UNLABELLED: The aim of this study was to identify parameters influencing DNA extraction and PCR amplification efficiencies in an attempt to standardize Mucorales qPCR. The Fungal PCR Initiative Mucorales Laboratory Working Group distributed two panels of simulated samples to 26 laboratories: Panel A (six sera spiked with Mucorales DNA and one negative control serum) and Panel B (six Mucorales DNA extracts). Panel A underwent DNA extraction in each laboratory according to the local procedure and were sent to a central laboratory for testing using three different qPCR techniques: one in-house qPCR assay and two commercial assays (MucorGenius and Fungiplex). Panel B DNA extracts were PCR amplified in each laboratory using local procedures: nine in-house qPCR assays and two commercial kits (MucorGenius and MycoGENIE). All data were compiled and anonymously analyzed at the central laboratory. For Panel A, a total of six different automated platforms and five manual extraction methods were used. Positive rates were 64%, 70%, and 89%, for the MucorGenius, Fungiplex, and the in-house qPCR assay, respectively. Using a large volume of serum for DNA extraction provided the highest analytical sensitivity (82.5% for 1 mL compared with 62.7% for smaller volumes, P < 0.01). For Panel B, five in-house qPCR assays and two commercial kits had >78% positivity. Using larger PCR input volumes (≥7 μL) was associated with the highest sensitivity at 95.5% compared to 58.3% when lower input volumes were used (P < 0.01). Using larger sample volumes for nucleic acid extraction and DNA template volumes for PCR amplification significantly improves the performance of Mucorales qPCR when testing serum. IMPORTANCE: Mucormycosis is a life-threatening mold infection affecting immunosuppressed patients but also other patients with diabetes or trauma. Better survival is linked to shorter delays in diagnosis and treatment initiation. Detection of Mucorales-free DNA in serum or plasma using quantitative PCR allows a prompt diagnosis and earlier treatment. Several techniques and protocols of quantitative Mucorales PCR are used in Europe, and improving performance remains a common objective of laboratories participating in the fungal PCR Initiative Working Group. This study, which combined results from 26 laboratories in Europe, showed that the main parameters underpinning sensitivity are the preanalytical variables (volume of serum used for DNA extraction and DNA template volume), irrespective of the extraction platforms and qPCR assay/platform.

• MeSH
• diagnostické techniky molekulární normy   metody   MeSH
• DNA fungální * krev   genetika   MeSH
• kvantitativní polymerázová řetězová reakce * normy   metody   MeSH
• lidé MeSH
• Mucorales * genetika   izolace a purifikace   MeSH
• mukormykóza * diagnóza   mikrobiologie   krev   MeSH
• senzitivita a specificita * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH

Brucellosis is a zoonosis with non-specific clinical symptoms involving multiple systems and organs. Its prevalence is low in most of EU countries, which can lead to the difficulties in laboratory and clinical diagnostic. Due to its relationship to the Ochrobactrum spp., it may be misclassified in rapid identification systems. We present a case of a 13-year-old immunocompetent girl who was examined several times for fever, fatigue, night sweats and weight loss; laboratory results showed mildly elevated C-reactive protein, anaemia and leukopenia. Four weeks before the onset of symptoms, she had been on a family holiday in Egypt. Given her symptoms, a haemato-oncological or autoimmune disease was considered more likely. The diagnosis of Brucella spondylitis was made after 4 months. The main reasons for this delay were as follows: low specificity of clinical symptoms, delay in completing the travel history, inconclusive initial serological results and misidentification of the blood culture isolate as Ochrobactrum sp. Even in countries with a low incidence of brucellosis, it is essential to educate healthcare professionals about the disease. Low specificity of symptoms and limited experience of laboratory staff may lead to late diagnosis with risk of complications and poor outcome. If Ochrobactrum spp. is detected in clinical specimens by rapid identification, careful re-evaluation must follow and all measures to prevent laboratory-acquired infections must be taken until Brucella spp. is unequivocally excluded.

• MeSH
• bakteriemie * diagnóza   mikrobiologie   MeSH
• Brucella izolace a purifikace   klasifikace   MeSH
• brucelóza * diagnóza   mikrobiologie   MeSH
• chybná diagnóza * MeSH
• gramnegativní bakteriální infekce diagnóza   mikrobiologie   MeSH
• horečka * mikrobiologie   etiologie   MeSH
• lidé MeSH
• mladiství MeSH
• Ochrobactrum * genetika   izolace a purifikace   MeSH
• spondylitida mikrobiologie   diagnóza   MeSH
• Check Tag
• lidé MeSH
• mladiství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• Geografické názvy
• Egypt MeSH

Penile squamous cell carcinoma (pSCC) represents an uncommon malignancy characterized by stagnant mortality, psychosexual distress, and a highly variable prognosis. Currently, the World Health Organization distinguishes between human papillomavirus (HPV)-related and HPV-independent pSCC. Recently, there has been an evolving line of research documenting the enrichment of HPV-independent pSCC with a high tumor mutational burden (TMB) and programmed death ligand-1 expression, as well as clusters of genes associated with HPV status. In this study, we conducted comprehensive next-generation sequencing DNA profiling of 146 pSCC samples using a panel consisting of 355 genes associated with tumors. This profiling was correlated with immunohistochemical markers and prognostic clinical data. A survival analysis of recurrent genomic events (found in ≥10 cases) was performed. TP53, CDKN2A, ATM, EPHA7, POT1, CHEK1, GRIN2A, and EGFR alterations were associated with significantly shortened overall survival in univariate and multivariate analysis. HPV positivity, diagnosed through both p16 immunohistochemistry and HPV DNA analysis, displayed no impact on survival but was associated with high-grade, lymphatic invasion, programmed death ligand-1 negativity/weak expression, and low TMB. FAT1, TP53, CDKN2A, CASP8, and HRAS were more often mutated in HPV-independent pSCC. In contrast, HPV-associated pSCCs were enriched by EPHA7, ATM, GRIN2A, and CHEK1 mutations. PIK3CA, FAT1, FBXW7, and KMT2D mutations were associated with high TMB. NOTCH1, TP53, CDKN2A, POT1, KMT2D, ATM, CHEK1, EPHA3, and EGFR alterations were related to adverse clinicopathologic signs, such as advanced stage, high tumor budding, and lymphovascular invasion. We detected 160 alterations with potential treatment implications, with 21.2% of samples showing alterations in the homologous recombination repair pathway. To the best of our knowledge, this study describes the largest cohort of pSCC with complex molecular pathologic, clinical, and prognostic analysis correlating with prognosis.

• MeSH
• ATM protein genetika   MeSH
• dospělí MeSH
• erbB receptory genetika   MeSH
• infekce papilomavirem MeSH
• inhibitor p16 cyklin-dependentní kinasy genetika   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mutace MeSH
• nádorové biomarkery * genetika   analýza   MeSH
• nádorový supresorový protein p53 genetika   MeSH
• nádory penisu * genetika   mortalita   patologie   virologie   MeSH
• prognóza MeSH
• proteiny vázající telomery MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• shelterinový komplex MeSH
• spinocelulární karcinom * genetika   mortalita   patologie   virologie   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• Publikační typ
• časopisecké články MeSH

The aim of this study is to evaluate opportunistic pathogenic bacteria of the genus Pseudomonas in anthropogenically impacted bathing waters, primarily focusing on bathing ponds. The findings include the detection of these bacteria, their susceptibility to selected antibiotics, and the determination of the Exotoxin A (exoA) gene using PCR method. P. aeruginosa was present in most samples, albeit in low concentrations (1-14 CFU/100 mL). The presence of P. otitidis, which is associated with ear infection, in this type of bathing water, was not rare (up to 90 CFU/100 mL). This species would not be detected by the standard methods, including tests on acetamid medium, used for P. aeruginosa in water. The isolated strains of P. otitidis lack the exoA gene and exhibited higher resistance to meropenem compared to P. aeruginosa.

• MeSH
• ADP-ribosatransferasy genetika   MeSH
• antibakteriální látky * farmakologie   MeSH
• bakteriální léková rezistence MeSH
• bakteriální proteiny genetika   MeSH
• bakteriální toxiny genetika   MeSH
• exotoxin A z Pseudomonas aeruginosa MeSH
• exotoxiny genetika   MeSH
• faktory virulence genetika   MeSH
• mikrobiální testy citlivosti * MeSH
• mikrobiologie vody * MeSH
• polymerázová řetězová reakce MeSH
• Pseudomonas * genetika   izolace a purifikace   klasifikace   účinky léků   MeSH
• rybníky * mikrobiologie   MeSH
• Publikační typ
• časopisecké články MeSH

Cervical cancer (CC) is the fourth most common malignant tumor in women worldwide. Detecting different biomarkers together on single cells by novel method mass cytometry could contribute to more precise screening. Liquid-based cytology (LBC) cervical samples were collected (N = 53) from women categorized as normal and precancerous lesions. Human papillomavirus was genotyped by polymerase chain reaction, while simultaneous examination of the expression of 29 proteins was done by mass cytometry (CyTOF). Differences in cluster abundances were assessed with Spearman's rank correlation as well as high dimensional data analysis (t-SNE, FlowSOM). Cytokeratin (ITGA6, Ck5, Ck10/13, Ck14, Ck7) expression patterns allowed determining the presence of different cells in the cervical epithelium. FlowSOM analysis enabled to phenotype cervical cells in five different metaclusters and find new markers that could be important in CC screening. The markers Ck18, Ck18, and CD63 (Metacluster 3) showed significantly increasing associated with severity of the precancerous lesions (Spearman rank correlation rho 0.304, p = 0.0271), while CD71, KLF4, LRIG1, E-cadherin, Nanog and p53 (Metacluster 1) decreased with severity of the precancerous lesions (Spearman rank correlation rho -0.401, p = 0.0029). Other metaclusters did not show significant correlation, but metacluster 2 (Ck17, MCM, MMP7, CD29, E-cadherin, Nanog, p53) showed higher abundance in low- and high-grade intraepithelial lesion cases. CyTOF appears feasible and should be considered when examining novel biomarkers on cervical LBC samples. This study enabled us to characterize different cells in the cervical epithelium and find markers and populations that could distinguish precancerous lesions.

• MeSH
• cervix uteri patologie   metabolismus   MeSH
• dospělí MeSH
• dysplazie děložního hrdla diagnóza   patologie   MeSH
• infekce papilomavirem patologie   diagnóza   virologie   MeSH
• Krüppel-like faktor 4 * MeSH
• lidé středního věku MeSH
• lidé MeSH
• nádorové biomarkery * genetika   metabolismus   MeSH
• nádory děložního čípku * diagnóza   patologie   genetika   MeSH
• prekancerózy * patologie   diagnóza   MeSH
• průtoková cytometrie * metody   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Acute kidney injury (AKI) due to gentamicin nephrotoxicity is a significant concern in clinical medicine, particularly in patients receiving prolonged or high-dose gentamicin therapy. Gentamicin is an aminoglycoside antibiotic frequently used in the treatment of a range of bacterial infections. However, its use is associated with nephrotoxicity which can manifest as AKI. Due to this, it is crucial to diagnose promptly and manage treatment effectively. Ongoing studies are therefore focusing on non-protein-coding RNAs as potential biomarkers for AKI. Numerous microRNAs (miRNAs) have been implicated in gentamicin-induced nephrotoxicity and AKI. They participate in pathways associated with inflammation, cell death, and oxidative stress and each of these factors play critical roles in the development of gentamicin-induced kidney injury. Research studies have demonstrated changes in the expression levels of these miRNAs in response to gentamicin exposure both in vitro and in in vivo models, as well as in human clinical trials involving patients receiving gentamicin therapy. The dysregulation of these miRNAs correlates with the severity of kidney injury and may serve as sensitive biomarkers for early detection and monitoring of AKI induced by gentamicin.

• MeSH
• akutní poškození ledvin * chemicky indukované   diagnóza   MeSH
• antibakteriální látky * škodlivé účinky   MeSH
• biologické markery * MeSH
• gentamiciny * škodlivé účinky   MeSH
• lidé MeSH
• mikro RNA * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Laboratorní diagnostika infekčního agens je založena na metodách přímého průkazu (kultivace, mikroskopie, PCR, antigenní test) a na metodách nepřímého průkazu (detekce protilátek). Sérologie zůstává důležitou součástí laboratorní diagnostiky infekčních onemocnění, přestože význam některých sérologických testů klesá s rozvojem molekulárně biologických metod. Pro pochopení, jakou roli hrají protilátky v procesu imunitní odpovědi na infekční agens, je nutné znát obecné mechanismy, které jsou s tvorbou protilátek spojené. Interpretace sérologických výsledků je složitá a závisí na fázi imunitní odpovědi, antigenních vlastnostech patogenu, použité metodě, senzitivitě a specificitě testů a klinickém kontextu konkrétního pacienta. Správná interpretace sérologických testů vyžaduje hlubokou znalost patogeneze infekce (specifická reakce na bakterie, viry nebo prvoky), metodických limitací a klinických souvislostí, což je zásadní pro efektivní diagnostiku a léčbu pacientů v klinické praxi.

Martinek J, Lochmanová A, Maďar R. Interpretation of serological results in the diagnosis of infectious diseases Laboratory diagnosis of infectious agents is based on direct detection methods (culture, microscopy, PCR, antigen test) and indirect detection methods (antibody detection). Serology remains an important part of the laboratory diagnosis of infectious diseases, although the importance of some serological tests is declining with the development of molecular biological methods. To understand the role of antibodies in the immune response to infectious agents, it is necessary to know the general mechanisms involved in antibody production. Interpretation of serological results is complex and depends on the phase of the immune response, the antigenic properties of the pathogen, the method used, the sensitivity and specificity of the tests, and the clinical context of the individual patient. Correct interpretation of serological tests requires an in-depth knowledge of the pathogenesis of infection (specific response to bacteria, viruses or protozoa), methodological limitations and clinical context, which is essential for effective diagnosis and treatment of patients in clinical practice.

BackgroundDuring the COVID-19 pandemic, non-pharmaceutical interventions (NPIs) such as social distancing, lockdowns and enhanced hygiene led to a decrease in respiratory pathogens. However, as NPIs were relaxed, a resurgence in several respiratory pathogens was observed including one local Chlamydia pneumoniae outbreak in Switzerland, prompting the need for a better understanding of C. pneumoniae epidemiology.AimTo assess temporal and geographical variations in C. pneumoniae detection before, during and after the COVID-19 pandemic.MethodsData on C. pneumoniae PCR detection ratios (number of positive tests/ total number of tests) across pre-pandemic (2018-2019), pandemic (2020-2022) and post-pandemic (2023) periods were collected via a global survey disseminated through various professional networks.ResultsC. pneumoniae detection ratios were analysed across 28 sites (27 in Europe, one in Taiwan) in 2023 (Dataset A, n = 172,223 tests) and 20 sites from 2018 to 2023 (Dataset B, n = 693,106 tests). Twenty-seven sites were laboratories (hospital or clinical) and one a surveillance system (Denmark). A significant decrease in detection ratios was observed during the pandemic period (from 1.05% to 0.23%, p < 0.001). In 2023, detection ratios increased to 0.28% (p < 0.002). Notable regional variations were found, with statistically significant increases in detection ratios at six sites located in Switzerland and Slovenia, where ratios ranged from 0.52% to 3.25%.DiscussionThe study highlights how NPIs influenced C. pneumoniae epidemiology, with reduced detection during the pandemic and partial resurgence afterwards. Regional variations suggest differing NPI impacts and underscore the need for continued surveillance.

• MeSH
• Chlamydophila pneumoniae * izolace a purifikace   genetika   MeSH
• COVID-19 * epidemiologie   MeSH
• infekce bakteriemi rodu Chlamydophila * epidemiologie   diagnóza   MeSH
• lidé MeSH
• pandemie MeSH
• polymerázová řetězová reakce MeSH
• SARS-CoV-2 MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Evropa MeSH
• Taiwan MeSH

BackgroundOn 29 January 2024, the European Centre for Disease Prevention and Control distributed an alert about a metronidazole-resistant Clostridioides difficile outbreak of PCR ribotype (RT) 955 in England.AimWe aimed to investigate the presence of RT955 in Czech, Slovak and Polish C. difficile isolates and evaluate different culture media for detecting its metronidazole resistance.MethodsIsolates with binary toxin genes identified as 'unknown' by the WEBRIBO PCR ribotyping database up to 2023 were re-analysed after adding the RT955 profile to the database. The RT955 isolates were characterised by whole genome sequencing and tested for susceptibility to 15 antimicrobials.ResultsWe did not find RT955 in Czech (n = 6,661, 2012-2023) and Slovak (n = 776, 2015-2023) isolates, but identified 13 RT955 cases (n = 303, 2021-2023) in three hospitals in Poland. By whole genome multilocus sequence typing, 10 isolates clustered into one clonal complex including a sequence of United Kingdom strain ERR12670107, and shared similar antimicrobial resistance genes/mutations. All 13 isolates were resistant to ciprofloxacin/moxifloxacin, erythromycin/clindamycin and ceftazidime. All isolates had a mutation in the nimB gene promoter and in NimB (Tyr130Ser and Leu155Ile). The metronidazole resistance was detected in all isolates using brain-heart-infusion agar supplemented with haemin and Chocolate agar. Results were discrepant with the European Committee on Antimicrobial Susceptibility Testing-recommended Fastidious anaerobe agar and Brucella blood agar.ConclusionThe identification of clonally related haem-dependent metronidazole-resistant C. difficile RT955 in multiple hospitals indicates a need for prospective surveillance to estimate its prevalence in Europe.

• MeSH
• antibakteriální látky * farmakologie   MeSH
• bakteriální léková rezistence * genetika   MeSH
• Clostridioides difficile * genetika   účinky léků   izolace a purifikace   klasifikace   MeSH
• epidemický výskyt choroby MeSH
• klostridiové infekce * epidemiologie   mikrobiologie   farmakoterapie   MeSH
• lidé MeSH
• metronidazol * farmakologie   MeSH
• mikrobiální testy citlivosti MeSH
• multilokusová sekvenční typizace MeSH
• polymerázová řetězová reakce MeSH
• ribotypizace * MeSH
• sekvenování celého genomu MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Česká republika MeSH
• Polsko MeSH
• Slovenská republika MeSH

Rationale: Early detection, standardized therapy, adequate infrastructure, and strategies for quality improvement should constitute essential components of every hospital's sepsis plan. Objectives: To investigate the extent to which recommendations from the sepsis guidelines are implemented and the availability of infrastructure for the care of patients with sepsis in acute-care hospitals. Methods: A multidisciplinary cross-sectional questionnaire was used to investigate sepsis care in hospitals. This included the use of sepsis definitions, the implementation of sepsis guideline recommendations, diagnostic and therapeutic infrastructure, antibiotic stewardship, and quality improvement initiatives (QIIs) in hospitals. Measurements and Main Results: A total of 1,023 hospitals in 69 countries were included. Most of them, 835 (81.6%), were in Europe. Sepsis screening was used in 54.2% of emergency departments (EDs), 47.9% of wards, and 61.7% of ICUs. Sepsis management was standardized in 57.3% of EDs, 45.2% of wards, and 70.7% of ICUs. The implementation of comprehensive QIIs was associated with increased screening (EDs, +33.3%; wards, +44.4%; ICUs, +23.8% absolute difference) and increased standardized sepsis management (EDs, +33.6%; wards, +40.0%; ICUs, +17.7% absolute difference) compared with hospitals without QIIs. A total of 9.8% of hospitals had implemented ongoing QIIs, and 4.6% had invested in sepsis programs. Conclusions: The findings indicate that there is considerable room for improvement in a large number of mainly European hospitals, particularly with regard to early identification and standardized management of sepsis, the availability of guidelines, diagnostic and therapeutic infrastructure, and the implementation of QIIs. Further efforts are required to implement a more comprehensive and appropriate quality of care.

• MeSH
• antibiotická politika MeSH
• dodržování směrnic * statistika a číselné údaje   MeSH
• jednotky intenzivní péče normy   MeSH
• lidé MeSH
• nemocnice * normy   statistika a číselné údaje   MeSH
• průřezové studie MeSH
• průzkumy a dotazníky MeSH
• sepse * terapie   diagnóza   MeSH
• směrnice pro lékařskou praxi jako téma MeSH
• urgentní služby nemocnice normy   MeSH
• zlepšení kvality * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Evropa MeSH