Balkan Journal of Medical Genetics

TWO YEARS OF NEWBORN SCREENING FOR CYSTIC FIBROSIS IN NORTH MACEDONIA: FIRST EXPERIENCE
Fustik S1,*, Anastasovska V2, Plaseska-Karanfilska D3, Stamatova A1, Spirevska L1, Pesevska M2, Terzikj M3, Vujovic M3
*Corresponding Author: Professor Stojka Fustik, M.D., Ph.D., Department for Cystic Fibrosis, University Clinic for Pediatrics, Faculty of Medicine, University “Ss. Cyril and Methodius,” Vodnjanska 17, 1000 Skopje, Republic of North Macedonia. Tel.: +389-23-147-716; +389-75-705-369. Fax: +389- 23-129-027. E-mail: stojkaf@yahoo.com
page: 41

MATERIALS AND METHODS

A two-step IRT-IRT algorithm is performed, and then a sweat test for confirmation/exclusion of the CF diagnosis when IRT values were both over the cutoff: 70.0 and 45.0 ng/mL, respectively (Figure 1). Heel prick blood samples are taken by a trained nurse, between 24 and 72 hours of life (when the baby is discharged from the maternity hospital) and spotted on Whatman 903 filter paper (Guthrie card). Premature or sick full-term neonates, who usually have a prolonged stay in neonatal intensive care units, are screened between the first and second week of life. The whole-blood samples are dried at room temperature within 2 hours and transported or mailed to the laboratory for neonatal screening twice per week from each birth center. The whole-blood IRT is measured by the DELFIA® time-resolved fluoroimmunoassay method, using a DELFIA® neonatal IRT kit (DELFIA®; PerkinElmer, Wallac Oy, Turku, Finland), and read by a 1420 VICTOR 2D Fluorometer (Wallac Oy, Turku, Finland). The IRT1 cutoff level of 70.0 ng/mL was established as 99.5th percentile of the IRT values obtained in a total of 2.058 newborns with birth weight >3000 g, gestational age >38 weeks, and age >48 hours. The second step in the algorithm is based on request of a second blood specimen from those neonates with elevated IRT-1, 3 weeks after the first screening. For IRT-2, a fixed cutoff level is used at 45.0 ng/mL. If the IRT-2 is over the cutoff value, the positive-screened child is referred for sweat testing. A doctor from the CF team meets the parents/baby before performing the sweat test and provide explanations about the screening results and the sweat test. The sweat test is performed according to a standardized protocol via quantitative pilocarpine iontophoresis with the Macroduct® collection method. Sweat chloride concentration is measured in a ChloroChek® Chloridometer® (ELITechGroup, Puteaux, France). The sweat test is analyzed immediately and the result is reported to the family on the same day. Based on the recommendations for sweat chloride test results in healthy and CF-affected infants [2,6,7], the following sweat chloride reference ranges are use: <29.0 mmol/L, CF rejected or unlikely; 30.0 to 59.0 mmol/L, intermediate; >60.0 mmol/L, consistent with a diagnosis of CF. Infants with a positive sweat test result are referred to the specialist CF doctor as soon as possible. A CF doctor informs the family about positive CF diagnosis, gives them information about the disease, the benefit of early diagnosis and treatment for the child’s health and arranges admission to the CF Unit for a comprehensive assessment of the child, introduction of therapy and parent training for chest physiotherapy. Parental written informed consent for CFTR mutation analysis is required at the first visit. DNA is isolated from peripheral blood using the standard phenol-chloroform extraction method. Mutation analysis of the CFTR gene is performed initially by SNaPshotreaction [8] for 11 most common regional CFTR mutations (F508del, G542X, N1303K, 621+1G>T, 2184insA, V456F, G126D, G1349D, E822K, R117C, 711+3A>G). For the patients whose genotypes were not determined with the initial molecular screening, several additional methods are applied, including multiplex ligation- dependent probe amplification (MLPA), for detection of deletions/duplications in the CFTR gene (SALSA MLPA KIT P091 CFTR; MRC-Holland, Amsterdam, The Netherlands); next generation sequencing (NGS), with TruSight Inherited gene panel performed on MiSeq Illumina Personal Sequencer and data analyses on the Illumina Variant Studio (Illumina Inc., San Diego, CA, USA) [8,9].

Number 27 VOL. 27 (2), 2024	Number 27 VOL. 27 (1), 2024
Number 26 Number 26 VOL. 26(2), 2023 All in one	Number 26 VOL. 26(2), 2023
Number 26 VOL. 26, 2023 Supplement	Number 26 VOL. 26(1), 2023
Number 25 VOL. 25(2), 2022	Number 25 VOL. 25 (1), 2022
Number 24 VOL. 24(2), 2021	Number 24 VOL. 24(1), 2021
Number 23 VOL. 23(2), 2020	Number 22 VOL. 22(2), 2019
Number 22 VOL. 22(1), 2019	Number 22 VOL. 22, 2019 Supplement
Number 21 VOL. 21(2), 2018	Number 21 VOL. 21 (1), 2018
Number 21 VOL. 21, 2018 Supplement	Number 20 VOL. 20 (2), 2017
Number 20 VOL. 20 (1), 2017	Number 19 VOL. 19 (2), 2016
Number 19 VOL. 19 (1), 2016	Number 18 VOL. 18 (2), 2015
Number 18 VOL. 18 (1), 2015	Number 17 VOL. 17 (2), 2014
Number 17 VOL. 17 (1), 2014	Number 16 VOL. 16 (2), 2013
Number 16 VOL. 16 (1), 2013	Number 15 VOL. 15 (2), 2012
Number 15 VOL. 15, 2012 Supplement	Number 15 Vol. 15 (1), 2012
Number 14 14 - Vol. 14 (2), 2011	Number 14 The 9th Balkan Congress of Medical Genetics
Number 14 14 - Vol. 14 (1), 2011	Number 13 Vol. 13 (2), 2010
Number 13 Vol.13 (1), 2010	Number 12 Vol.12 (2), 2009
Number 12 Vol.12 (1), 2009	Number 11 Vol.11 (2),2008
Number 11 Vol.11 (1),2008	Number 10 Vol.10 (2), 2007
Number 10 10 (1),2007	Number 9 1&2, 2006
Number 9 3&4, 2006	Number 8 1&2, 2005
Number 8 3&4, 2004	Number 7 1&2, 2004
Number 6 3&4, 2003	Number 6 1&2, 2003
Number 5 3&4, 2002	Number 5 1&2, 2002
Number 4 Vol.3 (4), 2000	Number 4 Vol.2 (4), 1999
Number 4 Vol.1 (4), 1998	Number 4 3&4, 2001
Number 4 1&2, 2001	Number 3 Vol.3 (3), 2000
Number 3 Vol.2 (3), 1999	Number 3 Vol.1 (3), 1998
Number 2 Vol.3(2), 2000	Number 2 Vol.1 (2), 1998
Number 2 Vol.2 (2), 1999	Number 1 Vol.3 (1), 2000
Number 1 Vol.2 (1), 1999	Number 1 Vol.1 (1), 1998

About the journal ::: Editorial ::: Subscription ::: Information for authors ::: Contact