Glioblastoma is the commonest primary malignant brain tumor, with a very poor prognosis and short overall survival. It is characterized by its high intra- and intertumoral heterogeneity, in terms of both the level of single-nucleotide variants, copy number alterations, and aneuploidy. Therefore, routine diagnosis can be challenging in some cases. We present a complicated case of glioblastoma, which was characterized with five cytogenomic methods: interphase fluorescence in situ hybridization, multiplex ligation-dependent probe amplification, comparative genomic hybridization array and single-nucleotide polymorphism, targeted gene panel, and whole-genome sequencing. These cytogenomic methods revealed classical findings associated with glioblastoma, such as a lack of IDH and TERT mutations, gain of chromosome 7, and loss of chromosome 10. At least three pathological clones were identified, including one with whole-genome duplication, and one with loss of 1p and suspected loss of 19q. Deletion and mutation of the TP53 gene were detected with numerous breakends on 17p and 20q. Based on these findings, we recommend a combined approach to the diagnosis of glioblastoma involving the detection of copy number alterations, mutations, and aneuploidy. The choice of the best combination of methods is based on cost, time required, staff expertise, and laboratory equipment. This integrated strategy could contribute directly to tangible improvements in the diagnosis, prognosis, and prediction of the therapeutic responses of patients with brain tumors.

• MeSH
• glioblastom * genetika   patologie   diagnóza   MeSH
• hybridizace in situ fluorescenční metody   MeSH
• jednonukleotidový polymorfismus MeSH
• lidé středního věku MeSH
• lidé MeSH
• mutace MeSH
• nádorové biomarkery genetika   MeSH
• nádory mozku * genetika   patologie   diagnóza   MeSH
• prognóza MeSH
• srovnávací genomová hybridizace metody   MeSH
• variabilita počtu kopií segmentů DNA MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH

The current study assessed the performance of the fully automated RT-PCR-based IdyllaTM GeneFusion Assay, which simultaneously covers the advanced non-small cell lung carcinoma (aNSCLC) actionable ALK, ROS1, RET, and MET exon 14 rearrangements, in a routine clinical setting involving 12 European clinical centers. The IdyllaTM GeneFusion Assay detects fusions using fusion-specific as well as expression imbalance detection, the latter enabling detection of uncommon fusions not covered by fusion-specific assays. In total, 326 archival aNSCLC formalin-fixed paraffin-embedded (FFPE) samples were included of which 44% were resected specimen, 46% tissue biopsies, and 9% cytological specimen. With a total of 179 biomarker-positive cases (i.e., 85 ALK, 33 ROS1, 20 RET fusions and 41 MET exon 14 skipping), this is one of the largest fusion-positive datasets ever tested. The results of the IdyllaTM GeneFusion Assay were compared with earlier results of routine reference technologies including fluorescence in situ hybridization, immunohistochemistry, reverse-transcription polymerase chain reaction, and next-generation sequencing, establishing a high sensitivity/specificity of 96.1%/99.6% for ALK, 96.7%/99.0% for ROS1, 100%/99.3% for RET fusion, and 92.5%/99.6% for MET exon 14 skipping, and a low failure rate (0.9%). The IdyllaTM GeneFusion Assay was found to be a reliable, sensitive, and specific tool for routine detection of ALK, ROS1, RET fusions and MET exon 14 skipping. Given its short turnaround time of about 3 h, it is a time-efficient upfront screening tool in FFPE samples, supporting rapid clinical decision making. Moreover, expression-imbalance-based detection of potentially novel fusions may be easily verified with other routine technologies without delaying treatment initiation.

• MeSH
• anaplastická lymfomová kináza * genetika   MeSH
• exony * genetika   MeSH
• fúzní onkogenní proteiny * genetika   MeSH
• genová přestavba MeSH
• hybridizace in situ fluorescenční metody   MeSH
• lidé MeSH
• multiplexová polymerázová řetězová reakce MeSH
• nádorové biomarkery genetika   analýza   MeSH
• nádory plic * genetika   patologie   MeSH
• nemalobuněčný karcinom plic * genetika   patologie   MeSH
• protoonkogenní proteiny c-met * genetika   MeSH
• protoonkogenní proteiny c-ret * genetika   MeSH
• protoonkogenní proteiny * genetika   MeSH
• tyrosinkinasy * genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• multicentrická studie MeSH

Úvod a cíl: Primární diagnostika distálních biliárních striktur může být někdy svízelná a obtížná. Je známo, že kartáčková cytologie (BC – brush cytology) má v těchto případech nízkou diagnostickou senzitivitu. Fluorescenční in situ hybridizace (FISH) byla popsána jako užitečná doplňková metoda u pacientů se strikturami žlučových cest. Cílem naší studie bylo stanovit diagnostickou výtěžnost BC, FISH a jejich kombinace (BC + FISH) v primární diagnostice distálních biliárních striktur. Metodika: Tato prospektivní studie byla provedena v období od dubna 2019 do ledna 2021. Do studie byli postupně zařazeni pacienti s dosud nebioptovanými biliárními strikturami, kteří podstoupili první endoskopickou retrográdní cholangiopankreatografii (ERCP) na našem pracovišti. Byly provedeny cytologické a FISH analýzy vzorků tkáně odebraných ze dvou standardizovaných transpapilárních odběrů „kartáčkem“ z distálních striktur. Postoperační histologie nebo výsledky 12měsíčního sledování pacientů byly považovány za zlatý standard pro stanovení konečné diagnózy. Výsledky: Do studie bylo zařazeno celkem 109 pacientů. Sedm pacientů bylo z konečné analýzy vyřazeno a 26 mělo proximální stenózu. Ze zbývajících 76 pacientů (61,8 % mužů, průměrný věk 67,6 let, rozmezí 25–89 let) s distální stenózou byl podíl benigních a maligních striktur 25 (32,9 %), resp. 51 (67,1 %). Z podskupiny maligních striktur tvořilo 17,7 % cholangiokarcinom, 74,5 % nádory pankreatu a 7,8 % jiné nádory. Ve srovnání se samotnou BC zvýšila FISH senzitivitu z 0,373 % na 0,706 % (p = 0,0007) při mírném poklesu specificity (p = 0,045). Závěr: Duální modalita hodnocení tkáně pomocí BC + FISH má lepší senzitivitu pro primární diagnózu distálních biliárních striktur ve srovnání se samotnou BC.

Background and aim: Primary diagnosis of the distal biliary stricture can be sometime difficult. Brush cytology (BC) is known to have low diagnostic sensitivity in these cases. Fluorescence in situ hybridization (FISH) has been reported as a useful adjunctive test in patients with biliary strictures. We aimed to determine performance characteristics of BC, FISH and their combination (BC + FISH) in the primary diagnosis of distal biliary strictures. Methods: This single-center prospective study was conducted between April 2019 and January 2021. Consecutive patients with unsampled biliary strictures undergoing first ERCP in our institution were included. Cytological and FISH analysis of tissue specimens from two standardized transpapillary brushings from the distal strictures were provided. Histopathological confirmation after surgery or 12-month follow-up was regarded as the reference standard for the final diagnosis. Results: A total of 109 patients were enrolled. Seven patients were lost from the final analysis and 26 suffered proximal stenosis. Of the 76 remaining patients (61.8% males, mean age 67.6, range 25–89 years) with distal stenosis, the proportions of benign and malignant strictures were 25 (32.9%) and 51 (67.1%), respectively. Of the subgroup of malignant strictures, 17.7% were cholangiocarcinoma, 74.5% were pancreatic tumors and 7.8% others. In comparison to BC alone, FISH increased the sensitivity from 0.373% to 0.706% (p = 0.0007) with a slight decrease in specificity (p = 0.045). Conclusions: Dual modality tissue evaluation using BC + FISH has better sensitivity for the primary diagnosis of distal biliary strictures, compared to BC alone.

• Klíčová slova
• biliární striktura,
• MeSH
• hybridizace in situ fluorescenční * metody   MeSH
• lidé MeSH
• nemoci žlučových cest * diagnóza   MeSH
• prospektivní studie MeSH
• stenóza diagnóza   MeSH
• žlučové cesty patologie   MeSH
• Check Tag
• lidé MeSH

Cervids are characterized by their greatest karyotypic diversity among mammals. A great diversity of chromosome numbers in notably similar morphological groups leads to the existence of several complexes of cryptic species and taxonomic uncertainties. Some deer lineages, such as those of Neotropical deer, stand out for a rapid chromosomal reorganization and intraspecific chromosome polymorphisms, which have not been properly explored yet. For that reason, we contribute to the study of deer karyotype diversity and taxonomy by producing and characterizing new molecular cytogenetic markers for the gray brocket deer (Subulo gouazoubira), a deer species that retained the hypothetical ancestral karyotype of Cervidae. We used bacterial artificial chromosome (BAC) clones derived from the cattle genome (Bos taurus) as markers, which were hybridized on S. gouazoubira metaphase chromosomes. In total, we mapped 108 markers, encompassing all gray brocket deer chromosomes, except the Y chromosome. The detailed analysis of fluorescent in situ hybridization results showed 6 fissions and 1 fusion as interchromosomal rearrangements that have separated cattle and gray brocket deer karyotypes. Each group of BAC probes derived from bovine chromosome pairs 1, 2, 5, 6, 8, and 9 showed hybridization signals on 2 different chromosomes, while pairs 28 and 26 are fused in tandem in a single acrocentric chromosome in S. gouazoubira. Furthermore, the BAC markers detected the occurrence of intrachromosomal rearrangements in the S. gouazoubira chromosomes homologous to pair 1 and the X chromosome of cattle. We present a karyotypic map of the 108 new markers, which will be of great importance for future karyotypic evolution studies in cervids and, consequently, help in their conservation and taxonomy resolution.

• MeSH
• chromozom X MeSH
• hybridizace in situ fluorescenční metody   MeSH
• karyotyp MeSH
• karyotypizace MeSH
• skot genetika   MeSH
• umělé bakteriální chromozomy genetika   MeSH
• vysoká zvěř * genetika   MeSH
• zvířata MeSH
• Check Tag
• skot genetika   MeSH
• zvířata MeSH
• Publikační typ
• zprávy MeSH

Most patients with chronic lymphocytic leukaemia (CLL) are nowadays diagnosed without any symptoms and do not require therapy. A prognostic score identifying patients within this large group who are at high risk of disease progression would be highly beneficial. The recently published International Prognostic Score for Early asymptomatic patients (IPS-E) uses combination of absolute lymphocyte count (ALC) >15 × 109 /l, palpable lymphadenopathy, and unmutated immunoglobulin heavy-chain variable-region (IGHV) gene to predict the time to first-line therapy (TTFT). Patients at low, intermediate, and high risk had estimated 5-year TTFT of 8%, 28%, and 61%. We performed an external validation of the IPS-E score using an unselected, consecutive group of 130 Binet A patients. The 5-year TTFT was 11%, 36%, and 78% (C-statistic 0·74). Furthermore, we propose an alternative system (AIPS-E) using cytogenetic aberrations instead of palpable lymphadenopathy. This system yielded 5-year TTFT of 14%, 40%, and 72%. These results were externally validated in 388 Binet A patients from five Czech centres; the 5-year TTFT was 16%, 37%, and 80% (C-statistic 0·74). In conclusion, we have successfully validated the IPS-E score for patients with early stage CLL. In addition, we propose a modified scoring system, the AIPS-E, combining IGHV, fluorescence in situ hybridisation, and ALC.

• MeSH
• čas zasáhnout při rozvinutí nemoci statistika a číselné údaje   MeSH
• chromozomální aberace statistika a číselné údaje   MeSH
• chronická lymfatická leukemie krev   diagnóza   mortalita   patologie   MeSH
• geny pro těžké řetězce imunoglobulinů genetika   MeSH
• hybridizace in situ fluorescenční metody   MeSH
• kohortové studie MeSH
• lidé MeSH
• lymfadenopatie diagnóza   MeSH
• palpace metody   MeSH
• počet lymfocytů metody   MeSH
• prognóza MeSH
• progrese nemoci MeSH
• rizikové faktory MeSH
• senioři MeSH
• staging nádorů metody   MeSH
• výzkumný projekt trendy   MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH

The widely distributed ray-finned fish genus Carassius is very well known due to its unique biological characteristics such as polyploidy, clonality, and/or interspecies hybridization. These biological characteristics have enabled Carassius species to be successfully widespread over relatively short period of evolutionary time. Therefore, this fish model deserves to be the center of attention in the research field. Some studies have already described the Carassius karyotype, but results are inconsistent in the number of morphological categories for individual chromosomes. We investigated three focal species: Carassius auratus, C. carassius and C. gibelio with the aim to describe their standardized diploid karyotypes, and to study their evolutionary relationships using cytogenetic tools. We measured length (q+plength) of each chromosome and calculated centromeric index (i value). We found: (i) The relationship between q+plength and i value showed higher similarity of C. auratus and C. carassius. (ii) The variability of i value within each chromosome expressed by means of the first quartile (Q1) up to the third quartile (Q3) showed higher similarity of C. carassius and C. gibelio. (iii) The fluorescent in situ hybridization (FISH) analysis revealed higher similarity of C. auratus and C. gibelio. (iv) Standardized karyotype formula described using median value (Q2) showed differentiation among all investigated species: C. auratus had 24 metacentric (m), 40 submetacentric (sm), 2 subtelocentric (st), 2 acrocentric (a) and 32 telocentric (T) chromosomes (24m+40sm+2st+2a+32T); C. carassius: 16m+34sm+8st+42T; and C. gibelio: 16m+22sm+10st+2a+50T. (v) We developed R scripts applicable for the description of standardized karyotype for any other species. The diverse results indicated unprecedented complex genomic and chromosomal architecture in the genus Carassius probably influenced by its unique biological characteristics which make the study of evolutionary relationships more difficult than it has been originally postulated.

• MeSH
• chromozomy genetika   MeSH
• diploidie MeSH
• fylogeneze MeSH
• genetická variace genetika   MeSH
• genom genetika   MeSH
• hybridizace in situ fluorescenční metody   MeSH
• kapři genetika   MeSH
• karas zlatý genetika   MeSH
• karyotyp MeSH
• karyotypizace metody   MeSH
• mapování chromozomů metody   MeSH
• polyploidie MeSH
• ryby genetika   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

BACKGROUND: The blood flukes of genus Schistosoma are the causative agent of schistosomiasis, a parasitic disease that infects more than 200 million people worldwide. Proteases of schistosomes are involved in critical steps of host-parasite interactions and are promising therapeutic targets. We recently identified and characterized a group of S1 family Schistosoma mansoni serine proteases, including SmSP1 to SmSP5. Expression levels of some SmSPs in S. mansoni are low, and by standard genome sequencing technologies they are marginally detectable at the method threshold levels. Here, we report their spatial gene expression patterns in adult S. mansoni by the high-sensitivity localization assay. METHODOLOGY: Highly sensitive fluorescence in situ RNA hybridization (FISH) was modified and used for the localization of mRNAs encoding individual SmSP proteases (including low-expressed SmSPs) in tissues of adult worms. High sensitivity was obtained due to specifically prepared tissue and probes in combination with the employment of a signal amplification approach. The assay method was validated by detecting the expression patterns of a set of relevant reference genes including SmCB1, SmPOP, SmTSP-2, and Sm29 with localization formerly determined by other techniques. RESULTS: FISH analysis revealed interesting expression patterns of SmSPs distributed in multiple tissues of S. mansoni adults. The expression patterns of individual SmSPs were distinct but in part overlapping and were consistent with existing transcriptome sequencing data. The exception were genes with significantly low expression, which were also localized in tissues where they had not previously been detected by RNA sequencing methods. In general, SmSPs were found in various tissues including reproductive organs, parenchymal cells, esophagus, and the tegumental surface. CONCLUSIONS: The FISH-based assay provided spatial information about the expression of five SmSPs in adult S. mansoni females and males. This highly sensitive method allowed visualization of low-abundantly expressed genes that are below the detection limits of standard in situ hybridization or by RNA sequencing. Thus, this technical approach turned out to be suitable for sensitive localization studies and may also be applicable for other trematodes. The results suggest that SmSPs may play roles in diverse processes of the parasite. Certain SmSPs expressed at the surface may be involved in host-parasite interactions.

• MeSH
• exprese genu * MeSH
• hybridizace in situ fluorescenční metody   normy   MeSH
• proteiny červů genetika   MeSH
• RNA metabolismus   MeSH
• Schistosoma mansoni enzymologie   genetika   MeSH
• serinové proteasy genetika   MeSH
• stanovení celkové genové exprese MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

So-called oncocytic papillary renal cell carcinoma (OPRCC) is a poorly defined variant of papillary renal cell carcinoma. Since its first description, several studies were published with conflicting results, and thus precise definition is lacking. A cohort of 39 PRCCs composed of oncocytic cells were analyzed. Cases were divided into 3 groups based on copy number variation (CNV) pattern. The first group consisted of 23 cases with CNV equal to renal oncocytoma. The second group consisted of 7 cases with polysomy of chromosomes 7 and 17 and the last group of 9 cases included those with variable CNV. Epidemiologic, morphologic and immunohistochemical features varied among the groups. There were not any particular histomorphologic features correlating with any of the genetic subgroups. Further, a combination of morphologic, immunohistochemical, and molecular-genetic features did not allow to precisely predict biologic behavior. Owing to variable CNV pattern in OPRCC, strict adherence to morphology and immunohistochemical profile is recommended, particularly in limited samples (i.e., core biopsy). Applying CNV pattern as a part of a diagnostic algorithm can be potentially misleading. OPRCC is a highly variable group of tumors, which might be misdiagnosed as renal oncocytoma. Using the term OPRCC as a distinct diagnostic entity is, thanks to its high heterogeneity, questionable.

• MeSH
• biopsie dutou jehlou normy   MeSH
• chromozomální aberace MeSH
• chybná diagnóza MeSH
• diferenciální diagnóza MeSH
• dospělí MeSH
• hybridizace in situ fluorescenční metody   MeSH
• imunohistochemie metody   MeSH
• karcinom z renálních buněk epidemiologie   genetika   patologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• nádorové biomarkery metabolismus   MeSH
• nádory ledvin diagnóza   genetika   patologie   MeSH
• oxyfilní adenom diagnóza   genetika   patologie   MeSH
• oxyfilní buňky metabolismus   patologie   MeSH
• překrývající se geny genetika   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• staging nádorů metody   MeSH
• variabilita počtu kopií segmentů DNA genetika   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Repetitive DNAs comprise large portion of eukaryote genomes. In genome projects, the assembly of repetitive DNAs is challenging due to the similarity between repeats, which generate ambiguities for alignment. Fluorescence in situ hybridization (FISH) is a powerful technique for the physical mapping of various sequences on chromosomes. This technique is thus very helpful in chromosome-based genome assemblies, providing information on the fine architecture of genomes and their evolution. However, various protocols are currently used for FISH mapping, most of which are relatively laborious and expensive, or work properly only with a specific type of probes or sequences, and there is a need for a universal and affordable FISH protocol. Here we tested a FISH protocol for mapping of different DNA repeats, such as multigene families (rDNAs, U snDNAs, histone genes), satellite DNAs, microsatellites, transposable elements, DOP-PCR products, and telomeric motif (TTAGG)n, on the chromosomes of various insects and other arthropods. Different cell types and stages obtained from diverse tissues were used. The FISH procedure proved high quality and reliable results in all experiments performed. We obtained data on the chromosomal distribution of DNA repeats in representatives of insects and other arthropods. Thus, our results allow us to conclude that the protocol is universal and requires only time adjustment for chromosome/DNA denaturation. The use of this FISH protocol will facilitate studies focused on understanding the evolution and role of repetitive DNA in arthropod genomes.

• MeSH
• členovci genetika   MeSH
• DNA genetika   MeSH
• fluorescence MeSH
• hmyz genetika   MeSH
• hybridizace in situ fluorescenční metody   MeSH
• mapování chromozomů metody   MeSH
• molekulární evoluce MeSH
• multigenová rodina genetika   MeSH
• repetitivní sekvence nukleových kyselin genetika   MeSH
• telomery genetika   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Duckweeds represent a small, free-floating aquatic family (Lemnaceae) of the monocot order Alismatales with the fastest growth rate among flowering plants. They comprise five genera (Spirodela, Landoltia, Lemna, Wolffiella, and Wolffia) varying in genome size and chromosome number. Spirodela polyrhiza had the first sequenced duckweed genome. Cytogenetic maps are available for both species of the genus Spirodela (S. polyrhiza and S. intermedia). However, elucidation of chromosome homeology and evolutionary chromosome rearrangements by cross-FISH using Spirodela BAC probes to species of other duckweed genera has not been successful so far. We investigated the potential of chromosome-specific oligo-FISH probes to address these topics. We designed oligo-FISH probes specific for one S. intermedia and one S. polyrhiza chromosome (Fig. 1a). Our results show that these oligo-probes cross-hybridize with the homeologous regions of the other congeneric species, but are not suitable to uncover chromosomal homeology across duckweeds genera. This is most likely due to too low sequence similarity between the investigated genera and/or too low probe density on the target genomes. Finally, we suggest genus-specific design of oligo-probes to elucidate chromosome evolution across duckweed genera.

• MeSH
• Araceae klasifikace   genetika   růst a vývoj   MeSH
• chromozomy rostlin genetika   MeSH
• druhová specificita MeSH
• fylogeneze MeSH
• genom rostlinný * MeSH
• hybridizace in situ fluorescenční metody   MeSH
• karyotypizace MeSH
• molekulární evoluce * MeSH
• oligonukleotidové sondy chemie   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH