Several methods of molecular analysis of microbial diversity, including terminal restriction fragment length polymorphism (T-RFLP) analysis are based on measurement of the DNA fragment length. Significant variation between sequence-determined and measured length of restriction fragments (drift) has been observed, which can affect the efficiency of the identification of microorganisms in the analyzed communities. In the past, this variation has been attributed to varying fragment length and purine content. In this study, principal component analysis and multiple regression analysis were applied to find the contributions of those and several other fragment characteristics. We conclude that secondary structure melting point and G+C nucleotide content, besides the fragment length, contribute to the variation observed, whereas the contribution of purine content is less important. Incomplete denaturation of the sample at the start of electrophoretic separation of fragments has been excluded as a major cause of the variation observed. Our regression model explains the observed drift variation by approximately 56%, with standard deviation of the prediction equal to approximately 1.3 bp.

• MeSH
• analýza polymorfismu délky amplifikovaných restrikčních fragmentů metody   normy   MeSH
• DNA fungální chemie   genetika   MeSH
• houby chemie   genetika   izolace a purifikace   MeSH
• konformace nukleové kyseliny MeSH
• Phalaris mikrobiologie   MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• tranzitní teplota MeSH
• zastoupení bazí MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• práce podpořená grantem MeSH

• MeSH
• Archaea genetika   metabolismus   MeSH
• Bacteria genetika   metabolismus   MeSH
• chlor metabolismus   MeSH
• houby genetika   metabolismus   MeSH
• mikrobiální společenstva fyziologie   MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• půda analýza   MeSH
• půdní mikrobiologie MeSH
• RNA ribozomální 16S analýza   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: Telomeres, nucleoprotein structures comprising short tandem repeats and delimiting the ends of linear eukaryotic chromosomes, play an important role in the maintenance of genome stability. Therefore, the determination of the length of telomeres is of high importance for many studies. Over the last years, new methods for the analysis of the length of telomeres have been developed, including those based on PCR or analysis of NGS data. Despite that, terminal restriction fragment (TRF) method remains the gold standard to this day. However, this method lacks universally accepted and precise tool capable to analyse and statistically evaluate TRF results. RESULTS: To standardize the processing of TRF results, we have developed WALTER, an online toolset allowing rapid, reproducible, and user-friendly analysis including statistical evaluation of the data. Given its web-based nature, it provides an easily accessible way to analyse TRF data without any need to install additional software. CONCLUSIONS: WALTER represents a major upgrade from currently available tools for the image processing of TRF scans. This toolset enables a rapid, highly reproducible, and user-friendly evaluation of almost any TRF scan including in-house statistical evaluation of the data. WALTER platform together with user manual describing the evaluation of TRF scans in detail and presenting tips and troubleshooting, as well as test data to demo the software are available at https://www.ceitec.eu/chromatin-molecular-complexes-jiri-fajkus/rg51/tab?tabId=125#WALTER and the source code at https://github.com/mlyc93/WALTER .

• MeSH
• software * MeSH
• telomery * genetika   MeSH
• Publikační typ
• časopisecké články MeSH

Spatial distribution of ectomycorrhizae-associated basidiomycetes was determined in oakbirch forest using terminal restriction fragment length polymorphism (T-RFLP) analysis. The data were correlated with actual soil humidity, pH, electric conductivity of the soil extract, absorbance A(465) and A(665) of water and alkali soil extracts and with the ratio A(465)/A(665) (parameter A4/A6). Natural non-homogeneity of the soil parameters was used as experimental gradient. Distance-based redundancy analysis of the T-RFLP data (with soil parameters being taken as environmental parameters) provided significant results when ITS1F-terminanted restriction fragments were analyzed. Among other fungi, a Mycena galericulata related fungus was observed to correlate negatively with A4/A6, indicating its association with highly humified soil organic matter. Positive association of other, unidentified fungi with A4/A6 was also observed. Several other unidentified fungi negatively correlated with electric conductivity of the soil extract. The results may explain nonhomogeneity of the spatial distribution of the fungi associated with ectomycorrhizae as a result of their interaction with non-homogeneous soil environment.

• MeSH
• Agaricales genetika   izolace a purifikace   růst a vývoj   MeSH
• Basidiomycota genetika   izolace a purifikace   klasifikace   růst a vývoj   MeSH
• bříza mikrobiologie   růst a vývoj   MeSH
• dub (rod) mikrobiologie   růst a vývoj   MeSH
• elektrolyty analýza   MeSH
• financování organizované MeSH
• kořeny rostlin mikrobiologie   MeSH
• mykorhiza MeSH
• organické látky analýza   MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• půda MeSH
• půdní mikrobiologie MeSH
• stromy mikrobiologie   růst a vývoj   MeSH
• Geografické názvy
• Česká republika MeSH

Common scab of potatoes is a disease, which is difficult to manage due to complex interactions of the pathogenic bacteria (Streptomyces spp.) with soil, microbial community and potato plants. In Bohemian-Moravian Highlands in the Czech Republic two sites (Vyklantice and Zdirec) were selected for a study of common scab disease suppressivity. At both sites, a field with low disease severity occurs next to one with high severity and the situation was regularly observed over four decades although all four fields undergo a crop rotation. In the four fields, quantities of bacteria, actinobacteria and the gene txtB from the biosynthetic gene cluster of thaxtomin, the main pathogenicity factor of common scab, were analyzed by real-time PCR. Microbial community structure was compared by terminal fragment length polymorphism analysis. Soil and potato periderm were characterized by contents of carbon, nitrogen, phosphorus, sulphur, calcium, magnesium, and iron. Quality of organic matter was assessed by high performance liquid chromatography of soil extracts. The study demonstrated that the suppressive character of the fields is locally specific. At Zdirec, the suppressivity was associated with low txtB gene copies in bulk soil, while at Vyklantice site it was associated with low txtB gene copies in the tuberosphere. The differences were discussed with respect to the effect of abiotic conditions at Zdirec and interaction between potato plant and soil microbial community at Vyklantice. Soil pH, Ca soil content or cation concentrations, although different were not in the range to predict the disease severity. Low severity of common scab was associated with low content of soil C, N, C/N, Ca and Fe suggesting that oligotrophic conditions may be favorable to common scab suppression.

• MeSH
• bakteriální geny * MeSH
• hlízy rostlin mikrobiologie   MeSH
• indoly metabolismus   MeSH
• multigenová rodina * MeSH
• nemoci rostlin mikrobiologie   MeSH
• piperaziny metabolismus   MeSH
• polymorfismus délky restrikčních fragmentů * MeSH
• půdní mikrobiologie * MeSH
• Solanum tuberosum mikrobiologie   MeSH
• Streptomyces genetika   metabolismus   patogenita   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Internal bacterial communities of synanthropic mites Acarus siro, Dermatophagoides farinae, Lepidoglyphus destructor, and Tyrophagus putrescentiae (Acari: Astigmata) were analyzed by culturing and culture-independent approaches from specimens obtained from laboratory colonies. Homogenates of surface-sterilized mites were used for cultivation on non-selective agar and DNA extraction. Isolated bacteria were identified by sequencing of the 16S rRNA gene. PCR amplified 16S rRNA genes were analyzed by terminal restriction fragment length polymorphism analysis (T-RFLP) and cloning sequencing. Fluorescence in situ hybridization using universal bacterial probes was used for direct bacterial localization. T-RFLP analysis of 16S rRNA gene revealed distinct species-specific bacterial communities. The results were further confirmed by cloning and sequencing (284 clones). L. destructor and D. farinae showed more diverse communities then A. siro and T. putrescentiae. In the cultivated part of the community, the mean CFUs from four mite species ranged from 5.2 × 10(2) to 1.4 × 10(3) per mite. D. farinae had significantly higher CFUs than the other species. Bacteria were located in the digestive and reproductive tract, parenchymatical tissue, and in bacteriocytes. Among the clones, Bartonella-like bacteria occurring in A. siro and T. putresecentiae represented a distinct group related to Bartonellaceae and to Bartonella-like symbionts of ants. The clones of high similarity to Xenorhabdus cabanillasii were found in L. destructor and D. farinae, and one clone related to Photorhabdus temperata in A. siro. Members of Sphingobacteriales cloned from D. farinae and A. siro clustered with the sequences of "Candidatus Cardinium hertigii" and as a separate novel cluster.

• MeSH
• Acari klasifikace   genetika   mikrobiologie   MeSH
• Bacteria klasifikace   genetika   izolace a purifikace   MeSH
• Dermatophagoides farinae mikrobiologie   MeSH
• druhová specificita MeSH
• hybridizace in situ fluorescenční MeSH
• klonování DNA MeSH
• molekulární sekvence - údaje MeSH
• polymerázová řetězová reakce MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• RNA ribozomální 16S genetika   MeSH
• roztoči mikrobiologie   MeSH
• sekvenční analýza DNA MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Genotypizace CMV u skupinových výskytů infekce u příjemců transplantátu krvetvorných kmenových buněk, léčených v UHKT v letech 2004-5, a následná epidemiologická analýza těchto případů, b) vypracování metod pro detekci mutací, spojených s rezistencí CMVna antivirotika, a jejich využití pro charakterizaci kmenů, získaných od pacientů s nedostatečnou odpovědí na antivirovou terapii. Genotypizace bude prováděna na základě analýzy virových glykoproteinů gB a gN pomocí polymorfismu fragmentů restrikčního štěpení genu UL55 (gB) a sekvenace N- koncové části genu UL73 (gN). Pro účely detekce rezistentních mutant bude využita sekvenace funkčně důležitých lokusů genů UL97 a UL54. Porovnání výsledků genetické analýzy s klinickými údaji o léčbě a průběhu infekce.; A) Genotyping of CMV strains identified in clustered cases of CMV infection in hematopoietic stem cell transplant recipients treated in Inst.Hematol. Blood Transfussion during the period 2004-5 using methods of RFLP analysis of gB gene UL55 and sequenation of N- terminal fragment of gN gene UL73.Subsequent epidemiological analysis of the infections. B) Setting of the methods for detection of antivirals resistence-associated CMV mutants by genetical analysis of UL97 and UL54 and their use for characterisation of CMV strains obtained from patients with poor response to antiviral therapy.

• MeSH
• cytomegalovirové infekce diagnóza   MeSH
• Cytomegalovirus genetika   MeSH
• genotyp MeSH
• molekulární epidemiologie MeSH
• mutace MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• sekvenční analýza MeSH
• virová léková rezistence MeSH

• Konspekt
• Biochemie. Molekulární biologie. Biofyzika
• NLK Obory
• biologie
• infekční lékařství
• transplantologie
• NLK Publikační typ
• závěrečné zprávy o řešení grantu IGA MZ ČR

Plant and microbial community composition in connection with soil chemistry determines soil nutrient cycling. The study aimed at demonstrating links between plant and microbial communities and soil chemistry occurring among and within four sites: two pine forests with contrasting soil pH and two grasslands of dissimilar soil chemistry and vegetation. Soil was characterized by C and N content, particle size, and profiles of low-molecular-weight compounds determined by high-performance liquid chromatography (HPLC) of soil extracts. Bacterial and actinobacterial community composition was assessed by terminal restriction fragment length polymorphism (T-RFLP) and cloning followed by sequencing. Abundances of bacteria, fungi, and actinobacteria were determined by quantitative PCR. In addition, a pool of secondary metabolites was estimated by erm resistance genes coding for rRNA methyltransferases. The sites were characterized by a stable proportion of C/N within each site, while on a larger scale, the grasslands had a significantly lower C/N ratio than the forests. A Spearman's test showed that soil pH was correlated with bacterial community composition not only among sites but also within each site. Bacterial, actinobacterial, and fungal abundances were related to carbon sources while T-RFLP-assessed microbial community composition was correlated with the chemical environment represented by HPLC profiles. Actinobacteria community composition was the only studied microbial characteristic correlated to all measured factors. It was concluded that the microbial communities of our sites were influenced primarily not only by soil abiotic characteristics but also by dominant litter quality, particularly, by percentage of recalcitrant compounds.

• MeSH
• Bacteria klasifikace   genetika   izolace a purifikace   MeSH
• bakteriální nálož MeSH
• biodiverzita MeSH
• DNA bakterií chemie   genetika   MeSH
• dusík analýza   MeSH
• fylogeneze MeSH
• houby klasifikace   genetika   izolace a purifikace   MeSH
• koncentrace vodíkových iontů MeSH
• methyltransferasy genetika   MeSH
• molekulární sekvence - údaje MeSH
• organické látky analýza   MeSH
• počet mikrobiálních kolonií MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• půda chemie   MeSH
• půdní mikrobiologie MeSH
• rostliny mikrobiologie   MeSH
• sekvenční analýza DNA MeSH
• shluková analýza MeSH
• uhlík analýza   MeSH
• vysokoúčinná kapalinová chromatografie MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Exfoliative toxin A (ETA)-coding temperate bacteriophages are leading contributors to the toxic phenotype of impetigo strains of Staphylococcus aureus. Two distinct eta gene-positive bacteriophages isolated from S. aureus strains which recently caused massive outbreaks of pemphigus neonatorum in Czech maternity hospitals were characterized. The phages, designated φB166 and φB236, were able to transfer the eta gene into a prophageless S. aureus strain which afterwards converted into an ETA producer. Complete phage genome sequences were determined, and a comparative analysis of five designed genomic regions revealed major variances between them. They differed in the genome size, number of open reading frames, genome architecture, and virion protein patterns. Their high mutual sequence similarity was detected only in the terminal regions of the genome. When compared with the so far described eta phage genomes, noticeable differences were found. Thus, both phages represent two new lineages of as yet not characterized bacteriophages of the Siphoviridae family having impact on pathogenicity of impetigo strains of S. aureus.

• MeSH
• DNA virů chemie   genetika   MeSH
• DNA viry genetika   izolace a purifikace   MeSH
• epidemický výskyt choroby MeSH
• exfoliatiny genetika   MeSH
• fylogeneze MeSH
• genom virový * MeSH
• impetigo epidemiologie   mikrobiologie   MeSH
• infekce spojené se zdravotní péčí epidemiologie   MeSH
• lidé MeSH
• molekulární sekvence - údaje MeSH
• novorozenec MeSH
• otevřené čtecí rámce MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• pořadí genů MeSH
• porodnice MeSH
• přenos genů horizontální MeSH
• profágy klasifikace   genetika   izolace a purifikace   MeSH
• sekvenční analýza DNA MeSH
• sekvenční homologie MeSH
• shluková analýza MeSH
• stafylokokové bakteriofágy klasifikace   genetika   izolace a purifikace   MeSH
• stafylokokové infekce epidemiologie   mikrobiologie   MeSH
• Staphylococcus aureus izolace a purifikace   virologie   MeSH
• syntenie MeSH
• transdukce genetická MeSH
• Check Tag
• lidé MeSH
• novorozenec MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH

Biologická terapie představuje v gastroenterologii významnou součást standardní terapie idiopatických střevních zánětů, kolorektálního karcinomu a uplatňuje se také v léčbě po transplantaci jater. U idiopatických střevních zánětů v současné době lze použít v léčbě Crohnovy nemoci infliximab či adalimumab, při onemocnění ulcerózní kolitidou pouze infliximab. Terapie je soustředěna do center biologické léčby, která garantují správnost indikace a realizaci léčby. Současné indikace umožňují použití biologické terapie u idiopatických střevních zánětů po vyčerpání možností konvenční léčby kortikosteroidy a/nebo imunosupresivy. V terapii kolorektálního karcinomu se ukazují výhodné kombinace bevacizumabu a cetuximabu s konvenční chemoterapií, jejíž účinek potencuje. Použití biologické terapie v hepatologii je již na hranicích transplantologické problematiky, do jejíž kompetence patří také její indikace.

In gastroenterology, biological therapy represents a significant component in the standard treatment for both idiopathic inflammatory bowel disease and colorectal cancer and is also employed following liver transplantation. Currently, the management of idiopathic inflammatory bowel diseases involves the use of infliximab or adalimumab to treat Crohn‘ s disease and the use of infliximab alone to treat ulcerative colitis. Therapy is restricted to biological therapy centres which guarantee correct indication and implementation of treatment. Current indications make possible the use of biological therapy in idiopathic inflammatory bowel diseases once conventional treatment options using corticosteroids and/or immunosuppressives have failed. The combination of bevacizumab and cetuximab with conventional chemotherapy the effect of which is thus potentiated has been shown beneficial in the treatment of colorectal cancer. The use of biological therapy in hepatology is rather an issue for transplantology which is also responsible for its indications.

• MeSH
• biologická terapie metody   škodlivé účinky   MeSH
• Crohnova nemoc farmakoterapie   MeSH
• idiopatické střevní záněty farmakoterapie   MeSH
• imunoglobuliny - Fab fragmenty škodlivé účinky   terapeutické užití   MeSH
• kolorektální nádory farmakoterapie   MeSH
• lidé MeSH
• monoklonální protilátky terapeutické užití   MeSH
• nádory chemicky indukované   MeSH
• neutropenie chemicky indukované   MeSH
• receptory TNF - typ I škodlivé účinky   terapeutické užití   MeSH
• rekombinantní proteiny škodlivé účinky   terapeutické užití   MeSH
• TNF-alfa antagonisté a inhibitory   MeSH
• tvorba protilátek účinky léků   MeSH
• ulcerózní kolitida farmakoterapie   MeSH
• Check Tag
• lidé MeSH