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
D fluorometer. The MS/MS method was performed using a NeoBase
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.
In the last two decades, the introduction of tandem mass spectrometry in clinical laboratories has enabled simultaneous testing of numerous acylcarnitines and amino acids from dried blood spots for ...detecting many aminoacidopathies, organic acidurias and fatty acid oxidation disorders. The expanded newborn screening was introduced in Slovenia in September 2018. Seventeen metabolic diseases have been added to the pre-existing screening panel for congenital hypothyroidism and phenylketonuria, and the newborn screening program was substantially reorganized and upgraded.
Tandem mass spectrometry was used for the screening of dried blood spot samples. Next-generation sequencing was introduced for confirmatory testing. Existing heterogeneous hospital information systems were connected to the same laboratory information system to allow barcode identification of samples, creating reports, and providing information necessary for interpreting the results.
In t he first y ear of t he expanded newborn screening a total of 15,064 samples w ere screened. Four patients were confirmed positive with additional testing.
An expanded newborn screening program was successfully implemented with the first patients diagnosed before severe clinical consequences.
Presejalno testiranje novorojenčkov (PTN) je ena svetovnih javnozdravstvenih zgodb o uspehu. Izvaja se v veliki večini razvitih držav sveta, razlikuje pa se le po obsegu in usmeritvah testiranja. ...Najbolj obsežni programi presejalnega testiranja na svetu obsegajo testiranje na več kot 50 različnih bolezni. PTN v Sloveniji je ključni del javnozdravstvenega sistema s pričetkom v letu 1979 s testiranjem na fenilketonurijo. V letu 1981 se je dodalo testiranje na kongenitalno hipotirozo. Program se je razvijal in širil z uvedbo novih tehnologij, kot so masna spektrometrija za bolezni presnove in imunoreaktivni ter genetski testi za bolezni, kot so cistična fibroza, kongenitalna adrenalna hiperplazija, spinalna mišična atrofija in primarne imunske pomanjkljivosti. Odvzem vzorca, posušen madež krvi na filter papirju, omogoča enostaven transport v laboratorij Pediatrične klinike v Ljubljani, kjer poteka analiza. Strokovno izvedeni odvzem, ki ga opravi osebje porodnišnic, zagotavlja ustrezno kakovost vzorcev in je pogoj za uspešno izvedbo testiranja. Vključenost v program je univerzalna za vse novorojenčke, z možnostjo zavrnitve s strani staršev. Do danes smo v Sloveniji uspešno odkrili in zdravili več 100 novorojenčkov, kar potrjuje njegov pomen. Z odkritjem novih zdravil in terapij bo obseg PTN naraščal. Uspešnost programa pa je neposredno odvisna od sinergije sodelujočih skupin – porodnišnic, pediatrične klinike in laboratorija.
As children spend up to 9 h a day in kindergarten, the main purpose of our study was to evaluate the effect of antioxidant-rich kindergarten meals on oxidative stress biomarkers (OSBs) in healthy ...children. In the randomized control trial with a follow-up, healthy 5–6-year-old children from six kindergartens were randomly divided into a prototype group (PG,
n
= 40) and a control group (CG,
n
= 17). PG followed a 2-week antioxidant-rich kindergarten meal plan (breakfast, lunch, and two snacks), and CG followed their standard kindergarten meal plans. Outside the kindergartens, participants ate as usual. We used a consecutive 7-day dietary record inside and outside the kindergarten and the national dietary assessment tool OPEN to assess the total dietary antioxidant capacity (dTAC) of the consumed foods. Malondialdehyde (MDA), 8-hydroxy-2-deoxyguanosine (8-OHdG), and four F2-isoprostane were measured in fasting urine on days 1 and 15. We also measured total antioxidant power (PAT) and hydroperoxides (d-ROMs) in fasting serum on day 15 and obtained the value of the oxidative stress index (OSI). We used a Welch two-sample
t
-test and multiple regression analysis to compare the prototype and control groups and a nonparametric Wilcoxon signed rank exact test to compare pre- and post-intervention results in urine. Antioxidant-rich kindergarten meals contributed to a significantly (
p
< 0.05) higher intake of dTAC in PG participants compared to standard meals in CG participants (8.6 vs. 2.8 mmol/day). We detected a negative correlation between dTAC intake and d-ROMs and between dTAC intake and OSI (
r
= − 0.29,
p
= 0.043 and
r
= − 0.31,
p
= 0.032, respectively). A significant decrease in urinary 8-iso-15-prostaglandin-F-2 alpha was detected in PG participants between days 1 and 15; however, no other intra-individual significant differences in urinary OSBs were found.
Conclusion
: Antioxidant-rich food in kindergarten is warranted due to its potential health-protective effect. Additionally, we present original data on the average levels of urinary and serum OSBs in healthy 5–6-year-old children.
Trial registration
: The study was registered at ClinicalTrials.gov, on February 5, 2020 (
https://clinicaltrials.gov/ct2/show/NCT04252105
).
What is Known:
• Kindergartens are recognized as promising environments for public health measures.
• A diet rich in antioxidants can reduce OSBs and, consequently, the risk of developing NCDs.
What is New:
• Antioxidant-rich kindergarten diet can ensure a protective intake of dTAC in children.
• Original data on serum oxidative stress biomarkers (d-ROMs, PAT, and OSI) and urinary oxidative stress biomarkers (MDA, 8-OHdG, and F2 isoprostanes) in healthy 5–6-year-old children.
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.
Newborn Screening in a Pandemic-Lessons Learned Mlinaric, Matej; Bonham, James R; Kožich, Viktor ...
International journal of neonatal screening,
04/2023, Letnik:
9, Številka:
2
Journal Article
Recenzirano
Odprti dostop
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.
Significant part of Southeastern Europe (with a population of 76 million) has newborn screening (NBS) programs non-harmonized with developed European countries. Initial survey was conducted in ...2013/2014 among 11 countries from the region (Albania, Bulgaria, Bosnia and Herzegovina (BIH), Croatia, Kosovo, Macedonia, Moldova, Montenegro, Romania, Serbia, and Slovenia) to assess the main characteristics of their NBS programs and their future plans. Their cumulative population at that time was ~52,5 million. At that time, none of the countries had an expanded NBS program, while phenylketonuria screening was not introduced in four and congenital hypothyroidism in three of 11 countries. We repeated the survey in 2020 inviting the same 11 countries, adding Cyprus, Greece, Hungary, and Malta (due to their geographical position in the wider region). The aims were to assess the current state, to evaluate the change in the period, and to identify the main obstacles impacting the implementation of expanded NBS and/or reaching a wider population. Responses were collected from 12 countries (BIH—Federation of BIH, BIH—Republic of Srpska, Bulgaria, Croatia, Greece, Hungary, Kosovo, North Macedonia, Malta, Montenegro, Romania, Serbia, Slovenia) with a population of 68.5 million. The results of the survey showed that the regional situation regarding NBS only modestly improved in this period. All of the surveyed countries except Kosovo screened for at least congenital hypothyroidism, while phenylketonuria was not screened in four of 12 countries. Croatia and Slovenia implemented an expanded NBS program using tandem mass spectrometry from the time of last survey. In conclusion, the current status of NBS programs in Southeastern Europe is very variable and is still underdeveloped (or even non-existent) in some of the countries. We suggest establishing an international task-force to assist with implementation and harmonization of basic NBS services where needed.
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.
We presented the follow up of the only two Slovenian patients diagnosed with HT1. Metabolic control was monitored by measuring tyrosine, phenylalanine and succinylacetone from dried blood spots (DBSs). Retrograde screening of HT1 was performed from DBSs taken at birth using tandem mass spectrometry.
First patient was diagnosed at the age of 6 months in the asymptomatic phase due to an abnormal liver echogenicity, the other presented at 2.5 months with an acute liver failure and needed a liver transplantation. The first was a compound heterozygote for a novel FAH intronic variant c.607-21A>G and c.192G>T whereas the second was homozygous for c.192G>T. At the non-transplanted patient, 66% of tyrosine and 79% of phenylalanine measurements were in strict reference ranges of 200–400 μmol/L and >30 μmol/L, respectively, which resulted in a favorable cognitive outcome at 3.6 years. On retrograde screening, both patients had elevated SA levels; on the other hand, tyrosine was elevated only at one.
We showed that non-coding regions should be analyzed when clinical and biochemical markers are characteristic of HT1. DBSs represent a convenient sample type for frequent amino acid monitoring. Retrograde diagnosis of HT1 was possible after more than three years of birth with SA as a primary marker, complemented by tyrosine.
•Non-coding region variants of FAH gene can result in a symptomatic HT1.•Retrograde screening for HT1 is technically possible even three years after birth.•DBS are convenient for monitoring HT1 patients and are family-friendly.•Regular monitoring in HT1 patients can result in a favorable cognitive outcome.
NEWBORN SCREENING PROGRAM IN SLOVENIA Jernej Kovač; Barbka Repič Lampret; Žiga Iztok Remec ...
Slovenska pediatrija,
05/2024, Letnik:
31, Številka:
2
Journal Article
Recenzirano
Odprti dostop
Newborn Screening (NBS) is one of the most significant and successful public health initiatives globally. Implemented in various capacities across most developed countries, NBS programs vary in scope ...and focus, with the most comprehensive screening for over 50 conditions. In Slovenia, NBS is a vital component of the public health system, initiated in 1979 with screenings for phenylketonuria. The program has evolved and expanded with the introduction of new technologies such as mass spectrometry for metabolic diseases and immunoreactive and genetic tests for conditions like cystic fibrosis, spinal muscular atrophy, and primary immune deficiencies. Dried blood spots on filter paper facilitate easy transportation to the University Children’s Hospital Ljubljana for the analysis. Professional sample collection by maternity ward staff ensures sample quality, which is critical for successful laboratory testing. Participation in the program is universal for all newborns, with an option for parental refusal. Hundreds of Slovenian newborns have been successfully diagnosed and treated, underscoring the program’s importance. With the advent of new medicines and therapies, the scope of NBS is set to expand. The program’s success relies on the synergy between participating groups - maternity hospitals, the pediatric clinic, and the laboratory.
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