Veronika Krulisova

Genetic characterization of northeastern Italian population isolates in the context of broader European genetic diversity

Population genetic studies on European populations have highlighted Italy as one of genetically most diverse regions. This is possibly due to the country’s complex demographic history and large variability in terrain throughout the territory. This is the reason why Italy is enriched for population isolates, Sardinia being the best-known example. As the population isolates have a great potential in disease-causing genetic variants identification, we aimed to genetically characterize a region from northeastern Italy, which is known for isolated communities. Total of 1310 samples, collected from six geographically isolated villages, were genotyped at >145 000 single-nucleotide polymorphism positions. Newly genotyped data were analyzed jointly with the available genome-wide data sets of individuals of European descent, including several population isolates. Despite the linguistic differences and geographical isolation the village populations still show the greatest genetic similarity to other Italian samples. The genetic isolation and small effective population size of the village populations is manifested by higher levels of genomic homozygosity and elevated linkage disequilibrium. These estimates become even more striking when the detected substructure is taken into account. The observed level of genetic isolation in Friuli-Venezia Giulia region is more extreme according to several measures of isolation compared with Sardinians, French Basques and northern Finns, thus proving the status of an isolate.

Comparison of different IRT-PAP protocols to screen newborns for cystic fibrosis in three central European populations

BACKGROUND In recent years different IRT/PAP protocols have been evaluated, but the individual performance remains unclear. To optimize the IRT/PAP strategy we compared protocols from three regional CF newborn screening centers (Heidelberg, Dresden, and Prague). METHODS We evaluated the effect of elevating the IRT-cut-off from 50 to 65 μg/l (~97.5th to ~99.0th percentile), the need of a failsafe protocol (FS, IRT ≥ 99.9th percentile) and the relative performance using either two IRT-dependent PAP-cut-offs or one PAP-cut-off. FINDINGS Elevation of the IRT cut-off to 65 μg/l (~99.0th percentile) increased the PPV significantly (Dresden: 0.065 vs. 0.080, p < 0.0001, Prague: 0.052 vs. 0.074, p < 0.0001) without reducing sensitivity. All three IRT/PAP protocols showed a trend towards a higher sensitivity with FS than without and when using one PAP-cut-off instead of two IRT-dependent PAP-cut-offs. CONCLUSIONS For best performance we suggest an IRT/PAP protocol with an IRT-cut-off close to the 99.0th percentile, FS, and a single PAP-cut-off.

A Genome-Wide Analysis of Populations from European Russia Reveals a New Pole of Genetic Diversity in Northern Europe

Several studies examined the fine-scale structure of human genetic variation in Europe. However, the European sets analyzed represent mainly northern, western, central, and southern Europe. Here, we report an analysis of approximately 166,000 single nucleotide polymorphisms in populations from eastern (northeastern) Europe: four Russian populations from European Russia, and three populations from the northernmost Finno-Ugric ethnicities (Veps and two contrast groups of Komi people). These were compared with several reference European samples, including Finns, Estonians, Latvians, Poles, Czechs, Germans, and Italians. The results obtained demonstrated genetic heterogeneity of populations living in the region studied. Russians from the central part of European Russia (Tver, Murom, and Kursk) exhibited similarities with populations from central–eastern Europe, and were distant from Russian sample from the northern Russia (Mezen district, Archangelsk region). Komi samples, especially Izhemski Komi, were significantly different from all other populations studied. These can be considered as a second pole of genetic diversity in northern Europe (in addition to the pole, occupied by Finns), as they had a distinct ancestry component. Russians from Mezen and the Finnic-speaking Veps were positioned between the two poles, but differed from each other in the proportions of Komi and Finnic ancestries. In general, our data provides a more complete genetic map of Europe accounting for the diversity in its most eastern (northeastern) populations.

A product of immunoreactive trypsinogen and pancreatitis-associated protein as second-tier strategy in cystic fibrosis newborn screening.

BACKGROUND In cystic fibrosis newborn screening (CFNBS), immunoreactive trypsinogen (IRT) and pancreatitis-associated protein (PAP) can be used as screening parameters. We evaluated the IRT×PAP product as second-tier parameter in CFNBS in newborns with elevated IRT. METHODS Data on 410,111 screened newborns including 78 patients with classical cystic fibrosis (CF) from two European centers were retrospectively analyzed by discrimination analysis to identify a screening protocol with optimal cutoffs. We also studied differences in PAP measurement methods and the association of IRT and PAP with age. RESULTS PAP values differed systematically between fluorometric and photometric assays. The IRT×PAP product showed better discrimination for classical CF than PAP only as second-tier screening parameter (p<0.001). In CF patients, IRT decreased while PAP values remained high over years. In newborns without CF, IRT decreased after birth over weeks while PAP increased within days. CONCLUSIONS The IRT×PAP product performs well as second-tier cutoff parameter for CFNBS. Screening quality parameters depend on the analytic method and on age at blood collection.

Under the mask of Kabuki syndrome: Elucidation of genetic-and phenotypic heterogeneity in patients with Kabuki-like phenotype

Kabuki syndrome is mainly caused by dominant de-novo pathogenic variants in the KMT2D and KDM6A genes. The clinical features of this syndrome are highly variable, making the diagnosis of Kabuki-like phenotypes difficult, even for experienced clinical geneticists. Herein we present molecular genetic findings of causal genetic variation using array comparative genome hybridization and a Mendeliome analysis, utilizing targeted exome analysis focusing on regions harboring rare disease-causing variants in Kabuki-like patients which remained KMT2D/KDM6A-negative. The aCGH analysis revealed a pathogenic CNV in the 14q11.2 region, while targeted exome sequencing revealed pathogenic variants in genes associated with intellectual disability (HUWE1, GRIN1), including a gene coding for mandibulofacial dysostosis with microcephaly (EFTUD2). Lower values of the MLL2-Kabuki phenotypic score are indicative of Kabuki-like phenotype (rather than true Kabuki syndrome), where aCGH and Mendeliome analyses have high diagnostic yield. Based on our findings we conclude that for new patients with Kabuki-like phenotypes it is possible to choose a specific molecular testing approach that has the highest detection rate for a given MLL2-Kabuki score, thus fostering more precise patient diagnosis and improved management in these genetically- and phenotypically heterogeneous clinical entities.

25 Cystic fibrosis (CF) prevalence derived from CF newborn screening (CFNBS) in the Czech Republic: comparison of previous epidemiological and current CFNBS-based disease prevalence data

Objectives: This study reviews parental views on how they were informed of the probable CF diagnosis, following neonatal screening, and explores which method parent’s found most acceptable. Methods: Retrospective questionnaires were used to find parents’ views on how they were told that their child had a probable diagnosis of CF, who told them initially, were they told at home or by the GP followed by hospital, how acceptable was the mode of information sharing? Results: 40 parents of children with CF were studied. Most parents (9 out of 12) who had been informed by their GP recall receiving inaccurate information about the condition. All parents (28 out of 28) who had received a home visit to be told about the CF diagnosis felt that the information received had been accurate and found it easier to accept. Conclusion: Our qualitative study confirms that parents who were given accurate information by CF professionals during a home visit found this an acceptable means of being informed of the probable diagnosis of CF. Parents reported less negative recollections of that time than parents who were informed by their GP and met CF professionals in hospital. We consider that this is a superior method of sharing the information about the diagnosis of CF.

17 Cystic fibrosis newborn screening (CF NBS): 4 year experience of the Prague Centre with the IRT/DNA/IRT protocol

Background: Cystic Fibrosis Newborn screening (CFNBS) as a pilot study started at the Institute of Mother and Child (IMC) Centre in 1999 and 444 063 newborns were examined until 2003. Current CFNBS has started gradually in Poland in 2006, covering the whole country in 2009 and is ongoing. 582 693 children were screened until the end of 2011. During CFNBS different protocol’s strategies were used. Aim: Impact of the implementation of NBS on the age of CF diagnosis. Methods: The study involved children diagnosed and treated only in IMC CF Centre (from 1999 until the end of 2011). Three groups were formed: 1. Pilot group (p-NBS) − 56 children, according to protocol strategy IRT/IRT and IRT/IRT/DNA, only F508del mutation was assessed; 2. Current group (c-NBS) − 92 children, IRT/IRT/DNA and IRT/DNA protocol, with expanded DNA analysis panel; 3. Patient diagnosed clinically (non-NBS) − 56 children. Moreover, DNA analysis in c-NBS group contains also frequent mutation in Polish population: 3849+10kbC>T, which is combined with low sweat test values. All patients underwent sweat tests. Results: The age of diagnosis was significantly lower (both p T than p-NBS and non-NBS group (both 0.9%). Conclusion: IRT/DNA strategy with extended DNA analysis provides an opportunity of earlier CF diagnosis even in children with normal sweat test values.