• MeSH
• prasata genetika   MeSH
• skot MeSH
• transportní proteiny genetika   MeSH
• zvířata MeSH
• Check Tag
• skot MeSH
• zvířata MeSH

AIMS: Brewing yeasts are classified into two species-Saccharomyces pastorianus and Saccharomyces cerevisiae. Most of the brewing yeast strains are natural interspecies hybrids typically polyploids and their identification is thus often difficult giving heterogenous results according to the method used. We performed genetic characterization of a set of the brewing yeast strains coming from several yeast culture collections by combination of various DNA-based techniques. The aim of this study was to select a method for species-specific identification of yeast and discrimination of yeast strains according to their technological classification. METHODS AND RESULTS: A group of 40 yeast strains were characterized using PCR-RFLP analysis of ITS-5·8S, NTS, HIS4 and COX2 genes, multiplex PCR, RAPD-PCR of genomic DNA, mtDNA-RFLP and electrophoretic karyotyping. Reliable differentiation of yeast to the species level was achieved by PCR-RFLP of HIS4 gene. Numerical analysis of the obtained RAPD-fingerprints and karyotype revealed species-specific clustering corresponding with the technological classification of the strains. Taxonomic position and partial hybrid nature of strains were verified by multiplex PCR. Differentiation among species using the PCR-RFLP of ITS-5·8S and NTS region was shown to be unreliable. Karyotyping and RFLP of mitochondrial DNA evinced small inaccuracies in strain categorization. CONCLUSIONS: PCR-RFLP of HIS4 gene and RAPD-PCR of genomic DNA are reliable and suitable methods for fast identification of yeast strains. RAPD-PCR with primer 21 is a fast and reliable method applicable for differentiation of brewing yeasts with only 35% similarity of fingerprint profile between the two main technological groups (ale and lager) of brewing strains. SIGNIFICANCE AND IMPACT OF THE STUDY: It was proved that PCR-RFLP method of HIS4 gene enables precise discrimination among three technologically important Saccharomyces species. Differentiation of brewing yeast to the strain level can be achieved using the RAPD-PCR technique.

• MeSH
• DNA fungální genetika   MeSH
• druhová specificita MeSH
• mitochondriální DNA genetika   MeSH
• pivo analýza   mikrobiologie   MeSH
• polymerázová řetězová reakce metody   MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• Saccharomyces genetika   izolace a purifikace   metabolismus   MeSH
• technika náhodné amplifikace polymorfní DNA metody   MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH

Several biochemical and molecular methods were used for discrimination of four Lactobacillus reuteri strains isolated from goatling and lamb stomach mucosa. Internal transcribed spacer (ITS)-PCR method and protein analysis by SDS-PAGE and MALDI-TOF showed to be suitable for strain discrimination whereas ITS-PCR/RFLP and enterobacterial repetitive intergenic consensus (ERIC)-PCR were not strain specific. The used methods differentiated tested strains into distinct groups; however, the location of strains in groups varied. Consistency in results was observed in the case of L. reuteri E and L. reuteri KO4m that were clustered into the same groups using all techniques, except of MALDI-TOF MS. The last one grouped goatling strains and lamb isolate into separate clusters. All investigated methods, except of ITS-PCR/RFLP and ERIC-PCR, were assessed as appropriate for distinguishing of L. reuteri strains.

• MeSH
• bakteriální proteiny analýza   MeSH
• bakteriologické techniky metody   MeSH
• DNA bakterií chemie   genetika   MeSH
• elektroforéza v polyakrylamidovém gelu MeSH
• kozy MeSH
• Limosilactobacillus reuteri klasifikace   genetika   izolace a purifikace   fyziologie   MeSH
• mezerníky ribozomální DNA chemie   genetika   MeSH
• ovce MeSH
• polymerázová řetězová reakce MeSH
• reprodukovatelnost výsledků MeSH
• shluková analýza MeSH
• spektrometrie hmotnostní - ionizace laserem za účasti matrice MeSH
• žaludeční sliznice mikrobiologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• srovnávací studie MeSH

Cutaneous leishmaniasis (CL) is an important public health problem in Turkey. CL has been most frequently seen in Sanliurfa. There is an expectation of increase in the population of leishmaniasis cases with the influence of Syrian refugees arriving in Turkey. In this study we aimed to diagnosis of CL and identifying of parasite from Leishmania isolates by using ITS 1 PCR RFLP. Samples were collected from 135 CL patients in Sanliurfa. After the specimens were inoculated in medium NNN, the ones which were cultures positive were cultivated in RPMI 1640 followed by PCR-RFLP. Genomic DNA was extracted phenol-chloroform procedure. Samples were examined by using ITS 1 PCR followed by RFLP analysis. Our results indicated that two species, L. tropica (132 samples) and L. major (3 samples), are responsible for cutaneous leishmaniasis in Sanlıurfa. Our study is the first scientific study in which it is reported molecular analyses of cutaneous leishmaniasis cases caused by L. major in Sanliurfa in Southestern Anatolia Region. Because CL cases caused by L.major are detected in our study, it is considered that genotyping is important for diagnosis of Leishmania and following change of epidemiology.

• MeSH
• dítě MeSH
• dospělí MeSH
• kojenec MeSH
• kůže mikrobiologie   patologie   MeSH
• Leishmania klasifikace   genetika   MeSH
• leishmanióza kožní diagnóza   epidemiologie   mikrobiologie   patologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mezerníky ribozomální DNA genetika   MeSH
• mladiství MeSH
• mladý dospělý MeSH
• polymerázová řetězová reakce MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• předškolní dítě MeSH
• protozoální DNA genetika   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
• Geografické názvy
• Turecko epidemiologie   MeSH

Zhodnocení genetické variability genů cytokinové sítě vzhledem k revmatoidní artritidě a jejímu intermediárnímu fenotypu, včetně analýzy plazmatických hladin vybraných cytokinů. Metod PCR a RFLP bude využito ke stanovení promotorových a intronových polymorfismů. Nové polymorfismy budou v regulačních oblastech genů pro IL-8, IL-13 a IL-15 detekovány pomocí heteroduplexní analýzy a SSCP. Proteinové hladiny vybraných cytokinů budou hodnoceny pomocí enzymaticky zesílené chemiluminiscence a ELISA metody. K vyjádření vtahu mezi genotypy a onemocněním bude využita tzv. "case-control" studie (asociační studie - "případ-kontroly"). Významnost rozdílů bude hodnocena pomocí chi-square testu a Fisher exact testu. Odds ratio (OR) s 95% konfidenčním intervalem budevyužito ke stanovení relativního rizika ve vztahu k RA.; Assesment of genetic variability of genes in cytokine net in respect to rheumatoid arthritis and its intermediate phenotype, inclusive of the analysis of selected cytokines plasma levels. For detection of pholymorphisms in promoter and introne regions, PCR and RFLP methods will be used. New polymorphisms in regulatory regions of IL-8, IL-13 and IL-15 genes will be detected by heteroduplex analysis and SSCP. The protein levels of cytokines will be measured by enzyme-amplified chemiluminiscence and ELISAmethod. The "case-control" (association) study will be used to assess the relation between genotype and disease. The significance of the differences will be calculated by chi-square test and Fisher's exact test. Odds ratio (OR) with 95% confidence interval will be used to estimate the relative risks.

• MeSH
• cytokiny analýza   genetika   MeSH
• ELISA metody   využití   MeSH
• interleukin-13 MeSH
• interleukin-15 MeSH
• interleukin-8 MeSH
• luminiscenční měření metody   využití   MeSH
• polymorfismus genetický MeSH
• revmatoidní artritida genetika   MeSH

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

This is the first exhaustive report on the fungal community biodiversity in hypersaline water in România. A total of 27 fungal strains (19 molds and eight yeast) have been isolated from Lopătari hypersaline water, Buzau County. Based on classical investigation, these strains have been identified as belonging to the genera Aureobasidium, Alternaria, Aspergillus, Penicillium, and Fusarium. The molecular characterization of fungal isolates at species level was performed using PCR-RFLP analysis of the 5.8S-ITS region. PCR products were digested with different combinations of endonucleases. The most frequently isolated species were Aspergillus niger (14.81% of all isolates), A. versicolor, (14.81%) and Penicillium crustosum (14.81%). In addition, ribosomal restriction patterns which exhibited profiles specific to Aureobasidium pullulans were derived, and to discriminate between Aureobasidium isolates, the elongase-encoding gene (ELO) was chosen as a genetic marker followed by digestion with endonuclease HhaI. Five yeast isolates displayed restriction patterns corresponding to Aureobasidium melanogenum (18.52%) and three isolates to Aureobasidium pullulans (11.11%). In addition, the RFLP types of Aureobasidium pullulans varieties with HhaI are clearly distinguished and could be applied to assess the intraspecific variability.

• MeSH
• Ascomycota genetika   izolace a purifikace   metabolismus   MeSH
• Aspergillus genetika   izolace a purifikace   metabolismus   MeSH
• biodiverzita MeSH
• DNA fungální genetika   MeSH
• fylogeneze MeSH
• houby klasifikace   genetika   izolace a purifikace   MeSH
• kvasinky genetika   izolace a purifikace   metabolismus   MeSH
• mikrobiologie vody * MeSH
• polymerázová řetězová reakce MeSH
• polymorfismus délky restrikčních fragmentů * MeSH
• řeky chemie   mikrobiologie   MeSH
• ribozomální DNA genetika   MeSH
• sekvenční analýza DNA MeSH
• slané vody * MeSH
• tolerance k soli * MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Rumunsko MeSH

Diagnostika legionelóz, hlavne závažnejšej život ohrozujúcej formy, tzv. Legionárskej choroby je zložitá, najmä pre netypické príznaky infekcie, nie vždy dominujúcu atypickú pneumóniu a často veľmi dramatický septický priebeh ochorenia so zlyhávaním orgánov. Stanovenie diagnózy v akútnej fáze ochorenia je možné detekciou legionelového antigénu v moči a dôkazom DNA legionel v PCR/real-time PCR v sére, vzorkách z dolných dýchacích ciest a v moči. Kultivácia na špecifických pôdach zostáva zlatým štandardom, ale pre svoju náročnosť sa využíva málo. Sérologické vyšetrenie si vyžaduje párové vzorky, a tak je prínosom pre diagnostiku v neskoršej fáze infekcie, pričom asi 20 % pacientov vôbec netvorí protilátky. Veľký pokrok sa zaznamenal v typizačných metódach (RFLP, PFGE, metódy založené na PCR, sekvenačné metódy) a rýchlych identifikačných metódach (MALDI-TOF).

The diagnosis of legionellosis, especially of its severe, life-threatening form, Legionnaires' disease, is complicated, primarily because of non-typical symptoms of the infection, not always dominating atypical pneumonia, and often a very dramatic septic course of the disease with multiorgan failures. The diagnosis of the acute phase of the disease can be established by the detection of Legionella antigen in urine and by PCR/real-time PCR detection of Legionella DNA in serum and lower respiratory tract and urine samples. Cultivation on specific media remains the gold standard, but this very demanding method is rarely used. Serological testing requires paired samples and thus is relevant to the diagnosis at a later stage of infection, although it is to be noted that about 20% of patients do not produce the antibodies. Great progress has been made in typing methods (RFLP, PFGE, or PCR based and sequence based methods) and rapid identification methods (MALDI-TOF).

• MeSH
• analýza moči MeSH
• antibakteriální látky aplikace a dávkování   terapeutické užití   MeSH
• časná diagnóza MeSH
• diferenciální diagnóza MeSH
• incidence MeSH
• klinické laboratorní techniky metody   MeSH
• legionářská nemoc * diagnóza   farmakoterapie   MeSH
• Legionella * izolace a purifikace   klasifikace   patogenita   MeSH
• lidé MeSH
• nukleové kyseliny analýza   MeSH
• sérologické testy MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• souhrny MeSH

OBJECTIVE: The motility and genotype of the flagellin fliC and fliD genes were investigated in 82 Clostridioides difficile isolates belonging to the ribotypes (RTs): 027 (n = 41), 176 (n = 17), 023 (n = 8), 017 (n = 6) and 046 (n = 10). The reference C. difficile strains 630 and M120 were included as controls for the motility assay. METHODS: A Multiple Locus Variable-number Tandem Repeat Analysis (MLVA) was used to exclude the genetic relatedness of C. difficile isolates belonging to the same RT. The variability of the fliC and fliD genes was determined by PCR-restriction fragment length polymorphism (RFLP) analysis and Sanger sequencing. The motility assay was carried out with 0.175% BHI agar tubes and BHI solid media plates with 0.4% agar. RESULTS: The highest motility was observed in C. difficile RT023 isolates (p < 0.01), followed by RTs 027 and 176. C. difficile isolates of RTs 017 and 046 were less motile than RTs 027, 176 and 023 (p < 0.01). The fliC and fliD genes were present in all clinical isolates irrespective of the motility results. In the fliC gene analysis, four different RFLP groups were identified (I, II, VII, X). The fliC group VII was identified in two RTs (027 and 176), whereas the remaining three groups (I, II and X) belonged to a single RT 046, 017 and 023, respectively. The fliD gene analysis identified four new RFLP groups (a, b, c and d). CONCLUSIONS: C. difficile RT023 is highly motile and its motility is comparable to the hypervirulent RT027 and its genetic relative RT176.

• MeSH
• bakteriální proteiny genetika   MeSH
• Clostridioides difficile * genetika   MeSH
• Clostridioides MeSH
• flagelin genetika   MeSH
• genotyp MeSH
• klostridiové infekce * MeSH
• lidé MeSH
• ribotypizace MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Zhodnocení genetické variability genů pro MMP, TIMP a ACE vzhledem k akutnímu infarktu myokardu a jeho intermediálnímu fenotypu, včetně analýzy sérovýchh hladin vybraných působků. Metod PCR a RFLP bude využito ke stanovení promotorových a intronových polymorfismů. Nové polymorfismy budou v regulačních oblastech genů detekovány pomocí heteroduplexní analýzy a SSCP. Proteinové hladiny vybraných působků budou hodnoceny pomocí ELISA metody. K vyjádření vtahu mezi genotypy a onemocněním bude využita tzv. "case-control" studie (asociační studie - "případ-kontroly"). Data budou sumarizována jednak běžnými statickým metodami (chi-kvadrát test, Fisher exakt test, odds ratio, ..) zaměřenými na hlavní cílové parametry projektu a dále je plánována vícerozměrná analýza rizikových faktorů.; Assesment of genetic variability of MMP, TIMP and ACE genes in respect to acute infarct myocardum and its intermediate phenotype, inclusive of the analysis of selected serum levels. For detection of pholymorphisms in promoter and intone regions, PCR andRFLP methods will be used. New polymorphisms in regulatory regions of MMP and TIMP genes will be detected by heteroduplex analysis and SSCP. The serum levels will be measured by ELISA method. The "case-control" (association) study will be used to assessthe relation between genotype and disease. The data will be summarised by common statistic methods (chi-square test, Fisher's exact test and odds ratio) and further the analysis of risk factors is planed.

• MeSH
• beta blokátory terapeutické užití   MeSH
• infarkt myokardu MeSH
• inhibitory ACE terapeutické užití   MeSH
• inhibitory matrixových metaloproteinas MeSH
• matrixová metaloproteinasa 12 MeSH
• polymerázová řetězová reakce MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• polymorfismus genetický MeSH
• polymorfismus konformace jednovláknové DNA MeSH
• prognóza MeSH
• remodelace komor MeSH
• tkáňové inhibitory metaloproteinas MeSH

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

Smithov-Lemliho-Opitzov syndróm (SLOS) je metabolicko-malformačný syndróm s autozómovo recesívnou dedičnosťou, spôsobený deficitom 7-dehydrocholesterolreduktázy, posledného enzýmu v biosyntéze cholesterolu. Najčastejšie sa vyskytuje v strednej Európe (1 : 15-20 000). Na diagnostiku SLOS nestačia klinické príznaky, pretože ochorenie má veľkú fenotypovú variabilitu. Objav poruchy biosyntézy cholesterolu umožnil v súčasnosti presnú biochemickú diagnostiku, ktorá je založená na detekcii vysokých hodnôt prekurzorov cholesterolu 7-dehydrocholesterolu, jeho izoméru 8-dehydrocholesterolu a nízkych hodnôt celkového cholesterolu pomocou plynovej chromatografie a hmotnostnej spektrometrie. Na skríningové vyšetrenie akumulovaného 7-dehydrocholesterolu je vhodná jednoduchá a lacná ultrafialová spektrofotometria lipidov séra. V molekulovej diagnostike SLOS sa využívajú dva prístupy, metóda PCR/RFLP a sekvenovanie celej kódujúcej oblasti DHCR7 génu.

Smith-Lemli-Opitz syndrome (SLOS) is autosomal recessive metabolic-malformation syndrome, caused by a deficiency of 7-dehydrocholesterol reductase affecting the last step of cholesterol biosynthesis. SLOS appears to be most frequent in central European populations (1 : 15-20,000). The clinical diagnosis of SLOS cannot always be made with certainty because of the remarkable variability of clinical phenotype. The discovery of primary defect of cholesterol biosynthesis had important implication for exact diagnosis of patients with SLOS. Biochemical diagnosis of SLOS is now confirmed by gas chromatography/mass spectrometry by finding low serum cholesterol and its abnormally high precursors 7- and 8-dehydro-cholesterol levels. The ultraviolet spectrophotometry determination of serum sterols with 7-dehydrocholesterol accumulation is a reliable and cheap test for screening SLOS. Two techniques, polymerase chain reaction together with restriction analysis (PCR/RFLP) and sequencing are used to detect mutations in the DHCR7 gene.