Gene sequencing of 16S, 18S, and ITS regions is a crucial tool in molecular diagnostics, especially in microbiology, pathology and forensic medicine. These genes contain conserved and variable regions and are widely used for the taxonomic classification of bacteria and eukaryotes. Sequencing of 16S rDNA helps detect bacterial infections, while sequencing of ITS regions and 18S rDNA is used to identify fungal or parasitic infections, especially when traditional methods are ineffective. This article focuses on the expanded possibilities of these methods, their application in clinical diagnostics and research, their advantages and disadvantages, and discusses potential future developments in the field of next-generation sequencing (NGS) technology.
- Klíčová slova
- 16S sequencing, 18S sequencing, ITS region sequencing, Infection diagnostics, Molecular pathology, Next-generation sequencing, next generation sequencing,
- MeSH
- lidé MeSH
- RNA ribozomální 16S * genetika MeSH
- RNA ribozomální 18S genetika MeSH
- sekvenční analýza DNA MeSH
- vysoce účinné nukleotidové sekvenování MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
- Názvy látek
- RNA ribozomální 16S * MeSH
- RNA ribozomální 18S MeSH
The ribotyping of Clostridioides difficile is one of the basic methods of molecular epidemiology for monitoring the spread of C. difficile infections. In the Czech Republic, this procedure is mainly available in university hospitals. The introduction of ribotyping in a tertiary health care facility such as Liberec Regional Hospital not only increases safety in the facility but also supports regional professional development. In our study, 556 stool samples collected between June 2017 and June 2018 were used for C. difficile infection screening, followed by cultivation, toxinotyping, and ribotyping of positive samples. The toxinotyping of 96 samples revealed that 44.8% of typed strains could produce toxins A and B encoded by tcdA and tcdB, respectively. The ribotyping of the same samples revealed two epidemic peaks, caused by the regionally most prevalent ribotype 176 (n = 30, 31.3). C. difficile infection incidence ranged between 5.5 and 4.2 cases per 10,000 patient-bed days. Molecular diagnostics and molecular epidemiology are the two most developing parts of clinical laboratories. The correct applications of molecular methods help ensure greater safety in hospitals.
- Klíčová slova
- Clonal spreading, Clostridioides difficile, Health care facility, Ribotyping, Toxinotyping,
- MeSH
- bakteriální toxiny * MeSH
- Clostridioides difficile * genetika MeSH
- Clostridioides MeSH
- klostridiové infekce * diagnóza epidemiologie MeSH
- lidé MeSH
- nemocnice univerzitní MeSH
- poskytování zdravotní péče MeSH
- ribotypizace MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- bakteriální toxiny * MeSH
The current progress and increasing knowledge about the genetic causes of cancer opens up new possibilities for its treatment. However, it is necessary to combine the results obtained using classical pathological methods with sensitive, multiplex molecular pathological methods. The method that meets the required criteria is MLPA based on multiplex PCR reaction. This method detects both changes in gene copy number and DNA methylation and, last but not least, point mutations. The MLPA reaction is applicable to even highly fragmented DNA. At the same time, it is a robust method that can be performed on standard thermocyclers, the fluorescent tip label requires automatic sequencers. Up to 50 genetic markers can be tested in one reaction, a number that allows a diagnostic and prognostic conclusion. All these features lead to the routine use of MLPA analysis not only in diagnosis but also in cancer research. The present article aims to summarize the different types of MLPA reactions, its benefits, but also the potential pitfalls.
- Klíčová slova
- MLPA, Molecular pathology, PCR, Tumors of the central nervous system,
- MeSH
- DNA genetika MeSH
- genetické markery MeSH
- genová dávka MeSH
- lidé MeSH
- metylace DNA * MeSH
- nádory centrálního nervového systému * genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- DNA MeSH
- genetické markery MeSH
The aim of the presented communication is to clearly inform the general professional public about the newly approved modifications in this classification, including the newly approved types of tumours. A significant change is the new grading system for these tumours, including the innovative involvement of tumour profiling at the molecular level in the system for determining the degree of tumour differentiation and the application of the principle of integrated diagnostics, i. e. the synthesis of available histopathological and molecular findings in CNS tumors.
- Klíčová slova
- Molecular pathology, WHO classification, grading, tumours of the central nervous system,
- MeSH
- lidé MeSH
- nádory centrálního nervového systému * diagnóza MeSH
- Světová zdravotnická organizace MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Halogenated organic compounds are naturally occurring in subsurface environments; however, accumulation of the degradative intermediate cis-1,2-dichloroethene (cDCE) at soil and groundwater sites contaminated with xenobiotic chlorinated ethenes is a global environmental and public health issue. Identifying microorganisms capable of cDCE degradation in these environments is of interest because of their potential application to bioremediation techniques. In this study, we sequenced, assembled, and analyzed the complete genome of Acinetobacter pittii CEP14, a strain isolated from chloroethene-contaminated groundwater, that has demonstrated the ability for aerobic cometabolic degradation of cDCE in the presence of n-hexane, phenol, and toluene. The A. pittii CEP14 genome consists of a 3.93 Mbp-long chromosome (GenBank accession no. CP084921) with a GC content of 38.9% and three plasmids (GenBank accession no. CP084922, CP084923, and CP084924). Gene function was assigned to 83.4% of the 3,930 coding DNA sequences. Functional annotation of the genome revealed that the CEP14 strain possessed all genetic elements to mediate the degradation of a range of aliphatic and aromatic compounds, including n-hexane and phenol. In addition, it harbors gene clusters involved in cytosol detoxification and oxidative stress resistance, which could play a role in the mitigation of toxic chemical intermediates that can arise during the degradation of cDCE. Gene clusters for heavy metal and antibiotic resistance were also identified in the genome of CEP14. These results suggest that CEP14 may be a versatile degrader of xenobiotic compounds and well-adapted to polluted environments, where a combination of heavy metal and organic compound pollution is often found.
- Klíčová slova
- Acinetobacter pittii, Chlorinated ethenes (CEs), Cometabolism, Oxygenase, Phenol, Whole-genome shotgun sequencing, cis-1,2-dichloroethene (cDCE) biodegradation,
- MeSH
- Acinetobacter MeSH
- biodegradace MeSH
- dichlorethyleny MeSH
- fenoly * MeSH
- genomika MeSH
- xenobiotika * MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- 1,2-dichloroethylene MeSH Prohlížeč
- dichlorethyleny MeSH
- fenoly * MeSH
- xenobiotika * MeSH
Shortly after the WHOs first notice a suspected case of omicron SARS-CoV-2 was reported in Liberec, Czech Republic. The primary goal of the following actions was to test the presence of the variant and stop the spread of the virus variant. On November 25 a sixty-year-old lady, who had recently returned from Namibia, visited a GP with flu-like symptoms and a rash on her chest. The antigen test was positive for SARS-CoV-2, a PCR test was planned. At that time, it was not known that a new variant of concern was spreading from Africa. On November 26 in the morning the GP announced a suspected omicron case to the Regional public health authority, who organized the following steps. A mobile sampling team was sent to the patient's home immediately, sample transported into the regional hospital and analyzed with the help of the national reference laboratory. The captured virus SARS-CoV-2 fitted the description of the omicron variant, was shared in the GISAID database and named hCoV-19/Czech Republic/KNL_2021-110119140/2021. Contact tracing was started immediately, eleven persons were tested and quarantined. One of them positive with no further spread. It is the first documented omicron case in the Czech Republic and one of the first cases in Europe, with an excellent systemic response to the alert. The laboratory was able to detect the omicron variant instantly after the request. This case also demonstrates how easily the virus spreads on long distances and how important it might be to increase the uptake of the booster vaccine.
- Klíčová slova
- COVID-19, SARS-CoV-2, case report, omicron,
- MeSH
- COVID-19 * MeSH
- lidé středního věku MeSH
- lidé MeSH
- SARS-CoV-2 * MeSH
- výzkum MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
- Geografické názvy
- Evropa MeSH
Sampling of microbial biomass is crucial for understanding and controlling remediation processes ongoing at contaminated sites in general, particularly when molecular genetic analyses are employed. In this study, fiber-based carriers with a nanofiber layer were developed and tested as a method to sample microbial biomass in groundwater for molecular genetic analysis. Nanofiber carriers, varying in the shape and the linear density of nanofibers, were examined throughout a 27-month monitoring period in groundwater contaminated with benzene, toluene, ethylbenzene and xylene isomers (BTEX), and chlorinated ethenes. The effect of carrier shape and nanofiber layer density on the microbial surface colonization and composition of the microbial biofilm was determined using real-time PCR and next-generation sequencing (NGS) analysis. Differences in microbial community composition between nanofiber carriers, groundwater, and soil samples were also analyzed to assess the applicability of carriers for biomass sampling at contaminated sites. The nanofiber carriers showed their applicability as a sampling tool, particularly because of their easy manipulation that facilitates DNA isolation. The majority of taxa (Proteobacteria, Firmicutes, and Bacteroidetes) present on the carrier surfaces were also detected in the groundwater. Moreover, the microbial community on all nanofiber carriers reflected the changes in the chemical composition of groundwater. Although the carrier characteristics (shape, nanofiber layer) did not substantially influence the microbial community on the carrier surface, the circular and planar carriers with a nanofiber layer displayed faster microbial surface colonization. However, the circular carrier was the most suitable for biomass sampling in groundwater because of its high contact area and because it does not require pre-treatment prior to DNA extraction.
- Klíčová slova
- Biomass carrier, Biomass sampling, Groundwater, Nanofibers, Next-generation sequencing, Real-time PCR,
- MeSH
- biodegradace MeSH
- biomasa MeSH
- chemické látky znečišťující vodu * analýza MeSH
- nanovlákna * MeSH
- podzemní voda * MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- chemické látky znečišťující vodu * MeSH
The benefits of plant-microbe interactions have been exploited extensively for nutrient removal. Radial oxygen loss in aquatic macrophytes potentially promotes nitrification and accelerates nitrogen removal through coupled nitrification-denitrification process. Nitrification is likely the limiting activity for an effective nitrogen removal in wetlands. In this work, we have quantified the effect of radial oxygen losses in Typha angustifolia plants in environments of contrasting salinities, including a temporary lagoon, a constructed wetland, and a river estuary. In all sites, radial oxygen diffusion occurred mainly at a narrow band, from 1 to 5 cm from the root tip, and were almost absent at the tip and basal sections of the root (> 5 cm). Root sections with active oxygen diffusion tended to show higher bacterial and archaeal densities in the rhizoplane according to 16S rRNA gene abundance data, except at higher salinities. Archaeal amoA /bacterial amoA gene ratios were highly variable among sites. Archaeal nitrifiers were only favoured over bacteria on the root surface of Typha collected from the constructed wetland. Collectively, radial oxygen loss had little effect on the nitrifying microbial community at the smaller scale (differences according to root-section), and observed differences were more likely related to prevailing physicochemical conditions of the studied environments or to long-term effects of the root microenvironment (root vs sediment comparisons).
- MeSH
- amoniak metabolismus MeSH
- kořeny rostlin metabolismus MeSH
- kyslík metabolismus MeSH
- mikrobiota MeSH
- nitrifikace fyziologie MeSH
- orobincovité metabolismus MeSH
- RNA ribozomální 16S genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- amoniak MeSH
- kyslík MeSH
- RNA ribozomální 16S MeSH
Biomolecular and hydrochemical tools were used to evaluate natural attenuation of chlorinated ethenes in a Quaternary alluvial aquifer located close to a historical source of large-scale tetrachloroethylene (PCE) contamination. Distinct stratification of redox zones was observed, despite the aquifer's small thickness (2.8 m). The uppermost zone of the target aquifer was characterised by oxygen- and nitrate-reducing conditions, with mixed iron- to sulphate-reducing conditions dominant in the lower zone, along with indications of methanogenesis. Natural attenuation of PCE was strongly influenced by redox heterogeneity, while higher levels of PCE degradation coincided with iron- to sulphate reducing conditions. Next generation sequencing of the middle and/or lower zones identified anaerobic bacteria (Firmicutes, Chloroflexi, Actinobacteria and Bacteroidetes) associated with reductive dechlorination. The relative abundance of dechlorinators (Dehalococcoides mccartyi, Dehalobacter sp.) identified by real-time PCR in soil from the lower levels supports the hypothesis that there is a significant potential for reductive dechlorination of PCE. Local conditions were insufficiently reducing for rapid complete dechlorination of PCE to harmless ethene. For reliable assessment of natural attenuation, or when designing monitoring or remedial systems, vertical stratification of key biological and hydrochemical markers should be analysed as standard, even in shallow aquifers.
- Klíčová slova
- Biological markers, Chlorinated solvents, Hydrochemical markers, Indigenous microorganisms, Natural attenuation,
- MeSH
- biodegradace MeSH
- chemické látky znečišťující vodu analýza MeSH
- Chloroflexi MeSH
- ethyleny analýza MeSH
- halogenace MeSH
- monitorování životního prostředí * MeSH
- podzemní voda chemie MeSH
- tetrachlorethylen chemie MeSH
- vysoce účinné nukleotidové sekvenování MeSH
- železo analýza MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- chemické látky znečišťující vodu MeSH
- ethylene MeSH Prohlížeč
- ethyleny MeSH
- tetrachlorethylen MeSH
- železo MeSH
Cis-1,2-dichloroethylene (cDCE), which is a common hazardous compound, often accumulates during incomplete reductive dechlorination of higher chlorinated ethenes (CEs) at contaminated sites. Simple monoaromatics, such as toluene and phenol, have been proven to induce biotransformation of cDCE in microbial communities incapable of cDCE degradation in the absence of other carbon sources. The goal of this microcosm-based laboratory study was to discover non-toxic natural monoaromatic secondary plant metabolites (SPMEs) that could enhance cDCE degradation in a similar manner to toluene and phenol. Eight SPMEs were selected on the basis of their monoaromatic molecular structure and widespread occurrence in nature. The suitability of the SPMEs chosen to support bacterial growth and to promote cDCE degradation was evaluated in aerobic microbial cultures enriched from cDCE-contaminated soil in the presence of each SPME tested and cDCE. Significant cDCE depletions were achieved in cultures enriched on acetophenone, phenethyl alcohol, p-hydroxybenzoic acid and trans-cinnamic acid. 16S rRNA gene sequence analysis of each microbial community revealed ubiquitous enrichment of bacteria affiliated with the genera Cupriavidus, Rhodococcus, Burkholderia, Acinetobacter and Pseudomonas. Our results provide further confirmation of the previously stated secondary compound hypothesis that plant metabolites released into the rhizosphere can trigger biodegradation of environmental pollutants, including cDCE.
- MeSH
- acetofenony metabolismus MeSH
- aerobióza MeSH
- Bacteria genetika metabolismus MeSH
- biodegradace MeSH
- cinnamáty metabolismus MeSH
- dichlorethyleny metabolismus MeSH
- fenethylalkohol metabolismus MeSH
- fenoly metabolismus MeSH
- fylogeneze MeSH
- hydroxybenzoáty metabolismus MeSH
- látky znečišťující půdu metabolismus MeSH
- mikrobiální společenstva genetika MeSH
- půdní mikrobiologie MeSH
- RNA ribozomální 16S MeSH
- rostliny metabolismus MeSH
- sekundární metabolismus MeSH
- toluen metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- 1,2-dichloroethylene MeSH Prohlížeč
- acetofenony MeSH
- acetophenone MeSH Prohlížeč
- cinnamáty MeSH
- cinnamic acid MeSH Prohlížeč
- dichlorethyleny MeSH
- fenethylalkohol MeSH
- fenoly MeSH
- hydroxybenzoáty MeSH
- látky znečišťující půdu MeSH
- phenolic acid MeSH Prohlížeč
- RNA ribozomální 16S MeSH
- toluen MeSH
