Neonatal Screening in Europe Revisited: An ISNS Perspective on the Current State and Developments Since 2010.

Neonatal screening (NBS) was initiated in Europe during the 1960s with the screening for phenylketonuria. The panel of screened disorders ("conditions") then gradually expanded, with a boost in the late 1990s with the introduction of tandem mass spectrometry (MS/MS), making it possible to screen for 40-50 conditions using a single blood spot. The most recent additions to screening programmes (screening for cystic fibrosis, severe combined immunodeficiency and spinal muscular atrophy) were assisted by or realised through the introduction of molecular technologies. For this survey, we collected data from 51 European countries. We report the developments between 2010 and 2020 and highlight the achievements reached with the progress made in this period. We also identify areas where further progress can be made, mainly by exchanging knowledge and learning from experiences in neighbouring countries. Between 2010 and 2020, most NBS programmes in geographical Europe matured considerably, both in terms of methodology (modernised) and with regard to the panel of conditions screened (expanded). These developments indicate that more collaboration in Europe through European organisations is gaining momentum. We can only accomplish the timely detection of newborn infants potentially suffering from one of the many rare diseases and take appropriate action by working together.

A dual colour FISH method for routine validation of sexed Bos taurus semen.

BACKGROUND: Usage of sexed semen that allows to choose the gender of the calves, is commonly practiced in livestock industry as a profitable breeding alternative, especially in dairy farming. The flow cytometric cell sorting is the only commercially available method for bovine sperm sexing. For validation of the sexing procedure several methods have been developed including sperm fluorescence in situ hybridisation techniques. Latter usually include the use of pre-labelled nucleotides for probe synthesis which is relatively expensive approach compared to combined application of aminoallyl-dUTP and chemical binding of fluorescent dyes. Here a sex determining dual colour bovine sperm fluorescence in situ hybridisation method is presented which is considered more cost-effective technique than the previously reported approaches. RESULTS: The reliability of sex chromosome identifying probes, designed in silico, was proven on bovine metaphase plate chromosomes and through comparison with a commercially available standard method. In the dual colour FISH experiments of unsexed and sexed bovine sperm samples the hybridisation efficiency was at least 98%, whereas the determined sex ratios were not statistically different from the expected. Very few cells carried both of the sex chromosome-specific signals (less than 0.2%). CONCLUSIONS: A protocol for a dual colour bovine sperm FISH method is provided which is cost-effective, simple and fast for sex determination of spermatozoa in bull semen samples.

Epitope of titin A-band-specific monoclonal antibody Tit1 5 H1.1 is highly conserved in several Fn3 domains of the titin molecule. Centriole staining in human, mouse and zebrafish cells.

BACKGROUND: Previously we have reported on the development of a new mouse anti-titin monoclonal antibody, named MAb Titl 5 H1.1, using the synthetic peptide N-AVNKYGIGEPLESDSVVAK-C which corresponds to an amino acid sequence in the A-region of the titin molecule as immunogen. In the human skeletal muscles, MAb Titl 5 H1.1 reacts specifically with titin in the A-band of the sarcomere and in different non-muscle cell types with nucleus and cytoplasm, including centrioles. In this report we have studied the evolutionary aspects of the binding of MAb Tit1 5 H1.1 with its target antigen (titin). RESULTS: We have specified the epitope area of MAb Tit1 5 H1.1 by subpeptide mapping to the hexapeptide N-AVNKYG-C. According to protein databases this amino acid sequence is located in the COOH-terminus of several different Fn3 domains of the A-region of titin molecule in many organisms, such as human being, mouse, rabbit, zebrafish (Danio rerio), and even in sea squirt (Ciona intestinalis). Our immunohisto- and cytochemical studies with MAb Tit1 5 H1.1 in human, mouse and zebrafish tissues and cell cultures showed a striated staining pattern in muscle cells and also staining of centrioles, cytoplasm and nuclei in non-muscle cells. CONCLUSIONS: The data confirm that titin can play, in addition to the known roles in striated muscle cells also an important role in non-muscle cells as a centriole associated protein. This phenomenon is highly conserved in the evolution and is related to Fn3 domains of the titin molecule. Using titin A-band-specific monoclonal antibody MAb Tit1 5 H1.1 it was possible to locate titin in the sarcomeres of skeletal muscle cells and in the centrioles, cytoplasm and nuclei of non-muscle cells in phylogenetically so distant organisms as Homo sapiens, Mus musculus and zebrafish (Danio rerio).

Balanced reciprocal translocation t(5;13)(q33;q12) and 9q31.1 microduplication in a man suffering from infertility and pollinosis.

We describe the first case of two chromosomal abnormalities, balanced reciprocal translocation t(5;13)(q33;q12.1) and a microduplication in the region 9q31.1, in a man suffering from infertility and pollinosis. In the region 13q12.1 is located the TUBA3C (tubulin, alpha 3c) gene, which plays an important dynamic role in the motility of flagella. This case might support the opinion that haploinsufficiency of the TUBA3C gene could be the cause of sperm immotility and abnormal sperm morphology, resulting in infertility in the patient. Single-nucleotide polymorphism (SNP) array analysis revealed a novel 9q31.1 microduplication inherited from both parents, which contributes to the genomic instability.

Small supernumerary marker chromosome (sSMC) derived from chromosome 22 in an infertile man with hypogonadotropic hypogonadism.

We describe the first case of a supernumerary inv dup(22)(q11.1) in an infertile male with hypogonadotropic hypogonadism. This case supports the opinion that supernumerary inv dup(22)(q11.1) could play a role in male infertility. We suggest that the breakpoint in the region 22q11.1 and/or fourfold dosage of centromeric/pericentromeric sequences of the chromosome 22 may be the cause of hypogonadotropic hypogonadism resulting in impaired spermatogenesis and infertility in our patient.

Titin A-band-specific monoclonal antibody Tit1 5H1.1. Cellular Titin as a centriolar protein in non-muscle cells.

We report the development of a new mouse anti-titin monoclonal antibody, named MAb Tit1 5H1.1, using the synthetic peptide corresponding to an amino acid sequence in the A-band of the titin molecule as immunogen. In the human skeletal muscle, MAb Tit1 5H1.1 reveals a clearly striated staining pattern, reacting with the A-band of the sarcomere. Electrophoretic, immunoblotting, and amino acid sequence analyses with ESI-MS/MS of human skeletal muscle tissue proved the target antigen of MAb Tit1 5H1.1 to be titin. The antibody reacts with titin also in non-muscle cells, producing a punctate pattern in cytoplasm and the nucleus. The most striking finding was a clear reaction of MAb Tit1 5H1.1 with centrioles in all cell types investigated so far. Immunocytochemical co-localization study with ninein-specific antibodies confirmed that the target antigen of MAb Tit1 5H1.1 is a centriole-associated protein. Experiments of the inhibition of synthesis of titin using titin siRNA duplex for the destruction of titin mRNA have shown a decreased staining of centrioles by MAb Tit1 5H1.1 in non-muscle cells and support the proposal that the target antigen of MAb is indeed titin. We suggest this anti-titin monoclonal antibody could be a valuable tool in the study of titin function and its subcellular location, both in muscle and non-muscle cells.

Two novel deletions (array CGH findings) in pigment dispersion syndrome.

PURPOSE: We report the first male with pigment dispersion syndrome and a balanced translocation t(10;15)(p11.1;q11.1). METHODS: Cytogenetic analyses using Giemsa banding and FISH methods, and array CGH were performed. RESULTS: Array CGH analyses did not show altered DNA sequences in the breakpoints of the translocation, but revealed two novel deletions in 2q22.1 and 18q22.1. CONCLUSION: We suppose that the coexistence of t(10;15) and pigment dispersion syndrome in our patient is a coincidence. The deletion in 2q22.1, where the gene LRP1B has been located, may play a major role in the dysembryogenesis of the eye and cause the disorder.

Reciprocal translocation t(7;16)(q21.2;p13.3) in an infertile man.

OBJECTIVE: To report the first case of reciprocal translocation t(7;16)(q21.2;p13.3) associated with male factor infertility. DESIGN: Case report. SETTING: University genetics laboratory and university andrology unit. PATIENT(S): A 38-year-old man with infertility and oligoasthenoteratozoospermia, but otherwise healthy. INTERVENTION(S): Chromosome analyses from peripheral blood lymphocyte cultures using Giemsa (G)-banding (GTG) and C-banding (CBG) and fluorescent in situ hybridization (FISH) were performed. MAIN OUTCOME MEASURE(S): Sperm count, motility, morphology, GTG and CBG banding, and FISH. RESULT(S): We report an apparently unique reciprocal translocation t(7;16)(q21.2;p13.3) confirmed by FISH in an infertile man. Semen analyses showed oligoasthenoteratozoospermia, with sperm count ranging from 2 x 10(6)/mL to 5 x 10(6)/mL and head defects (98%) in sperm morphology. CONCLUSION(S): In the present patient the breakpoint at 16p13.3 could have disrupted or harbored the PRM1, PRM2, or TNP2 genes responsible for the replacement of the histones involved in packaging the DNA into the sperm head. Resulting haploinsufficiency of these genes is likely to be the cause of sperm head defects and infertility in the patient. This case supports the opinion that alterations in the expression of protamine genes may be one of the causes of male factor infertility.