AIM: Romania ranks first in Europe in terms of mortality from cervical cancer, recording 6.3 times more deaths than the mean in EU countries. Although vaccination campaigns were launched by health officials in Romania, the acceptance rate remained insignificant and programmes were discontinued. A successful vaccination programme requires a high rate of acceptance and accurate information for health professionals and parents. The aim of the study was to evaluate the level of parental knowledge about human papilomavirus (HPV) infection and HPV vaccination including the information obtained from general practitioners and identification of barriers in implementing a vaccination strategy. METHODS: We performed a cross-sectional study using a self-administered questionnaire for the parents of pupils in grades 5-8, in three randomly selected secondary schools in Tîrgu Mureş, Romania. RESULTS: We surveyed 918 parents. Of the respondents, 85.8% have heard of HPV infection. Most reported an average level of knowledge about HPV infection and HPV vaccination. The two main sources of information were specialized healthcare professionals (42.8% for HPV infection, 39.1% for HPV vaccination) and the Internet browsing (42.3% and 42.9%, respectively). Based on current knowledge, only one third of parents would have their child vaccinated against HPV infection. According to most parents surveyed, the main reasons for not wanting to have their child vaccinated is the fear of side effects; the vaccine is new and insufficiently studied; or parents do not know details about the vaccine. CONCLUSIONS: The parents showed the average level of knowledge about HPV infection and HPV vaccination. The implementation of an effective programme to increase the acceptance rate of HPV vaccination requires educational strategies aimed at involving parents and their children and supported by general practitioners and public health professionals.

• Klíčová slova
• HPV infection, HPV vaccination, cervical cancer, parents, questionnaire,
• MeSH
• dítě MeSH
• dospělí MeSH
• infekce papilomavirem epidemiologie   prevence a kontrola   MeSH
• lidé středního věku MeSH
• lidé MeSH
• nádory děložního čípku epidemiologie   prevence a kontrola   MeSH
• pacientův souhlas se zdravotní péčí MeSH
• průřezové studie MeSH
• průzkumy a dotazníky MeSH
• rodiče psychologie   MeSH
• vakcíny proti papilomavirům aplikace a dávkování   MeSH
• zdraví - znalosti, postoje, praxe MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Rumunsko epidemiologie   MeSH
• Názvy látek
• vakcíny proti papilomavirům MeSH

The EMDataResource Ligand Model Challenge aimed to assess the reliability and reproducibility of modeling ligands bound to protein and protein-nucleic acid complexes in cryogenic electron microscopy (cryo-EM) maps determined at near-atomic (1.9-2.5 Å) resolution. Three published maps were selected as targets: Escherichia coli beta-galactosidase with inhibitor, SARS-CoV-2 virus RNA-dependent RNA polymerase with covalently bound nucleotide analog and SARS-CoV-2 virus ion channel ORF3a with bound lipid. Sixty-one models were submitted from 17 independent research groups, each with supporting workflow details. The quality of submitted ligand models and surrounding atoms were analyzed by visual inspection and quantification of local map quality, model-to-map fit, geometry, energetics and contact scores. A composite rather than a single score was needed to assess macromolecule+ligand model quality. These observations lead us to recommend best practices for assessing cryo-EM structures of liganded macromolecules reported at near-atomic resolution.

• MeSH
• beta-galaktosidasa chemie   metabolismus   MeSH
• COVID-19 virologie   MeSH
• elektronová kryomikroskopie * metody   MeSH
• Escherichia coli MeSH
• konformace proteinů MeSH
• ligandy MeSH
• molekulární modely * MeSH
• reprodukovatelnost výsledků MeSH
• SARS-CoV-2 MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• beta-galaktosidasa MeSH
• ligandy MeSH

The EMDataResource Ligand Model Challenge aimed to assess the reliability and reproducibility of modeling ligands bound to protein and protein/nucleic-acid complexes in cryogenic electron microscopy (cryo-EM) maps determined at near-atomic (1.9-2.5 Å) resolution. Three published maps were selected as targets: E. coli beta-galactosidase with inhibitor, SARS-CoV-2 RNA-dependent RNA polymerase with covalently bound nucleotide analog, and SARS-CoV-2 ion channel ORF3a with bound lipid. Sixty-one models were submitted from 17 independent research groups, each with supporting workflow details. We found that (1) the quality of submitted ligand models and surrounding atoms varied, as judged by visual inspection and quantification of local map quality, model-to-map fit, geometry, energetics, and contact scores, and (2) a composite rather than a single score was needed to assess macromolecule+ligand model quality. These observations lead us to recommend best practices for assessing cryo-EM structures of liganded macromolecules reported at near-atomic resolution.

• Publikační typ
• časopisecké články MeSH
• preprinty MeSH

Helminths and bacteria are major players in the mammalian gut ecosystem and each influences the host immune system and health. Declines in helminth prevalence and bacterial diversity appear to play a role in the dramatic rise of immune mediated inflammatory diseases (IMIDs) in western populations. Helminths are potent modulators of immune system and their reintroduction is a promising therapeutic avenue for IMIDs. However, the introduction of helminths represents a disturbance for the host and it is important to understand the impact of helminth reintroduction on the host, including the immune system and gut microbiome. We tested the impact of a benign tapeworm, Hymenolepis diminuta, in a rat model system. We find that H. diminuta infection results in increased interleukin 10 gene expression in the beginning of the prepatent period, consistent with induction of a type 2 immune response. We also find induction of humoral immunity during the patent period, shown here by increased IgA in feces. Further, we see an immuno-modulatory effect in the small intestine and spleen in patent period, as measured by reductions in tissue immune cells. We observed shifts in microbiota community composition during the patent period (beta-diversity) in response to H. diminuta infection. However, these compositional changes appear to be minor; they occur within families and genera common to both treatment groups. There was no change in alpha diversity. Hymenolepis diminuta is a promising model for helminth therapy because it establishes long-term, stable colonization in rats and modulates the immune system without causing bacterial dysbiosis. These results suggest that the goal of engineering a therapeutic helminth that can safely manipulate the mammalian immune system without disrupting the rest of the gut ecosystem is in reach.

• MeSH
• Bacteria klasifikace   izolace a purifikace   MeSH
• biodiverzita MeSH
• biologické modely * MeSH
• feces chemie   MeSH
• fylogeneze MeSH
• Hymenolepis diminuta imunologie   MeSH
• imunitní systém * MeSH
• imunoglobulin A analýza   imunologie   MeSH
• interakce hostitele a parazita imunologie   MeSH
• interleukin-10 genetika   metabolismus   MeSH
• krysa rodu Rattus MeSH
• potkani Wistar MeSH
• regulace genové exprese imunologie   MeSH
• slezina imunologie   MeSH
• střeva mikrobiologie   parazitologie   MeSH
• střevní mikroflóra * MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• imunoglobulin A MeSH
• interleukin-10 MeSH

The clinical value of assessing immunoglobulin (Ig)G and IgA subclasses in addition to the isotypes of soluble Igs in serum has been well established. >20years ago, the International Union of Immunological Societies and the World Health Organization performed collaborative studies in order to validate antibody (Ab) clones for the detection of IgG and IgA subclasses for a broad range of laboratory assays, except for flow cytometry. Here we analyzed the performance of commercially available Ab clones to detect IgG and IgA subclasses in memory B-cells and plasma cells (PCs) by flow cytometry. In a first step, 28 Ab clones were evaluated in peripheral blood from healthy donors. Only 17/28 clones showed reactivity against IgG and IgA subclasses expressed on the B-cell and PC surface membrane, including Ab clones for IgG1 (SAG1, HP6188, HP6001 and HP6186), IgG2 (SAG2, HP6014 and HP6002), IgG3 (SAG3, HP6095 and HP6050), IgG4 (SAG4), IgA1 (SAA1, H69-11.4 and B3506B4) and IgA2 (SAA2, 2E2, and A9604D2). In a second step, for each Ig subclass a single clone was selected according to its specificity and fluorescence intensity (resolution power), for further more detailed validation (SAG1, SAG2, SAG3, SAG4, SAA1 and SAA2). This validation process was carried out in 4 different laboratories by testing the selected Ab clones in human peripheral blood, bone marrow and tonsil samples, using different staining protocols (e.g. surface membrane and/or cytoplasmic staining). All selected Ab clones displayed strong positivity, high specificity and optimal resolution between negative and positive cells. Alternative Ab clones were also validated. Thus, our results show the feasibility of using the validated Ig subclass Ab clones in combination with other B cell-associated markers for detailed dissection of the memory B-cell and PC compartments that express distinct Ig subclasses in different human tissues.

• Klíčová slova
• Antibody clones, EuroFlow, Flow cytometry, IgG and IgA subclasses, Immune monitoring, Immunophenotyping,
• MeSH
• B-lymfocyty imunologie   MeSH
• dospělí MeSH
• imunofenotypizace metody   MeSH
• imunoglobulin A analýza   MeSH
• imunoglobulin G analýza   MeSH
• lidé středního věku MeSH
• lidé MeSH
• monoklonální protilátky * MeSH
• plazmatické buňky imunologie   MeSH
• předškolní dítě MeSH
• průtoková cytometrie metody   normy   MeSH
• senioři MeSH
• specificita protilátek * MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• validační studie MeSH
• Názvy látek
• imunoglobulin A MeSH
• imunoglobulin G MeSH
• monoklonální protilátky * MeSH

The fluorescence detected using fluorochrome-labelled monoclonal antibodies depends not only on the abundance of the target antigen, but amongst many other factors also on the effective fluorochrome-to-antibody ratio. The diagnostic approach of the EuroFlow consortium relies on reproducible fluorescence intensities over time. A capture bead system for mouse immunoglobulin light chains was utilized to compare the mean fluorescence intensity of 1323 consecutive antibody lots to the currently used lot of the same monoclonal antibody. In total, 157 different monoclonal antibodies were assessed over seven years. Median relative difference between consecutive lots was 3.8% (range: 0.01% to 164.7%, interquartile range: 1.3% to 10.1%). The relative difference exceeded 20% in 8.8% of all comparisons. FITC labelled monoclonal antibodies (median relative difference: 2.1%) showed a significantly smaller variation between lots than antibodies conjugated to PE (3.5%), PECy7 (3.9%), PerCPCy5.5 (5.8%), APC (5.8%), APCH7 (7.4%), and APCC750 (14.5%). Reagents labelled with Pacific Blue (1.4%), Pacific Orange (2.4%), HV450 (0.7%), and HV500 (1.7%) demonstrated more consistent results compared to conjugates of BV421 (4.1%) and BV510 (16.2%). Additionally, significant differences in lot-to-lot fluorescence stability amongst antibodies labelled with the same fluorochrome were observed between manufacturers. These observations might guide future quality recommendations for the production and application of fluorescence-labelled monoclonal antibodies in multicolor flow cytometry.

• Klíčová slova
• Flow cytometry, Fluorescence intensity, Fluorochrome-to-antibody ratio, Variability,
• MeSH
• fluorescenční barviva * normy   MeSH
• monoklonální protilátky * MeSH
• myši MeSH
• průtoková cytometrie metody   normy   MeSH
• reprodukovatelnost výsledků MeSH
• stabilita proteinů MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fluorescenční barviva * MeSH
• monoklonální protilátky * MeSH

A fully-standardized EuroFlow 8-color antibody panel and laboratory procedure was stepwise designed to measure minimal residual disease (MRD) in B-cell precursor (BCP) acute lymphoblastic leukemia (ALL) patients with a sensitivity of ≤10-5, comparable to real-time quantitative polymerase chain reaction (RQ-PCR)-based MRD detection via antigen-receptor rearrangements. Leukocyte markers and the corresponding antibodies and fluorochromes were selected based on their contribution in separating BCP-ALL cells from normal/regenerating BCP cells in multidimensional principal component analyses. After 5 multicenter design-test-evaluate-redesign phases with a total of 319 BCP-ALL patients at diagnosis, two 8-color antibody tubes were selected, which allowed separation between normal and malignant BCP cells in 99% of studied patients. These 2 tubes were tested with a new erythrocyte bulk-lysis protocol allowing acquisition of high cell numbers in 377 bone marrow follow-up samples of 178 BCP-ALL patients. Comparison with RQ-PCR-based MRD data showed a clear positive relation between the percentage concordant cases and the number of cells acquired. For those samples with >4 million cells acquired, concordant results were obtained in 93% of samples. Most discordances were clarified upon high-throughput sequencing of antigen-receptor rearrangements and blind multicenter reanalysis of flow cytometric data, resulting in an unprecedented concordance of 98% (97% for samples with MRD < 0.01%). In conclusion, the fully standardized EuroFlow BCP-ALL MRD strategy is applicable in >98% of patients with sensitivities at least similar to RQ-PCR (≤10-5), if sufficient cells (>4 × 106, preferably more) are evaluated.

• MeSH
• dítě MeSH
• dospělí MeSH
• genová přestavba MeSH
• kojenec MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• novorozenec MeSH
• pre-B-buněčná leukemie diagnóza   MeSH
• předškolní dítě MeSH
• průtoková cytometrie metody   normy   MeSH
• receptory antigenů B-buněk genetika   MeSH
• reziduální nádor diagnóza   MeSH
• senioři MeSH
• senzitivita a specificita MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• kojenec MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• práce podpořená grantem MeSH
• Názvy látek
• receptory antigenů B-buněk MeSH

Reproducible expert-independent flow-cytometric criteria for the differential diagnoses between mature B-cell neoplasms are lacking. We developed an algorithm-driven classification for these lymphomas by flow cytometry and compared it to the WHO gold standard diagnosis. Overall, 662 samples from 662 patients representing 9 disease categories were analyzed at 9 laboratories using the previously published EuroFlow 5-tube-8-color B-cell chronic lymphoproliferative disease antibody panel. Expression levels of all 26 markers from the panel were plotted by B-cell entity to construct a univariate, fully standardized diagnostic reference library. For multivariate data analysis, we subsequently used canonical correlation analysis of 176 training cases to project the multidimensional space of all 26 immunophenotypic parameters into 36 2-dimensional plots for each possible pairwise differential diagnosis. Diagnostic boundaries were fitted according to the distribution of the immunophenotypes of a given differential diagnosis. A diagnostic algorithm based on these projections was developed and subsequently validated using 486 independent cases. Negative predictive values exceeding 92.1% were observed for all disease categories except for follicular lymphoma. Particularly high positive predictive values were returned in chronic lymphocytic leukemia (99.1%), hairy cell leukemia (97.2%), follicular lymphoma (97.2%), and mantle cell lymphoma (95.4%). Burkitt and CD10+ diffuse large B-cell lymphomas were difficult to distinguish by the algorithm. A similar ambiguity was observed between marginal zone, lymphoplasmacytic, and CD10- diffuse large B-cell lymphomas. The specificity of the approach exceeded 98% for all entities. The univariate immunophenotypic library and the multivariate expert-independent diagnostic algorithm might contribute to increased reproducibility of future diagnostics in mature B-cell neoplasms.

• MeSH
• difúzní velkobuněčný B-lymfom * MeSH
• dospělí MeSH
• folikulární lymfom * diagnóza   MeSH
• imunofenotypizace MeSH
• lidé MeSH
• průtoková cytometrie metody   MeSH
• reprodukovatelnost výsledků MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• práce podpořená grantem MeSH

Presence of minimal residual disease (MRD), detected by flow cytometry, is an important prognostic biomarker in the management of B-cell precursor acute lymphoblastic leukemia (BCP-ALL). However, data-analysis remains mainly expert-dependent. In this study, we designed and validated an Automated Gating & Identification (AGI) tool for MRD analysis in BCP-ALL patients using the two tubes of the EuroFlow 8-color MRD panel. The accuracy, repeatability, and reproducibility of the AGI tool was validated in a multicenter study using bone marrow follow-up samples from 174 BCP-ALL patients, stained with the EuroFlow BCP-ALL MRD panel. In these patients, MRD was assessed both by manual analysis and by AGI tool supported analysis. Comparison of MRD levels obtained between both approaches showed a concordance rate of 83%, with comparable concordances between MRD tubes (tube 1, 2 or both), treatment received (chemotherapy versus targeted therapy) and flow cytometers (FACSCanto versus FACSLyric). After review of discordant cases by additional experts, the concordance increased to 97%. Furthermore, the AGI tool showed excellent intra-expert concordance (100%) and good inter-expert concordance (90%). In addition to MRD levels, also percentages of normal cell populations showed excellent concordance between manual and AGI tool analysis. We conclude that the AGI tool may facilitate MRD analysis using the EuroFlow BCP-ALL MRD protocol and will contribute to a more standardized and objective MRD assessment. However, appropriate training is required for the correct analysis of MRD data.

• Klíčová slova
• BCP‐ALL, B‐cell differentiation, MRD, automation, database,
• MeSH
• automatizace MeSH
• dítě MeSH
• dospělí MeSH
• hematopoéza * MeSH
• imunofenotypizace metody   MeSH
• kojenec MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• pre-B-buněčná leukemie * patologie   diagnóza   MeSH
• předškolní dítě MeSH
• průtoková cytometrie * metody   MeSH
• reprodukovatelnost výsledků MeSH
• reziduální nádor * patologie   diagnóza   MeSH
• senioři MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• kojenec MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH

BACKGROUND: Following the publication of the European consensus statement on standards for essential colposcopy in 2020, the need for standards relating to more complex and challenging colposcopy practice was recognised. These standards relate to colposcopy undertaken in patients identified through cervical screening and tertiary referrals from colposcopists who undertake standard colposcopy only. This set of recommendations provides a review of the current literature and agreement on care for recognised complex cases. With good uptake of human papillomavirus (HPV) immunisation, we anticipate a marked reduction in cervical disease over the next decade. Still, the expert colposcopist will continue to be vital in managing complex cases, including previous cervical intraepithelial neoplasia (CIN)/complex screening histories and multi-zonal disease. AIMS: To provide expert guidance on complex colposcopy cases through published evidence and expert consensus. MATERIAL & METHODS: Members of the EFC and ESGO formed a working group to identify topics considered to be the remit of the expert rather than the standard colposcopy service. These were presented at the EFC satellite meeting, Helsinki 2021, for broader discussion and finalisation of the topics. RESULTS & DISCUSSION: The agreed standards included colposcopy in pregnancy and post-menopause, investigation and management of glandular abnormalities, persistent high-risk HPV+ with normal/low-grade cytology, colposcopy management of type 3 transformation zones (TZ), high-grade cytology and normal colposcopy, colposcopy adjuncts, follow-up after treatment with CIN next to TZ margins and follow-up after treatment with CIN with persistent HPV+, and more. These standards are under review to create a final paper of consensus standards for dissemination to all EFC and ESGO members.

• Klíčová slova
• Cancer, Cervix, Colposcopy, European guideline, Screening,
• MeSH
• časná detekce nádoru MeSH
• dysplazie děložního hrdla * diagnóza   MeSH
• infekce papilomavirem * diagnóza   MeSH
• kolposkopie MeSH
• lidé MeSH
• nádory děložního čípku * MeSH
• Papillomaviridae MeSH
• těhotenství MeSH
• Check Tag
• lidé MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• konsensus - konference MeSH
• přehledy MeSH