Molecular and cellular pediatrics最新文献

{"title":"Correction to: Novel ways to monitor immunosuppression in pediatric kidney transplant recipients-underlying concepts and emerging data.","authors":"Thurid Ahlenstiel-Grunow, Lars Pape","doi":"10.1186/s40348-021-00124-w","DOIUrl":"https://doi.org/10.1186/s40348-021-00124-w","url":null,"abstract":"","PeriodicalId":74215,"journal":{"name":"Molecular and cellular pediatrics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2021-09-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8484395/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39472684","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"How FGF23 shapes multiple organs in chronic kidney disease.","authors":"Maren Leifheit-Nestler,&nbsp;Dieter Haffner","doi":"10.1186/s40348-021-00123-x","DOIUrl":"https://doi.org/10.1186/s40348-021-00123-x","url":null,"abstract":"<p><p>Chronic kidney disease (CKD) is associated with distinct alterations in mineral metabolism in children and adults resulting in multiple organ dysfunctions. Children with advanced CKD often suffer from impaired bone mineralization, bone deformities and fractures, growth failure, muscle weakness, and vascular and soft tissue calcification, a complex which was recently termed CKD-mineral and bone disorder (CKD-MBD). The latter is a major contributor to the enhanced cardiovascular disease comorbidity and mortality in these patients. Elevated circulating levels of the endocrine-acting phosphaturic hormone fibroblast growth factor (FGF) 23 are the first detectable alteration of mineral metabolism and thus CKD-MBD. FGF23 is expressed and secreted from osteocytes and osteoblasts and rises, most likely due to increased phosphate load, progressively as kidney function declines in order to maintain phosphate homeostasis. Although not measured in clinical routine yet, CKD-mediated increased circulating levels of FGF23 in children are associated with pathological cardiac remodeling, vascular alterations, and increased cognitive risk. Clinical and experimental studies addressing other FGF23-mediated complications of kidney failure, such as hypertension and impaired bone mineralization, show partly conflicting results, and the causal relationships are not always entirely clear. This short review summarizes regulators of FGF23 synthesis altered in CKD and the main CKD-mediated organ dysfunctions related to high FGF23 levels.</p>","PeriodicalId":74215,"journal":{"name":"Molecular and cellular pediatrics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2021-09-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8449753/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39427624","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"ESPED survey on newly diagnosed immune thrombocytopenia in childhood: how much treatment do we give?","authors":"Hannah von Lukowicz,&nbsp;Paul-Gerhardt Schlegel,&nbsp;Christoph Härtel,&nbsp;Henner Morbach,&nbsp;Imme Haubitz,&nbsp;Verena Wiegering","doi":"10.1186/s40348-021-00121-z","DOIUrl":"https://doi.org/10.1186/s40348-021-00121-z","url":null,"abstract":"<p><strong>Background: </strong>Immune thrombocytopenia (ITP) is an autoimmune disease associated with isolated thrombocytopenia, which is caused by an imbalance between platelet production and platelet destruction. Petechial and mucous membrane hemorrhages are characteristic of ITP, but life-threatening bleeding rarely occurs. Depending on the bleeding symptoms, ITP can be treated with glucocorticoids (GC), intravenous immunoglobulins (IVIG), or in severe cases, platelet transfusions. Mild bleeding does not necessarily require therapy. Using the German Surveillance Unit for rare Pediatric Diseases (ESPED) we conducted a prospective survey on ITP patients in all German Children's Hospitals between September 2018 and August 2019. We collected data on ITP, including the clinical course, therapy implementation recommendations (according to the Association of German Scientific Medical Societies guidelines), outcome, and influence of treatment regimens depending on the treating physician´s experience with ITP patients.</p><p><strong>Results: </strong>Of the 287 recorded cases of children with ITP, 268 questionnaires were sent to the authors. Two hundred seventeen of the questionnaires fulfilled the inclusion criteria. ITP affected boys and girls similarly, and the median age of manifestation was 3.5 years. The main reasons for hospitalization were thrombocytopenia, bleeding signs, hematomas, and/or petechiae. Bleeding scores were ≤ 3 in 96% of children, which corresponded to a low-to-moderately low risk of bleeding. No life-threatening bleeding was documented. The most common therapies were IVIG (n = 59), GC (n = 33), or a combination of these (n = 17). Blood products (i.e., red blood cells, platelet concentrate, and fresh frozen plasma) were given to 13 patients. Compared to the established guidelines, 67 patients were over-treated, and 2 patients were under-treated.</p><p><strong>Conclusions: </strong>Adherence to German ITP treatment guidelines is currently limited. To improve patient safety and medical care, better medical training and dissemination of the guidelines are required in line with targeted analyses of patients with serious bleeding events to identify potential risk constellations.</p>","PeriodicalId":74215,"journal":{"name":"Molecular and cellular pediatrics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2021-09-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8419130/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39387895","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A fresh look to the phenotype in mono-allelic likely pathogenic variants of the leptin and the leptin receptor gene.","authors":"Ingrid Koerber-Rosso,&nbsp;Stephanie Brandt,&nbsp;Julia von Schnurbein,&nbsp;Pamela Fischer-Posovszky,&nbsp;Josef Hoegel,&nbsp;Hannah Rabenstein,&nbsp;Reiner Siebert,&nbsp;Martin Wabitsch","doi":"10.1186/s40348-021-00119-7","DOIUrl":"https://doi.org/10.1186/s40348-021-00119-7","url":null,"abstract":"<p><p>Leptin (LEP) and leptin receptor (LEPR) play a major role in energy homeostasis, metabolism, and reproductive function. While effects of biallelic likely pathogenic variants (-/-) on the phenotype are well characterized, effects of mono-allelic likely pathogenic variants (wt/-) in the LEP and LEPR gene on the phenotype compared to wild-type homozygosity (wt/wt) have not been systematically investigated. We identified in our systematic review 44 animal studies (15 on Lep, 29 on Lepr) and 39 studies in humans reporting on 130 mono-allelic likely pathogenic variant carriers with 20 distinct LEP variants and 108 heterozygous mono-allelic likely pathogenic variant carriers with 35 distinct LEPR variants. We found indications for a higher weight status in carriers of mono-allelic likely pathogenic variant in the leptin and in the leptin receptor gene compared to wt/wt, in both animal and human studies. In addition, animal studies showed higher body fat percentage in Lep and Lepr wt/- vs wt/wt. Animal studies provided indications for lower leptin levels in Lep wt/- vs. wt/wt and indications for higher leptin levels in Lepr wt/- vs wt/wt. Data on leptin levels in human studies was limited. Evidence for an impaired metabolism in mono-allelic likely pathogenic variants of the leptin and in leptin receptor gene was not conclusive (animal and human studies). Mono-allelic likely pathogenic variants in the leptin and in leptin receptor gene have phenotypic effects disposing to increased body weight and fat accumulation.</p>","PeriodicalId":74215,"journal":{"name":"Molecular and cellular pediatrics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2021-08-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8390564/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39356575","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The mutual patterning between the developing nephron and its covering tissues-valid reasons to rethink the search for traces left by impaired nephrogenesis.","authors":"Will W Minuth","doi":"10.1186/s40348-021-00120-0","DOIUrl":"https://doi.org/10.1186/s40348-021-00120-0","url":null,"abstract":"<p><strong>Background: </strong>The impairment of nephrogenesis can cause the termination of nephron formation in preterm and low birth weight babies. This leads to oligonephropathy with severe health consequences in later life. Although many clinical parameters are known, surprisingly little information is available regarding the initial damage on the developing nephron. Equally astounding, the first morphological data regarding the specifics of nephron formation in the nephrogenic zone of the fetal human kidney during late gestation has only been published within the past few years. In this context, it was observed that each stage of nephron anlage is surrounded by a specific set of tissues. Although highly relevant for the normal progress of nephron formation, the mutual patterning has not been systematically described.</p><p><strong>Results: </strong>To contribute, the different stages of nephron anlage in the nephrogenic zone of the fetal human kidney during late gestation were screened by the optical microscope and documented by images. Following this, magnifications (28 × 18 cm) were produced to trace the contours of the developing nephron and its covering tissues. The resulting sketches, almost true to scale, were scanned, edited, and processed by a design program. As a base, first the individual position, size, and shape of the nephrogenic niche, pretubular aggregate, renal vesicles, comma- and S-shaped bodies are presented. Secondly, their structural relations to the renal capsule, collecting duct ampulla, perforating radiate artery, and expanding interstitium are shown. Third of all, the focus is on less considered configurations, such as site-specific approximation, local distancing, punctual adhesion, integration, separation, delamination, formation of congruent and divergent surfaces, and folding and opening of interstitial clefts.</p><p><strong>Conclusions: </strong>The present contribution illuminates the mutual patterning between the developing nephron and its covering tissues. It is indispensable to know about the microanatomical relations, in order to identify whether the noxae impairing nephrogenesis targets only the developing nephron or also its covering tissues as interacting and controlling instances.</p>","PeriodicalId":74215,"journal":{"name":"Molecular and cellular pediatrics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2021-08-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8371049/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39319614","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Novel ways to monitor immunosuppression in pediatric kidney transplant recipients-underlying concepts and emerging data.","authors":"Thurid Ahlenstiel-Grunow,&nbsp;Lars Pape","doi":"10.1186/s40348-021-00118-8","DOIUrl":"https://doi.org/10.1186/s40348-021-00118-8","url":null,"abstract":"<p><p>After pediatric kidney transplantation, immunosuppressive therapy is given to avoid acute and chronic rejections. However, the immunosuppression causes an increased risk of severe viral complications and bacterial infections and is associated with serious side effects. It is therefore crucial to achieve the optimal individual balance between over- and under-immunosuppression and thereby avoid unnecessary exposure to immunosuppressive drugs. In routine use, steering of immunosuppressants is performed primarily by monitoring of trough levels that mirror pharmacokinetics (although not, however, pharmacodynamics). Other diagnostic and prognostic markers to assess the individual intensity of immunosuppression are missing. Potential methods to determine immune function and grade of immunosuppression, such as analysis of the torque teno virus (TTV) load, QuantiFERON Monitor®, and ImmuKnow® as well as virus-specific T cells (Tvis), are currently being evaluated. In some studies TTV load, QuantiFERON Monitor® and ImmuKnow® were associated with the risk for post-transplant rejections and infections, but randomized controlled trials after pediatric kidney transplantation are not available. Post-transplant monitoring of Tvis levels seem to be promising because Tvis control virus replication and have been shown to correlate with virus-specific as well as general cellular immune defense, which represents the individual's susceptibility to infections. Additional Tvis-monitoring provides an innovative opportunity to personalize the antiviral management and the dosing of the immunosuppressive therapy after pediatric kidney transplantation to avoid unnecessary therapeutic interventions and identify over-immunosuppression.</p>","PeriodicalId":74215,"journal":{"name":"Molecular and cellular pediatrics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2021-07-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s40348-021-00118-8","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39222964","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Aggressive infantile myofibromatosis with intestinal involvement.","authors":"Tristan Römer,&nbsp;Norbert Wagner,&nbsp;Till Braunschweig,&nbsp;Robert Meyer,&nbsp;Miriam Elbracht,&nbsp;Udo Kontny,&nbsp;Olga Moser","doi":"10.1186/s40348-021-00117-9","DOIUrl":"https://doi.org/10.1186/s40348-021-00117-9","url":null,"abstract":"<p><strong>Background: </strong>Infantile myofibromatosis (IM) is the most common cause of multiple fibrous tumors in infancy. Multicentric disease can be associated with life-threatening visceral lesions. Germline gain-of-function mutations in PDGFRB have been identified as the most common molecular defect in familial IM.</p><p><strong>Case presentation: </strong>We here describe an infant with PDGFRB-driven IM with multiple tumors at different sites, including intestinal polyposis with hematochezia, necessitating temporary chemotherapy.</p><p><strong>Conclusions: </strong>PDGFRB-driven IM is clinically challenging due to its fluctuating course and multiple organ involvement in the first years of life. Early molecular genetic analysis is necessary to consider tyrosine kinase inhibitor treatment in case of aggressive visceral lesions.</p>","PeriodicalId":74215,"journal":{"name":"Molecular and cellular pediatrics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2021-06-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s40348-021-00117-9","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39235683","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Identification of a novel MICU1 nonsense variant causes myopathy with extrapyramidal signs in an Iranian consanguineous family.","authors":"Fatemeh Bitarafan,&nbsp;Mehrnoosh Khodaeian,&nbsp;Elham Amjadi Sardehaei,&nbsp;Fatemeh Zahra Darvishi,&nbsp;Navid Almadani,&nbsp;Yalda Nilipour,&nbsp;Masoud Garshasbi","doi":"10.1186/s40348-021-00116-w","DOIUrl":"https://doi.org/10.1186/s40348-021-00116-w","url":null,"abstract":"<p><strong>Background: </strong>Ca²⁺ as a universal second messenger regulates basic biological functions including cell cycle, cell proliferation, cell differentiation, and cell death. Lack of the protein mitochondrial calcium uptake1 (MICU1), which has been regarded as a gatekeeper of Ca ions, leads to the abnormal mitochondrial Ca²⁺ handling, excessive production of reactive oxygen species (ROS), and increased cell death. Mutations in MICU1 gene causes a very rare neuromuscular disease, myopathy with extrapyramidal signs (MPXPS), due to primary alterations in mitochondrial calcium signaling which demonstrates the key role of mitochondrial Ca²⁺ uptake. To date, 13 variants have been reported in MICU1 gene in 44 patients presented with the vast spectrum of symptoms.</p><p><strong>Case presentation: </strong>Here, we report a 44-year-old Iranian patient presented with learning disability, muscle weakness, easy fatigability, reduced tendon reflexes, ataxia, gait disturbance, elevated hepatic transaminases, elevated serum creatine kinase (CK), and elevated lactate dehydrogenase (LDH). We identified a novel nonsense variant c.385C>T; p.(R129*) in MICU1 gene by whole exome sequencing (WES) and segregation analysis.</p><p><strong>Conclusions: </strong>Our finding along with previous studies provides more evidence on the clinical presentation of the disease caused by pathogenic mutations in MICU1. Finding more variants and expanding the spectrum of the disease increases the diagnostic rate of molecular testing in screening of this kind of diseases and in turn improves the quality of counseling for at risk couples and helps them to minimize the risks of having affected children.</p>","PeriodicalId":74215,"journal":{"name":"Molecular and cellular pediatrics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2021-05-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8107061/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"38976644","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Daratumumab therapy for post-HSCT immune-mediated cytopenia: experiences from two pediatric cases and review of literature.","authors":"Lina Driouk,&nbsp;Robert Schmitt,&nbsp;Anke Peters,&nbsp;Sabine Heine,&nbsp;Hermann Josef Girschick,&nbsp;Brigitte Strahm,&nbsp;Charlotte M Niemeyer,&nbsp;Carsten Speckmann","doi":"10.1186/s40348-021-00114-y","DOIUrl":"https://doi.org/10.1186/s40348-021-00114-y","url":null,"abstract":"<p><strong>Background: </strong>Immune-mediated cytopenias (AIC) are challenging complications following allogeneic hematopoietic stem cell transplantation (HSCT). While broad-acting immunosuppressive agents like corticosteroids are often standard of care, several novel therapies which target specific immunological pathways have recently been developed and provide hope for patients with steroid-refractory courses and may limit long-term toxicity. The successful off-label use of the plasma cell depleting anti-CD38 antibody daratumumab was published in several case reports, suggesting efficacy, i.e., in patients with antibody-mediated AIC refractory to previous B cell depletion. We want to share our experience with two children, whom we treated with daratumumab, including one fatal course with uncontrolled disease. Given the absence of substantial data from HSCT registries or prospective trials, we furthermore provide a critical review of the literature on daratumumab treatment of AIC.</p><p><strong>Case presentations: </strong>Patient 1 (P1), an 11-year-old girl with lipopolysaccharide-responsive and beige-like anchor protein (LRBA) deficiency who developed immune-mediated thrombocytopenia (AIT) from day +58 after HSCT, showed a complete response to daratumumab after the fourth of six total daratumumab doses. She remains transfusion independent for over a year of follow-up. Previously, her thrombocytopenia was refractory to corticosteroids, rituximab, intravenous immunoglobulins (IVIG), eltrombopag, cyclosporine A, and sirolimus. Patient 2 (P2), a 6-year-old boy with CD40 ligand (CD40L) deficiency, developed both AIT and hemolytic anemia (AIHA) after HSCT on days +58 and +83, respectively, and was also treated with daratumumab after being previously refractory to prednisolone, rituximab, and IVIG. Yet, he did neither respond to daratumumab nor the concomitantly administered methyprednisolone pulse, plasmapheresis, and eculizumab and succumbed due to refractory disease.</p><p><strong>Conclusion: </strong>Reviewing the literature on the use of daratumumab for refractory AIC post-HSCT, we consider daratumumab a promising agent for this life-threatening disorder: ten of the twelve patients reached transfusion independency in the literature. However, treatment failures are likely to be underreported. Thus, controlled trials are needed to explore the safety and efficacy of daratumumab in this rare post-HSCT complication.</p>","PeriodicalId":74215,"journal":{"name":"Molecular and cellular pediatrics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2021-04-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s40348-021-00114-y","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"38920477","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Combined RT-qPCR and pyrosequencing of a Spike glycoprotein polybasic cleavage motif can uncover pediatric SARS-CoV-2 infections associated with heterogeneous presentation.","authors":"Patrick Philipp Weil, Jacqueline Hentschel, Frank Schult, Anton Pembaur, Beniam Ghebremedhin, Olivier Mboma, Andreas Heusch, Anna-Christin Reuter, Daniel Müller, Stefan Wirth, Malik Aydin, Andreas C W Jenke, Jan Postberg","doi":"10.1186/s40348-021-00115-x","DOIUrl":"10.1186/s40348-021-00115-x","url":null,"abstract":"<p><strong>Background: </strong>Reverse transcription of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (+)RNA genome and subgenomic RNAs (sgRNAs) and subsequent quantitative polymerase chain reaction (RT-qPCR) is the reliable diagnostic gold standard for COVID-19 diagnosis and the identification of potential spreaders. Apart from clinical relevance and containment, for specific questions, it might be of interest to (re)investigate cases with low SARS-CoV-2 load, where RT-qPCR alone can deliver conflicting results, even though these cases might neither be clinically relevant nor significant for containment measures, because they might probably not be infectious. In order to expand the diagnostic bandwidth for non-routine questions, particularly for the reliable discrimination between negative and false-negative specimens associated with high C_T values, we combined the RT-qPCR workflow with subsequent pyrosequencing of a S-gene amplicon. This expansion can help to confirm SARS-CoV-2 infections without the demand of confirmative antibody testing, which requires to summon patients again for blood sampling few to several weeks after symptom onset.</p><p><strong>Results: </strong>We successfully established a combined RT-qPCR and S-gene pyrosequencing method which can be optionally exploited after routine diagnostics. This allows a reliable interpretation of RT-qPCR results in specimens with relatively low viral loads and close to the detection limits of qPCR. After laboratory implementation, we tested the combined method in a large pediatric cohort from two German medical centers (n=769). Pyrosequencing after RT-qPCR enabled us to uncover 5 previously unrecognized cases of pediatric SARS-CoV-2-associated diseases, mainly exhibiting mild and heterogeneous presentation-apart from a single case of multisystem inflammatory syndrome in children (MIS-C) associated with SARS-CoV-2, who was hospitalized in the course of the study.</p><p><strong>Conclusions: </strong>The proposed protocol allows a specific and sensitive confirmation of SARS-CoV-2 infections close to the detection limits of RT-qPCR. The tested biotinylated primers do not negatively affect the RT-qPCR pipeline and thus can be optionally applied to enable deeper inspection of RT-qPCR results by subsequent pyrosequencing. Moreover, due to the incremental transmission of SARS-CoV-2 variants of concern, we note that the used strategy can uncover (Spike) P681H allowing the pre-selection of SARS-CoV-2 B.1.1.7 candidate specimens for deep sequencing.</p>","PeriodicalId":74215,"journal":{"name":"Molecular and cellular pediatrics","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2021-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s40348-021-00115-x","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"38824990","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}