.2 deletion syndrome and 22q11.2 duplication syndrome in a series of 103,903 newborns included in the newborn screening program of Catalonia (Spain). RESULTS: Thirty newborns tested were positive (low TREC levels) and five were found to have copy number variations at the 22q11 region (4 deletions and 1 duplication) when investigated with array comparative genomic hybridization technology and MLPA. CONCLUSION: Newborn screening for SCID enables detection of several conditions, such as 22q syndromes, which should be managed by prompt, proactive approaches with adequate counseling for families by a multidisciplinary team.

Propuesta para mejorar la sensibilidad del algoritmo diagnóstico de evaluación de la función tiroidea / Proposal to improve the sensitivity of the diagnostic algorithm for thyroid function evaluation

La medida en suero de la concentración de tirotropina y tiroxina es la base para la evaluación bioquímica de la función tiroidea. Con frecuencia, el intervalo de referencia de la tirotropina sirve como cribado inicial para valorar la necesidad de añadir la medida de tiroxina. Este trabajo se ha realizado con el objetivo de mejorar la sensibilidad diagnóstica del cribado. Se seleccionaron todos los resultados de tirotropina y tiroxina solicitados de manera simultánea a pacientes de consultas externas: para la primera parte del estudio se usaron los del año 2008 (n=10.900) y para la segunda parte, los de pacientes del año 2009 sin seguimiento en el año previo (n=5.367). Se realizaron dos curvas ROC para delimitar el intervalo de decisión del algoritmo con una sensibilidad del 90% y se contabilizó el número de resultados falsos negativos obtenidos. Los intervalos de tirotropina obtenidos en el primer y segundo estudio fueron (2,11-3,50) mint.u./L y (2,04-3,41) mint.u/L respectivamente. En ambos estudios la sensibilidad aumentó aproximadamente de un 70% de media con el intervalo de referencia a un 90% con el intervalo del algoritmo. El número de falsos negativos se redujo de 75 a 30 en el primer caso, y de 37 a 13 en el segundo. La aplicación de un intervalo de tirotropina calculado para la evaluación de la función tiroidea, en pacientes ambulatorios con o sin seguimiento previo, supone un aumento en la sensibilidad diagnóstica, respecto al empleo del intervalo de referencia de tirotropina (AU)

The measurement of thyrotropin and thyroxine concentrations in serum is the basis of the biochemical evaluation of thyroid function. The reference interval of thyrotropin is frequently used as an initial screening to assess the need for thyroxine measurement. This study was carried out to obtain a different and more adjusted interval of thyrotropin, in order to improve the diagnostic sensitivity. All of the results of thyrotropin and thyroxine requested at the same time on outpatients were selected: for the first part of the study, those from year 2008 (n=10,900), and for the second part, those from 2009 with no follow-up in the previous year (n=5,367). Two ROC curves were used to define the algorithm decision interval with a sensitivity of 90% and the number of false negative results was calculated. The thyrotropin intervals obtained in the first and second studies were (2.11-3.50) mIU/L and (2.04-3.41) mIU/L, respectively. In both studies, the sensitivity increased approximately from an average of 70% to 90% of the confidence interval using the algorithm interval. The number of false negatives was reduced from 75 to 30 in the first case, and from 37 to 13 in the second case. The application of a calculated thyrotropin interval to assess thyroid function in outpatients with or without prior monitoring, leads to an increase of the diagnostic sensitivity with regard to the use of the thyrotropin reference interval (AU)