Newborn screening (NBS) for inborn errors of metabolism is one of the most advanced tools for secondary prevention in medicine, as it allows early diagnosis and prompt treatment initiation. The expanded newborn screening was introduced in Italy between 2016 and 2017 (Law 167/2016; DM 13 October 2016; DPCM 12-1-2017). A total of 1,586,578 infants born in Italy were screened between January 2017 and December 2020. For this survey, we collected data from 15 Italian screening laboratories, focusing on the metabolic disorders identified by tandem mass spectrometry (MS/MS) based analysis between January 2019 and December 2020. Aminoacidemias were the most common inborn errors in Italy, and an equal percentage was observed in detecting organic acidemias and mitochondrial fatty acids beta-oxidation defects. Second-tier tests are widely used in most laboratories to reduce false positives. For example, second-tier tests for methylmalonic acid and homocysteine considerably improved the screening of CblC without increasing unnecessary recalls. Finally, the newborn screening allowed us to identify conditions that are mainly secondary to a maternal deficiency. We describe the goals reached since the introduction of the screening in Italy by exchanging knowledge and experiences among the laboratories.

Niemann-Pick type C (NPC) disease is an autosomal recessive lysosomal storage disorder caused by mutations in NPC1 or NPC2 genes. In 2009, the molecular characterization of 44 NPC Italian patients has been published. Here, we present an update of the genetic findings in 105 Italian NPC patients belonging to 83 unrelated families (77 NPC1 and 6 NPC2). NPC1 and NPC2 genes were studied following an algorithm recently published. Eighty-four different NPC1 and five NPC2 alleles were identified. Only two NPC1 alleles remained non detected. Sixty-two percent of NPC1 alleles were due to missense variants. The most frequent NPC1 mutation was the p.F284Lfs*26 (5.8% of the alleles). All NPC2 mutations were found in the homozygous state, and all but one was severe. Among newly diagnosed patients, 18 novel NPC1 mutations were identified. The pathogenic nature of 7/9 missense alleles and 3/4 intronic variants was confirmed by filipin staining and NPC1 protein analysis or mRNA expression in patient’s fibroblasts. Taken together, our previous published data and new results provide an overall picture of the molecular characteristics of NPC patients diagnosed so far in Italy.

Mitochondrial fatty acid oxidation disorders (FAODs) are a heterogeneous group of hereditary autosomal recessive diseases included in newborn screening (NBS) program in Italy. The aim of this study was to analyse FAODs cases, identified either clinically or by NBS,for clinical and genetic characterization and to evaluate a five years' experience of NBS, in the attempt to figure out the complexity of genotype-phenotype correlation and to confirm the clinical impact of NBS in our centre experience. We analysed FAODs patients diagnosed either by NBS or clinically, followed since February 2014 to April 2019 at the Regional Screening Centre and Inherited Metabolic Diseases Unit of Verona. Diagnosis was confirmed by plasma acylcarnitines, urinary organic acids, enzymatic and genetic testing. For not clear genotypes due to the presence of variants of uncertain significance, in silico predictive tools have been used as well as enzymatic activity assays. Patients underwent clinical, nutritional and biochemical follow up. We diagnosed 30 patients with FAODs. 20 by NBS: 3 CUD, 6 SCADD, 5 MCADD, 4 VLCADD, 2 MADD. Overall incidence of FAODs diagnosed by NBS was 1:4316 newborns. No one reported complications during the follow up period. 10 patients were diagnosed clinically: 2 CUD, 2 CPT2D, 1 VLCADD, 5 MADD. Mean age at diagnosis was 29.3 years. Within this group, complications or symptoms were reported at diagnosis, but not during follow-up. 12 mutations not previously reported in literature were found, all predicted as pathogenic or likely pathogenic. Our study highlighted the great phenotypic variability and molecular heterogeneity of FAODs and confirmed the importance of a tailored follow up and treatment. Despite the short duration of follow up, early identification by NBS prevented diseases related complications and resulted in normal growth and psycho-motor development as well.

Introduction: Biotinidase deficiency (BD) is an autosomal recessive disease causing a defect in the biotin-releasing enzyme. Newborn screening (NBS) allows early diagnosis and treatment, ensuring excellent prognosis. The aim of this study was to describe our experience in the diagnosis, treatment, and follow-up showing key strategies and unsolved questions of the management of BD patients.Methods: We analyzed data of patients identified by the Regional Centre for Newborn Screening of Verona and followed by the Inherited Metabolic Disease Unit of Verona and Neonatal Intensive Care Unit of Bolzano, Italy, from 2014 to 2020.Results: Thirty-seven patients were diagnosed by NBS (five profound and 32 partial BD), with a total incidence of 1:5,996. All were started on biotin at diagnosis and presented no symptoms at follow-up. Analysis of parents and siblings led to identification of five asymptomatic patients with partial BD: one asymptomatic parent and four young siblings. Genetic analysis of the BTD gene identified 17 different genotypes and one mutation not previously known.Discussion: Our data confirm that NBS introduction had a dramatic impact on BD diagnosis, and the incidence has increased significantly compared to other areas. Partial defects are more common than profound and have a distinctive genotype. Partial BD treatment is still controversial even at what dose of biotin and for how long. At the end, BD treatment is very easy and inexpensive and prevents severe neurological damage. Sharing experiences is essential to achieving guidelines for treatment and follow-up and a better genotype–phenotype correlation.

Recent studies suggest the efficacy of behavioral therapy for enuresis, even in cases of minor daytime voiding problems. We describe our experience with the clinical followup and behavioral therapy of children with primary enuresis. We followed 159 boys and 91 girls 5 to 17 years old with primary enuresis who were treated at 3 medical centers with a pediatric nephrology clinic during the last 3 years. A detailed voiding history was obtained. Each child was treated with a bladder training session, including an explanation of the enuretic process, daily diary recording and training to recognize bladder distention and increase voiding frequency. A total of 226 children (90%) presented with 1 or more symptoms of bladder maturation delay and 13% reported behavioral constipation. Of the patients 185 (74%) completed the proposed treatment, including 111 (60%) who reported a positive and 21 (11%) who reported a partial response. In 53 children (29%) the treatment failed. Most children with enuresis have daytime symptoms when an accurate history is recorded. As shown by our data, the efficacy of behavioral therapy is comparable to that of desmopressin or alarm therapy but it requires good compliance of the child with the therapeutic plan. Age is not a determining factor in the success rate.

BCKDK is an important key regulator of branched-chain ketoacid dehydrogenase complex activity by phosphorylating and so inactivating branched-chain ketoacid dehydrogenases, the rate-limiting enzyme of the branched-chain amino acid metabolism. We identified, by whole exome-sequencing analysis, the p.His162Gln variant of the BCKDK gene in a neonate, picked up by newborn screening, with a biochemical phenotype of a mild form of maple syrup urine disease (MSUD). The same biochemical and genetic picture was present in the father. Computational analysis of the mutation was performed to better understand its role. Extensive atomistic molecular dynamics simulations showed that the described mutation leads to a conformational change of the BCKDK protein, which reduces the effect of inhibitory binding bound to the protein itself, resulting in its increased activity with subsequent inactivation of BCKDC and increased plasmatic branched-chain amino acid levels. Our study describes the first evidence of the involvement of the BCKDK gene in a mild form of MSUD. Although further data are needed to elucidate the clinical relevance of the phenotype caused by this variant, awareness of this regulatory activation of BCKDK is very important, especially in newborn screening data interpretation.