Plevova, Kristina* Dotaz Zobrazit nápovědu
Studying bacterial population diversity is important to understand healthcare associated infections' epidemiology and has a significant impact on dealing with multidrug resistant bacterial outbreaks. We characterised the extended-spectrum beta-lactamase producing K. pneumoniae (ESBLp KPN) population in our hospital using mini-MLST. Then we used whole genome sequencing (WGS) to compare selected isolates belonging to the most prevalent melting types (MelTs) and the colonization/infection pair isolates collected from one patient to study the ESBLp KPN population's genetic diversity. A total of 922 ESBLp KPN isolates collected between 7/2016 and 5/2018 were divided into 38 MelTs using mini-MLST with only 6 MelTs forming 82.8% of all isolates. For WGS, 14 isolates from the most prominent MelTs collected in the monitored period and 10 isolates belonging to the same MelTs collected in our hospital in 2014 were randomly selected. Resistome, virulome and ST were MelT specific and stable over time. A maximum of 23 SNV per core genome and 58 SNV per core and accessory genome were found. To determine the SNV relatedness cut-off values, 22 isolates representing colonization/infection pair samples obtained from 11 different patients were analysed by WGS with a maximum of 22 SNV in the core genome and 40 SNV in the core and accessory genome within pairs. The mini-MLST showed its potential for real-time epidemiology in clinical practice. However, for outbreak evaluation in a low diversity bacterial population, mini-MLST should be combined with more sensitive methods like WGS. Our findings showed there were only minimal differences within the core and accessory genome in the low diversity hospital population and gene based SNV analysis does not have enough discriminatory power to differentiate isolate relatedness. Thus, intergenic regions and mobile elements should be incorporated into the analysis scheme to increase discriminatory power.
- MeSH
- bakteriální proteiny genetika MeSH
- beta-laktamasy genetika metabolismus MeSH
- dítě MeSH
- DNA bakterií genetika MeSH
- dospělí MeSH
- infekce bakteriemi rodu Klebsiella enzymologie epidemiologie genetika mikrobiologie MeSH
- infekce spojené se zdravotní péčí enzymologie epidemiologie genetika mikrobiologie MeSH
- Klebsiella pneumoniae enzymologie genetika izolace a purifikace MeSH
- kojenec MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mnohočetná bakteriální léková rezistence * MeSH
- multilokusová sekvenční typizace * MeSH
- novorozenec MeSH
- předškolní dítě MeSH
- sekvenování celého genomu * MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- Check Tag
- dítě MeSH
- dospělí MeSH
- kojenec MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mužské pohlaví MeSH
- novorozenec MeSH
- předškolní dítě MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
MicroRNA (miRNA) expression is deregulated in many tumors including chronic lymphocytic leukemia (CLL). Although the particular mechanism(s) responsible for their aberrant expression is not well characterized, the presence of mutations and single-nucleotide polymorphisms (SNPs) in miRNA genes, possibly affecting their secondary structure and expression, has been described. In CLL; however, the impact and frequency of such variations have yet to be elucidated. Using a custom resequencing microarray, we screened sequence variations in 109 cancer-related pre-miRNAs in 98 CLL patients. Additionally, the primary regions of miR-29b-2/29c and miR-16-1 were analyzed by Sanger sequencing in another cohort of 213 and 193 CLL patients, respectively. Altogether, we describe six novel miR-sequence variations and the presence of SNPs (n = 27), most of which changed the miR-secondary structure. Moreover, some of the identified SNPs have a significantly different frequency in CLL when compared with a control population. Additionally, we identified a novel variation in miR-16-1 that had not been described previously in CLL patients. We show that this variation affects the expression of mature miR-16-1. We also show that the expression of another miRNA with pathogenetic relevance for CLL, namely miR-29b-2, is influenced by the presence of a polymorphic insertion, which is more frequent in CLL than in a control population. Altogether, these data suggest that sequence variations may occur during CLL development and/or progression.
- MeSH
- alely MeSH
- chromozomální aberace MeSH
- chronická lymfatická leukemie genetika MeSH
- dospělí MeSH
- frekvence genu MeSH
- genetická variace * MeSH
- jednonukleotidový polymorfismus MeSH
- konformace nukleové kyseliny MeSH
- lidé středního věku MeSH
- lidé MeSH
- mikro RNA chemie genetika MeSH
- mutace MeSH
- regulace genové exprese u leukemie MeSH
- sekvenční analýza DNA MeSH
- senioři MeSH
- těžké řetězce imunoglobulinů genetika MeSH
- zárodečné mutace MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Publikační typ
- abstrakt z konference MeSH
Chronic lymphocytic leukemia (CLL) patients may acquire new chromosome abnormalities during the course of their disease. Clonal evolution (CE) has been detected by conventional chromosome banding (CBA), several groups also confirmed CE with fluorescence in situ hybridization (FISH). At present, there are minimal prospective data on CE frequency determined using a combination of both methods. Therefore, the aim of our study was to prospectively assess CE frequency using a combination of FISH and CBA after stimulation with CpG oligonucleotides and interleukin-2. Between 2008 and 2012, we enrolled 140 patients with previously untreated CLL in a prospective trial evaluating CE using FISH and CBA after stimulation. Patients provided baseline and regular follow-up peripheral blood samples for testing. There was a median of 3 cytogenetic examinations (using both methods) per patient. CE was detected in 15.7% (22/140) of patients using FISH, in 28.6% (40/140) using CBA, and in 34.3% (48/140) of patients by combining both methods. Poor-prognosis CE (new deletion 17p, new deletion 11q or new complex karyotype) was detected in 15% (21/140) of patients and was significantly associated with previous CLL treatment (p=0.013). CBA provides more complex information about cytogenetic abnormalities in CLL patients than FISH and confirms that many patients can acquire new abnormalities during the course of their disease in a relatively short time period.
- MeSH
- aktivace lymfocytů účinky léků MeSH
- chronická lymfatická leukemie genetika patologie MeSH
- dospělí MeSH
- hybridizace in situ fluorescenční * MeSH
- interleukin-2 farmakologie MeSH
- klonální evoluce * MeSH
- lidé středního věku MeSH
- lidé MeSH
- oligodeoxyribonukleotidy farmakologie MeSH
- prospektivní studie MeSH
- pruhování chromozomů * MeSH
- senioři MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- hodnotící studie MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
Recent evidence suggests that complex karyotype (CK) defined by the presence of ≥3 chromosomal aberrations (structural and/or numerical) identified by using chromosome-banding analysis (CBA) may be relevant for treatment decision-making in chronic lymphocytic leukemia (CLL). However, many challenges toward the routine clinical application of CBA remain. In a retrospective study of 5290 patients with available CBA data, we explored both clinicobiological associations and the clinical impact of CK in CLL. We found that patients with ≥5 abnormalities, defined as high-CK, exhibit uniformly dismal clinical outcomes, independently of clinical stage, TP53 aberrations (deletion of chromosome 17p and/or TP53 mutations [TP53abs]), and the expression of somatically hypermutated (M-CLL) or unmutated immunoglobulin heavy variable genes. Thus, they contrasted with CK cases with 3 or 4 aberrations (low-CK and intermediate-CK, respectively) who followed aggressive disease courses only in the presence of TP53abs. At the other end of the spectrum, patients with CK and +12,+19 displayed an exceptionally indolent profile. Building upon CK, TP53abs, and immunoglobulin heavy variable gene somatic hypermutation status, we propose a novel hierarchical model in which patients with high-CK exhibit the worst prognosis, whereas those with mutated CLL lacking CK or TP53abs, as well as CK with +12,+19, show the longest overall survival. Thus, CK should not be axiomatically considered unfavorable in CLL, representing a heterogeneous group with variable clinical behavior. High-CK with ≥5 chromosomal aberrations emerges as prognostically adverse, independent of other biomarkers. Prospective clinical validation is warranted before ultimately incorporating high-CK in risk stratification of CLL.
- MeSH
- chromozomální aberace * MeSH
- chronická lymfatická leukemie genetika mortalita patologie MeSH
- cytogenetika metody MeSH
- lidé středního věku MeSH
- lidé MeSH
- míra přežití MeSH
- mutace * MeSH
- nádorové biomarkery genetika MeSH
- nádorový supresorový protein p53 genetika MeSH
- následné studie MeSH
- prognóza MeSH
- retrospektivní studie MeSH
- senioři MeSH
- somatická hypermutace imunoglobulinových genů genetika MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Východiska:Chromozomové změny patří u chronické lymfocytární leukemie (CLL) mezi významné prognostické faktory. Hlavní metodou využívanou k detekci těchto chromozomových změn je fluorescenční in situ hybridizace (FISH), klasické cytogenetické vyšetření vyžadující buňky v metafázi je u CLL problematické kvůli nízké proliferační schopnosti maligních B-lymfocytů in vitro. V roce 2006 byla publikována metoda umožňující získat metafázní B-lymfocyty u CLL s využitím stimulace pomocí IL‐2 a CpG oligonukleotidem DSP30. Cílem naší studie bylo ověřit účinnost stimulace a zhodnotit využitelnost této metody v rutinní praxi. Soubor pacientů a metody: Celkem u 369 pacientů s diagnózou CLL byla vyšetřována periferní krev klasickou cytogenetickou metodou a současně metodou FISH pro oblasti 13q14, 11q22–23, CEP12 a 17p13. Výsledky: Pro klasické cytogenetické vyšetření se stimulací podařilo získat metafázní buňky u 307 (83 %) z 369 pacientů. Chromozomové změny byly nalezeny u 243 (79 %) ze 307 hodnocených pacientů. Kromě aberací vyšetřovaných metodou FISH byly nalezeny další specifické změny v karyotypu např. del(6q), del(14q), t(14;18)(q32;q21), t(11;14)(q13;q32) a t(18;22)(q21;q11). U 103 (42 %) pacientů byly cytogenetickým vyšetřením nalezeny komplexní změny karyotypu, které metodou FISH detekovány nebyly. Závěr: Stimulace pomocí IL‐2 a oligonukleotidu DSP30 účinně navozuje dělení maligních B‐lymfocytů a umožňuje zachytit velké množství chromozomových změn u CLL, které by při vyšetřování metodou FISH pro základní čtyři aberace zůstaly skryty. Používání této metody v rutinní praxi se osvědčilo hlavně při identifikaci pacientů s komplexními změnami karyotypu.
Background: Chromosomal aberrations play an important role as prognostic factors in chronic lymphocytic leukemia (CLL). These aberrations are mostly detected by fluorescent in situ hybridization (FISH), as chromosomal banding analysis has been scarce due to low proliferative activity of malignant B-lymphocytes in vitro. In 2006, a new method using stimulation with IL-2 and CpG oligonucleotide DSP30 for metaphase generation in CLL was published [1]. The objective of our study was to verify the efficacy of stimulation and to evaluate if the method is suitable for routine diagnostics. Patients and Methods: In total, peripheral blood samples of 369 CLL patients were analyzed in parallel by chromosomal banding analysis and by FISH probes for 13q14, 11q22–23, CEP12 and 17p13. Results: Out of 369 patients, 307 (83%) were successfully stimulated for metaphase generation. Chromosomal aberrations were detected in 243 (79%) out of 307 patients evaluated by chromosomal banding analysis. Other aberrations that are not included into standard FISH panel were detected in patients’ karyotypes, e.g. del(6q), del(14q), t(14;18)(q32;q21), t(11;14)(q13;q32) and t(18;22)(q21;q11). One hundred and three (42%) patients showed complex aberrant karyotype not detected by FISH analysis. Conclusion: Stimulation with IL-2 and oligonucleotide DSP30 is an efficient method how to induce proliferation of malignant B-lymphocytes and allows detection of a substantial number of chromosomal aberrations in addition to those detected by standard FISH panel. Using this method in routine diagnostics is helpful particularly in identification of patients with complex aberrant karyotype.
- Klíčová slova
- CpG‐ODN DSP30,
- MeSH
- chromozomální aberace * MeSH
- chromozomální delece MeSH
- chronická lymfatická leukemie * genetika MeSH
- cytogenetické vyšetření * statistika a číselné údaje využití MeSH
- dospělí MeSH
- hybridizace in situ fluorescenční * MeSH
- interleukin-2 MeSH
- karyotyp MeSH
- krev MeSH
- kultivované buňky MeSH
- lidé středního věku MeSH
- lidé MeSH
- oligonukleotidy MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- translokace genetická 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
- práce podpořená grantem MeSH