1. Quality assurance for next-generation sequencing diagnostics of rare neurological diseases in the European Reference NetworkAleš Maver, Katja Lohmann, Fran Borovečki, Nicola Wolstenholme, Rachel L. Taylor, Malte Spielmann, Tobias B. Haack, Matthias Gerberding, Borut Peterlin, Holm Graessner, 2024, izvirni znanstveni članek Povzetek: In the past decade, next-generation sequencing (NGS) has revolutionised genetic diagnostics for rare neurological disorders (RND). However, the lack of standardised technical, interpretative, and reporting standards poses a challenge for ensuring consistent and high-quality diagnostics globally. To address this, the European Reference Network for Rare Neurological Diseases (ERN-RND) collaborated with the European Molecular Genetics Quality Network (EMQN) to establish an external quality assessment scheme for NGS-based diagnostics in RNDs. The scheme, initiated in 2021 with a pilot involving 29 labs and followed by a second round in 2022 with 42 labs, aimed to evaluate the performance of laboratories in genetic testing for RNDs. Each participating lab analysed genetic data from three hypothetical cases, assessing genotyping, interpretation, and clerical accuracy. Despite a majority of labs using exome or genome sequencing, there was considerable variability in gene content, sequencing quality, adherence to standards, and clinical guidance provision. Results showed that while most labs provided correct molecular diagnoses, there was significant variability in reporting technical quality, adherence to interpretation standards, reporting strategies, and clinical commentary. Notably, some labs returned results with the potential for adverse medical outcomes. This underscores the need for further harmonisation, guideline development, and external quality assessment in the evolving landscape of genomic diagnostics for RNDs. Overall, the experience with the scheme highlighted the generally good quality of participating labs but emphasised the imperative for ongoing improvement in data analysis, interpretation, and reporting to enhance patient safety.
Ključne besede: next-generation sequencing, neurological disorders, genetic diagnostics, rare diseases
Objavljeno v DiRROS: 10.03.2026; Ogledov: 46; Prenosov: 28
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2. Reverse phenotyping after whole-exome sequencing in children with developmental delay/intellectual disability : an exception or a necessity?Nikola Ilic, Nina Maric, Aleš Maver, Lluis Armengol, Ružica Kravljanac, Jana Cirkovic, Jovana Krstić, Borut Peterlin, 2024, izvirni znanstveni članek Povzetek: This study delves into the diagnostic yield of whole-exome sequencing (WES) in pediatric patients presenting with developmental delay/intellectual disability (DD/ID), while also exploring the utility of Reverse Phenotyping (RP) in refining diagnoses. A cohort of 100 pediatric patients underwent WES, yielding a diagnosis in 66% of cases. Notably, RP played a significant role in cases with negative prior genetic testing, underscoring its significance in complex diagnostic scenarios. The study revealed a spectrum of genetic conditions contributing to DD/ID, illustrating the heterogeneity of etiological factors. Despite challenges, WES demonstrated effectiveness, particularly in cases with metabolic abnormalities. Reverse phenotyping was indicated in half of the patients with positive WES findings. Neural network models exhibited moderate-to-exceptional predictive abilities for aiding in patient selection for WES and RP. These findings emphasize the importance of employing comprehensive genetic approaches and RP in unraveling the genetic underpinnings of DD/ID, thereby facilitating personalized management and genetic counseling for affected individuals and families. This research contributes insights into the genetic landscape of DD/ID, enhancing our understanding and guiding clinical practice in this particular field of clinical genetics.
Ključne besede: developmental delay, intellectual disability, whole-exome sequencing, reverse phenotyping, neural network analysis
Objavljeno v DiRROS: 10.03.2026; Ogledov: 39; Prenosov: 24
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3. Microbiome analysis of groundwaters in the Slovenian Dinaric Karst, a recognized biodiversity hotspotKatarina Bačnik, Denis Kutnjak, Cene Gostinčar, Rok Kostanjšek, Janez Mulec, Monika Novak Babič, Ion Gutiérrez-Aguirre, Nina Gunde-Cimerman, Martina Turk, 2026, izvirni znanstveni članek Povzetek: Groundwater is the most important source of drinking water supply, irrigation, and industrial use, as well as a habitat for a diverse range of (micro)organisms. Some of the first studies of underground environments were carried out in the Dinaric Karst region of Slovenia, which harbors one of the highest subterranean biodiversities in the world. Despite their ecological importance, groundwater microbial communities remain underexplored. In this study, we conducted a comprehensive microbiome analysis of karst groundwater, investigating the abundance and diversity of viruses, archaea, bacteria, fungi, and protozoa. We detected a high diversity of both prokaryote- and eukaryote-infecting viruses, indicating the connection of subterranean environments with surface waters. In the archaeal community, seven different phyla were identified, dominated by members of "Candidatus Woesearchaeota" and Nitrososphaerota (syn. Thermoproteota). In the bacterial community of majority of the samples, the Pseudomonadota was the most abundant phylum, followed by Bacteroidota, Actinomycetota, and Verrucomicrobiota. Fungal abundance and diversity differed greatly between cave water samples, with the Ascomycota families Cladosporiaceae, Didymellaceae, Aspergillaceae and saprotrophic Basidiomycota detected in all samples. Besides fungi, microscopic eukaryotic diversity consisted of ciliates, copepods and golden-brown algae present in all samples. Differences in microbial communities between the sampling sites highlight the need for tailored site-specific groundwater conservation and monitoring strategies. Moreover, the identification of microbes associated with anthropogenic activities underscores their potential as source-specific indicators of groundwater pollution. As the first holistic overview of microbial diversity of subterranean water, the study establishes a valuable baseline for future research and conservation of these unique karst ecosystems.
Ključne besede: Karst cave water, high-throughput sequencing, viruses, archaea, bacteria, fungi, protozoa
Objavljeno v DiRROS: 02.03.2026; Ogledov: 128; Prenosov: 65
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4. Microbial DNA extraction of high-host content and low biomass samples : optimized protocol for nasopharynx metagenomic studiesPolona Rajar, Achal Dhariwal, Gabriela Salvadori, Roger Junges, Heidi Aarø Åmdal, Dag Berild, Drude Fugelseth, Ola Didrik Saugstad, 2022, izvirni znanstveni članek Povzetek: Introduction: Low microbial biomass and high human DNA content in nasopharyngeal aspirate samples hinder comprehensive characterization of microbiota and resistome. We obtained samples from premature infants, a group with increased risk of developing respiratory disorders and infections, and consequently frequent exposure to antibiotics. Our aim was to devise an optimal protocol for handling nasopharyngeal aspirate samples from premature infants, focusing on host DNA depletion and microbiome and resistome characterization. Methods: Three depletion and three DNA extraction protocols were compared, using RT-PCR and whole metagenome sequencing to determine the efficiency of human DNA removal, taxonomic profiling and assignment of antibiotic resistance genes. Protocols were tested using mock communities, as well as pooled and individual patient samples. Results: The only extraction protocol to retrieve the expected DNA yield from mock community samples was based on a lytic method to improve Gram positive recovery (MasterPure™). Host DNA content in non-depleted aliquots from pooled patient samples was 99%. Only samples depleted with MolYsis™ showed satisfactory, but varied reduction in host DNA content, in both pooled and individual patient samples, allowing for microbiome and resistome characterisation (host DNA content from 15% to 98%). Other depletion protocols either retrieved too low total DNA yields, preventing further analysis, or failed to reduce host DNA content. By using Mol_MasterPure protocol on aliquots from pooled patient samples, we increased the number of bacterial reads by 7.6 to 1,725.8-fold compared to non-depleted reference samples. PCR results were indicative of achieved microbial enrichment. Individual patient samples processed with Mol_MasterPure protocol varied greatly in total DNA yield, host DNA content (from 40% to 98%), species and antibiotic resistance gene richness. Discussion: Despite high human DNA and low microbial biomass content in nasopharynx aspirates of preterm infants, we were able to reduce host DNA content to levels compatible with downstream shotgun metagenomic analysis, including bacterial species identification and coverage of antibiotic resistance genes. Whole metagenomic sequencing of microbes colonizing the nasopharynx may contribute to explaining the possible role of airway microbiota in respiratory conditions and reveal carriage of antibiotic resistance genes.
Ključne besede: antimicrobial resistance, host DNA depletion, low biomass, microbiome, premature infant, resistome, respiratory microbiome, whole metagenomic sequencing
Objavljeno v DiRROS: 25.02.2026; Ogledov: 150; Prenosov: 42
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6. Genetic characterization of buckwheat accessions through genome-wide allele frequency fingerprintsMichelle M. Nay, Stephen L. Byrne, Eduardo A. Pérez, Achim Walter, Bruno Studer, 2020, izvirni znanstveni članek Povzetek: Genomics-assisted breeding of buckwheat (Fagopyrum esculentum Moench) depends on robust genotyping methods. Genotyping by sequencing (GBS) has evolved as a flexible and cost-effective technique frequently used in plant breeding. Several GBS pipelines are available to genetically characterize single genotypes but these are not able to represent the genetic diversity of buckwheat accessions that are maintained as genetically heterogeneous, open-pollinating populations. Here we report the development of a GBS pipeline which, rather than reporting the state of bi-allelic single nucleotide polymorphisms (SNPs), resolves allele frequencies within populations on a genome-wide scale. These genome-wide allele frequency fingerprints (GWAFFs) from 100 pooled individual plants per accession were found to be highly reproducible and revealed the genetic similarity of 20 different buckwheat accessions analysed in our study. The GWAFFs cannot only be used as an efficient tool to precisely describe buckwheat breeding material, they also offer new opportunities to investigate the genetic diversity between different buckwheat accessions and establish variant databases for key material. Furthermore, GWAFFs provide the opportunity to associate allele frequencies to phenotypic traits and quality parameters that are most reliably described on population level. This is the key to practically implement powerful genomics-assisted breeding concepts such as marker-assisted selection and genomic selection in future breeding schemes of allogamous buckwheat.
Ključne besede: Buckwheat (Fagopyrum esculentum Moench), genotyping by sequencing (GBS), population genomics, genome-wide allele frequency fingerprints (GWAFFs)
Objavljeno v DiRROS: 20.01.2026; Ogledov: 249; Prenosov: 125
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7. Changes in motor unit conduction velocity after unilaterallower-limb suspension and active recovery are correlated withmuscle ion channel gene expressionGiacomo Valli, Fabio Sarto, Francesco Negro, Elena Monti, Giuseppe Sirago, Matteo Paganini, Sandra Zampieri, Martino V. Franchi, Andrea Casolo, Julián Candia, Luigi Ferrucci, Marco Vincenzo Narici, Giuseppe De Vito, 2025, izvirni znanstveni članek Povzetek: The effects of muscle disuse on the propagation of action potentials along motorunit (MU) muscle fibres, a key process for effective muscle activation and forcegeneration, remain poorly understood. The aim of this study was to investigate changesin action potential propagation and to identify biological factors influencing thesechanges following unilateral lower-limb suspension (ULLS) and active recovery (AR).Eleven young males underwent 10 days of ULLS followed by 21 days of AR involvingresistance exercise. Maximal force of the knee extensors (MVC), high-density surfaceEMG recordings and muscle biopsies of the vastus lateralis muscle were collectedbefore ULLS, after ULLS and after AR. EMG recordings collected during submaximalisometric contractions were decomposed to estimate single-MU conduction velocity(CV). Biopsies were used to measure muscle fibre diameters via histochemical analysisand ion channel transcriptomic profiles via mRNA sequencing. The MVC declined by29% after ULLS and returned to baseline after AR. MU CV decreased after ULLSand recovered fully, even exceeding baseline values after AR. Muscle fibre diametersdid not change across the interventions and showed no correlation with MU CV.Conversely, a feature importance analysis revealed that mRNA expression levels ofspecific ion channel genes, particularly those involved in K+ transport, were correlatedwith MU CV at baseline and across the interventions. This study highlights the crucialrole of K+ ion channels in influencing MU CV in humans, offering new insights into MUCV modulation and the mechanisms of changes in muscle force after disuse and activerecovery.
Ključne besede: ion channel, mRNA sequencing, muscle fibre diameter, muscle unloading, neuromuscular impairment
Objavljeno v DiRROS: 12.01.2026; Ogledov: 185; Prenosov: 114
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8. Creating the Slovenian genome database and browser as a source of comprehensive variation of the Slovenian populationAleš Maver, Peter Juvan, Urška Kotnik, Luca Lovrečić, Gaber Bergant, Borut Peterlin, 2025, izvirni znanstveni članek Povzetek: The genomic data of Central European populations is underrepresented in the publicly available databases. We present the comprehensive genomic variation of the Slovenian population, based on the genomic sequencing of 9425 non-related individuals, i.e. more than 0.44% of the Slovenian population. Over 30 million unique single nucleotide and small indel (30.8 million), copy number (217.6 thousand), and mitochondrial variants (3.3 thousand) were uncovered and annotated by analysing the whole genome of 619 individuals and the whole exome of 8806 individuals. This population variation, including 3,9 million novel variants, is presented in a publicly available genome variant browser, the SloGenVar (https://slogenvar.si). We used this newly developed resource to reveal the population frequency of pathogenic variants in the genes associated with recessive conditions. The Slovenian genome database and browser offer the largest and the most comprehensive publicly available Central European population genomic variant resource, providing an important asset for genomic studies and as a control variant database for variant interpretation in the region and beyond.
Ključne besede: population genomic variation, whole genome sequencing, genome browser, Slovenian genome database
Objavljeno v DiRROS: 22.12.2025; Ogledov: 339; Prenosov: 139
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9. Prolonged hospitalization signature and early antibiotic effects on the nasopharyngeal resistome in preterm infantsAchal Dhariwal, Polona Rajar, Gabriela Salvadori, Heidi Aarø Åmdal, Dag Berild, Ola Didrik Saugstad, Drude Fugelseth, Gorm Greisen, 2024, izvirni znanstveni članek Povzetek: Respiratory pathogens, commonly colonizing nasopharynx, are among the leading causes of death due to antimicrobial resistance. Yet, antibiotic resistance determinants within nasopharyngeal microbial communities remain poorly understood. In this prospective cohort study, we investigate the nasopharynx resistome development in preterm infants, assess early antibiotic impact on its trajectory, and explore its association with clinical covariates using shotgun metagenomics. Our findings reveal widespread nasopharyngeal carriage of antibiotic resistance genes (ARGs) with resistomes undergoing transient changes, including increased ARG diversity, abundance, and composition alterations due to early antibiotic exposure. ARGs associated with the critical nosocomial pathogen Serratia marcescens persist up to 8–10 months of age, representing a long-lasting hospitalization signature. The nasopharyngeal resistome strongly correlates with microbiome composition, with inter-individual differences and postnatal age explaining most of the variation. Our report on the collateral effects of antibiotics and prolonged hospitalization underscores the urgency of further studies focused on this relatively unexplored reservoir of pathogens and ARGs.
Ključne besede: preterm infants, respiratory resistome, whole genome sequencing, genetics, drug effects
Objavljeno v DiRROS: 12.12.2025; Ogledov: 280; Prenosov: 157
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10. The role of the MTUS1 gene in the development of left ventricular noncompaction cardiomyopathy : a case reportTevž Gorjanc, Jaka Šikonja, Ana Drole Torkar, Mojca Žerjav-Tanšek, Jernej Kovač, Sara Bertok, Maruša Debeljak, Zvezdana Dolenc-Stražar, Marija Meznarič, Jernej Mlakar, Mirko Topalović, Gorazd Mlakar, Tadej Battelino, Urh Grošelj, 2025, drugi znanstveni članki Ključne besede: left ventricular noncompaction cardiomyopathy, microtubule-associated scaffold protein 1 (MTUS1), whole-genome sequencing
Objavljeno v DiRROS: 02.12.2025; Ogledov: 328; Prenosov: 151
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