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

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

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

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

• MeSH
• genetika MeSH

• Konspekt
• Obecná genetika. Obecná cytogenetika. Evoluce
• NLK Obory
• genetika, lékařská genetika