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Pyrosequencing analysis of fungal assemblages and the effect of ITS rDNA intragenomic variation on diversity estimates
Zelenka, Tomáš ; Kolařík, Miroslav (advisor) ; Větrovský, Tomáš (referee)
The ITS region of nuclear ribosomal DNA has recently become a frequently studied region for its use as a barcode marker. It is employed in environmental metagenomic analyses, the determination of fungi and in studies of fungal phylogenetics. In genomes, rDNA occurs in the form of large multicopy tandem arrays. Thus common Sanger sequencing leads to a consensus of many copies which in fact conceals most of the potential intragenomic variation. However 454 pyrosequencing reveals the sequence of single copies. Although it is believed that most of the variation among copies is reduced during a process called concerted evolution, some variation might still be conserved, including variation of non-functional pseudogenes. As a consequence, when using 454 pyrosequencing, we are not able to discriminate between this variation and real diversity. This could have a huge impact on the estimation of real diversity as well as on the correct assessment of pyrosequence studies. This thesis reviews current knowledge of intragenomic variation among fungi and summarizes some papers applying pyrosequencing in the research of fungal diversity. At the same time it indicates intragenomic variation as a potential cause of untrue diversity in diversity studies. Up until now there hasn't existed any experimental survey covering...
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Ericaceae root mycobionts: diversity, distribution and interactions with neighboring plants
Kolátková, Viktorie ; Vohník, Martin (advisor) ; Baláž, Milan (referee)
The roots of ericaceous plants provide an environment to many fungal endophytes and mycorrhizal fungi.We can observe at least four mycorrhizal types in the family, of which the ericoid mycorrhiza (ErM) is spread the most. The Ericaceae are able to survive in many different and often harsh habitats all over the world thanks to this symbiosis (from the tropical cloud forests, to the arctic tundra heathlands). The group of fungi, forming this relationship, used to be seen as a few very well circumscribed ascomycetes, namely Rhizoscyphus ericae (Read) Zhang & Zhuang and Oidiodendron maius Barron species. With the development of molecular methods, it was expanded of many other species, some of which belonging to Basidiomycota, particularly the order Sebacinales. The diversity of ErM fungal community varies at different geographic locations and it can be influenced by factors such as altitude or the surrounding ecosystem. However, due to lack of further information, we still know quite little about the species distribution. Although they are primarily symbionts of the Ericaceae, some ErM fungi can colonize wider spectrum of plant hosts. Their effect on these hosts is not yet fully understood. Most importantly, they increase the vitality of ericaceous plants and favor them in competition with adjacent...
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Characterization of fungal community in spruce (Picea abies) litter using cultivation and T-RFLP
Kolářová, Zuzana ; Koukol, Ondřej (advisor) ; Brabcová, Vendula (referee)
Fungi have a key role in the decomposition of coniferous litter and affect nutrient cycling in forest ecosystems. Therefore, great emphasis is placed on exploring the diversity of these organisms. The aim of this thesis was to describe fungal diversity in spruce litter and revealed temporal development of this community in a forest regenerating after bark beetle outbreak. Another objective was to compare sites with different length of bark beetle damage. The study area was located in the Bohemian Forest mountain range. Litter bags with spruce needles were placed on the forest soil and several samplings were performed in the course of three years. Diversity of fungi and changes in the fungal community were assessed by two methods: cultivation of needles on 2ř MEA and fingerprinting method T-RFLP. In total 71 fungal species were obtained from needles during a three-year succession by cultivation approach. Using T-RFLP 122 different fragments were generally recorded. The dominant species were Scleroconidioma sphagnicola, Thysanophora penicillioides, Hormonema dematiodes, Ceuthospora pinastri, species of genus Chalara, Trichoderma polysporum, Mycena galopus and unknown species Helotiales sp. 1. Primary saprotrophs occured in the community mainly in first 8 months and then were replaced by basidiomycetes....
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Ericaceae root mycobionts: diversity, distribution and interactions with neighboring plants
Kolátková, Viktorie ; Vohník, Martin (advisor) ; Baláž, Milan (referee)
The roots of ericaceous plants provide an environment to many fungal endophytes and mycorrhizal fungi.We can observe at least four mycorrhizal types in the family, of which the ericoid mycorrhiza (ErM) is spread the most. The Ericaceae are able to survive in many different and often harsh habitats all over the world thanks to this symbiosis (from the tropical cloud forests, to the arctic tundra heathlands). The group of fungi, forming this relationship, used to be seen as a few very well circumscribed ascomycetes, namely Rhizoscyphus ericae (Read) Zhang & Zhuang and Oidiodendron maius Barron species. With the development of molecular methods, it was expanded of many other species, some of which belonging to Basidiomycota, particularly the order Sebacinales. The diversity of ErM fungal community varies at different geographic locations and it can be influenced by factors such as altitude or the surrounding ecosystem. However, due to lack of further information, we still know quite little about the species distribution. Although they are primarily symbionts of the Ericaceae, some ErM fungi can colonize wider spectrum of plant hosts. Their effect on these hosts is not yet fully understood. Most importantly, they increase the vitality of ericaceous plants and favor them in competition with adjacent...
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Pyrosequencing analysis of fungal assemblages and the effect of ITS rDNA intragenomic variation on diversity estimates
Zelenka, Tomáš ; Kolařík, Miroslav (advisor) ; Větrovský, Tomáš (referee)
The ITS region of nuclear ribosomal DNA has recently become a frequently studied region for its use as a barcode marker. It is employed in environmental metagenomic analyses, the determination of fungi and in studies of fungal phylogenetics. In genomes, rDNA occurs in the form of large multicopy tandem arrays. Thus common Sanger sequencing leads to a consensus of many copies which in fact conceals most of the potential intragenomic variation. However 454 pyrosequencing reveals the sequence of single copies. Although it is believed that most of the variation among copies is reduced during a process called concerted evolution, some variation might still be conserved, including variation of non-functional pseudogenes. As a consequence, when using 454 pyrosequencing, we are not able to discriminate between this variation and real diversity. This could have a huge impact on the estimation of real diversity as well as on the correct assessment of pyrosequence studies. This thesis reviews current knowledge of intragenomic variation among fungi and summarizes some papers applying pyrosequencing in the research of fungal diversity. At the same time it indicates intragenomic variation as a potential cause of untrue diversity in diversity studies. Up until now there hasn't existed any experimental survey covering...
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Characterization of fungal community in spruce (Picea abies) litter using cultivation and T-RFLP
Kolářová, Zuzana ; Koukol, Ondřej (advisor) ; Brabcová, Vendula (referee)
Fungi have a key role in the decomposition of coniferous litter and affect nutrient cycling in forest ecosystems. Therefore, great emphasis is placed on exploring the diversity of these organisms. The aim of this thesis was to describe fungal diversity in spruce litter and revealed temporal development of this community in a forest regenerating after bark beetle outbreak. Another objective was to compare sites with different length of bark beetle damage. The study area was located in the Bohemian Forest mountain range. Litter bags with spruce needles were placed on the forest soil and several samplings were performed in the course of three years. Diversity of fungi and changes in the fungal community were assessed by two methods: cultivation of needles on 2ř MEA and fingerprinting method T-RFLP. In total 71 fungal species were obtained from needles during a three-year succession by cultivation approach. Using T-RFLP 122 different fragments were generally recorded. The dominant species were Scleroconidioma sphagnicola, Thysanophora penicillioides, Hormonema dematiodes, Ceuthospora pinastri, species of genus Chalara, Trichoderma polysporum, Mycena galopus and unknown species Helotiales sp. 1. Primary saprotrophs occured in the community mainly in first 8 months and then were replaced by basidiomycetes....
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