Phenylketonuria (PKU) was the first disease to be identified by the newborn screening (NBS) program. Currently, there are various methods for determining phenylalanine (Phe) values, with tandem mass spectrometry (MS/MS) being the most widely used method worldwide. We aimed to compare the MS/MS method with the fluorometric method (FM) for measuring Phe in the dried blood spot (DBS) and the efficacy of both methods in the NBS program. The FM was performed using a neonatal phenylalanine kit and a VICTOR2^TM D fluorometer. The MS/MS method was performed using a NeoBase^TM 2 kit and a Waters Xevo TQD mass spectrometer. The Phe values measured with the MS/MS method were compared to those determined by the FM. The cut-off value for the NBS program was set at 120 µmol/L for FM and 85 µmol/L for MS/MS. We analyzed 54,934 DBS. The measured Phe values varied from 12 to 664 µmol/L, with a median of 46 µmol/L for the MS/MS method and from 10 to 710 µmol/L, with a median of 70 µmol/L for the FM. The Bland–Altman analysis indicated a bias of −38.9% (−23.61 µmol/L) with an SD of 21.3% (13.89 µmol/L) when comparing the MS/MS method to the FM. The Phe value exceeded the cut-off in 187 samples measured with FM and 112 samples measured with MS/MS. The FM had 181 false positives, while the MS/MS method had 106 false positives. Our study showed that the MS/MS method gives lower results compared to the FM. Despite that, none of the true positives would be missed, and the number of false-positive results would be significantly lower compared to the FM.

Newborn screening for Phenylketonuria enables early detection and timely treatment with a phenylalanine-restricted diet to prevent severe neurological impairment. Although effective and in use for 60 years, screening, diagnostic, and treatment practices still vary widely across countries and centers. To evaluate the Phenylketonuria newborn screening practices internationally, we designed a survey with questions focusing on the laboratory aspect of the screening system. We analyzed 24 completed surveys from 23 countries. Most participants used the same sampling age range of 48–72 h; they used tandem mass spectrometry and commercial non-derivatized kits to measure phenylalanine (Phe), and had non-negative cut-off values (COV) set mostly at 120 µmol/L of Phe. Participants mostly used genetic analysis of blood and detailed amino acid analysis from blood plasma as their confirmatory methods and set the COV for the initiation of dietary therapy at 360 µmol/L of Phe. There were striking differences in practice as well. While most participants reported a 48–72 h range for age at sampling, that range was overall quite diverse Screening COV varied as well. Additional screening parameters, e.g., the phenylalanine/tyrosine ratio were used by some participants to determine the screening result. Some participants included testing for tetrahydrobiopterin deficiency, or galactosemia in their diagnostic process. Results together showed that there is room to select a best practice from the many practices applied. Such a best practice of PKU-NBS parameters and post-screening parameters could then serve as a generally applicable guideline.

Phenylketonuria (PKU) was the first disorder for which newborn screening (NBS) was introduced in the early 1960s. Slovenia started the NBS program for PKU in 1979, and the fluorimetric method was implemented in 1992, with a phenylalanine (Phe) cut-off set at 120 mol/L. This value has been in use for almost thirty years and has never been revised. We aimed to analyze the DBS samples and review the data from a large nationwide cohort of newborns to optimize the cut-off values for HFA screening to minimize the number of false positives while maintaining the highest level of sensitivity by detecting all those who needed to be treated. In the first prospective part of the study, we analyzed samples of all newborns in Slovenia in 2019 and 2020, and in the second retrospective part, we reviewed data from all known patients with hyperphenylalaninemia (HFA) in Slovenia born from 2000 to 2018. We defined true screening-positive cases as those that required a low-Phe diet. The sensitivity, specificity and positive predictive values of the modeling elevation of the Phe cut-off value from 120 µmol/L to 200 µmol/L were assessed. The number of recalls at the cut-off of 120 µmol/L was 108 out of 37,784 samples at NBS (2019–2020). Six newborns were defined as true positives and 102 samples as false positives. If the cut-off value was adjusted to 160 µmol/L, only 12 samples exceeded it and all six true positive newborns would be detected. Among the 360,000 samples collected at the NBS between 2000 and 2018, 72 HFA patients in need of a low-Phe diet were found. All the diagnosed cases would have been detected if the cut-off was set to 160 µmol/L. We demonstrated in a large group of newborns (400,000 in 20 years) that using the fluorimetric approach, a cut-off value of 160 µmol/L, rather than 120 mol/L, is safe and that there were no missing true positive patients who required treatment. By increasing the cut-off, this method becomes more precise, resulting in a significantly reduced rate of false positives and thus being less burdensome on both families and the healthcare system.

Introduction: Glycogen storage disease type 1b (GSD-1b) is characterized by neutropenia and neutrophil dysfunction generated by the accumulation of 1,5-anhydroglucitol-6-phosphate in neutrophils. Sodium-glucose co-transporter 2 inhibitors, such as empagliflozin, facilitate the removal of this toxic metabolite and ameliorate neutropenia-related symptoms, including severe infections and inflammatory bowel disease (IBD). Our case series presents the treatment of three pediatric GSD-1b patients with empagliflozin over a follow-up of three years; the most extended reported follow-up period to date.Cases description: A retrospective analysis of empagliflozin treatment of three pediatric GSD-1b patients (two male and one female; ages at treatment initiation: 4.5, 2.5 and 6 years) was performed. Clinical and laboratory data from a symmetrical period of up to three years before and after the therapy introduction was reported. Data on the clinical course of the treatment, IBD activity, the need for antibiotic treatment and hospitalizations, neutrophil count and function, and markers of inflammation were assessed. Prior the introduction of empagliflozin, patients had recurrent oral mucosa lesions and infections, abdominal pain, and anemia. During empagliflozin treatment, the resolution of aphthous stomatitis, termination of abdominal pain, reduced frequency and severity of infections, anemia resolution, increased appetite, and improved wound healing was observed in all patients, as well as an increased body mass index in two of them. In a patient with IBD, long-term deep remission was confirmed. An increased and stabilized neutrophil count and an improved neutrophil function enabled the discontinuation of G-CSF treatment in all patients. A trend of decreasing inflammation markers was detected.Conclusions: During the three-year follow-up period, empagliflozin treatment significantly improved clinical symptoms and increased the neutrophil count and function, suggesting that targeted metabolic treatment could improve the immune function in GSD-1b patients.

Tyrosinemia type 1 (HT1) is an inborn error of tyrosine catabolism that leads to severe liver, kidney, and neurological dysfunction. Newborn screening (NBS) can enable a timely diagnosis and early initiation of treatment.

The COVID-19 pandemic affected many essential aspects of public health, including newborn screening programs (NBS). Centers reported missing cases of inherited metabolic disease as a consequence of decreased diagnostic process quality during the pandemic. A number of problems emerged at the start of the pandemic, but from the beginning, solutions began to be proposed and implemented. Contingency plans were arranged, and these are reviewed and described in this article. Staff shortage emerged as an important issue, and as a result, new work schedules had to be implemented. The importance of personal protective equipment and social distancing also helped avoid disruption. Staff became stressed, and this needed to be addressed. The timeframe for collecting bloodspot samples was adapted in some cases, requiring reference ranges to be modified. A shortage of essential supplies and protective equipment was evident, and laboratories described sharing resources in some situations. The courier system had to be adapted to make timely and safe transport possible. Telemedicine became an essential tool to enable communication with patients, parents, and medical staff. Despite these difficulties, with adaptations and modifications, some centers evaluated candidate conditions, continued developments, or began new NBS. The pandemic can be regarded as a stress test of the NBS under real-world conditions, highlighting critical aspects of this multidisciplinary system and the need for establishing local, national, and global strategies to improve its robustness and reliability in times of shortage and overloaded national healthcare systems.