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Phylogenetic relationships of Auriculoscypha based on ultrastructural and molecular studies
Abstract
The phylogeny of Auriculoscypha anacardiicola, an associate of scale insects in India, is investigated using subcellular characters and MP and Bayesian analyses of combined nuLSU-rDNA, nuSSU-rDNA and 5.8S rDNA sequence data. It has simple septa with a pulley-wheel-shaped pore plug, which is diagnostic of phytoparasitic members of the Pucciniomycetes, and hyphal wall break on branching, a phenomenon unique to some simple septate heterobasidiomycetes. The septal ultrastructure of A. anacardiicola is similar to that of the genus Septobasidium. The close relationship to Septobasidium is also confirmed by rDNA sequence analyses. The polyphyletic nature of the order Platygloeales, noted in earlier studies, is evident from the present molecular analysis as well. The placement of Auriculoscypha in the Platygloeales can no longer be justified and both ultrastructural and molecular evidence strongly support the placement of Auriculoscypha in the Septobasidiales.
... This was followed by the observation by Lalitha et al. (1994) of a yeast phase in the life-cycle of Auriculoscypha, which supported its redisposition in Septobasidiales. Finally, based on molecular and ultrastructural characters, Kumar et al. (2007) unequivocally placed Auriculoscypha in the Septobasidiales. The only family of the order, the Septobasidiaceae, contains five genera, all of which are phytoparasitic insect symbionts (Swann et al., 2001). ...
... Auriculoscypha differs from all other genera of the family in having woody, stipitate-cupulate basidiomata. The basidiomata of all the other genera are simple mycelial mats that are resupinate on the host plant (Kumar et al., 2007). Lalitha (1992) provided a detailed account of the distribution of Auriculoscypha in southwest India. ...
Fungus-insect symbioses provide challenging and fascinating systems for studying biotic interactions between fungi, insects and other associated organisms (Klepzig et al., 2001). The types of interactions among these organisms are often difficult to pigeonhole as mutualism, antagonism, parasitism or commensalism. A remarkable but little-known interaction involving a fungus, a coccid and some tree species in southwest India is highlighted here. The sap of the plant is fed by the coccid that in turn is parasitized by the fungus, thus involving interactions between three trophic levels.
... as the only known species. 4 Several Basidiomycetes are known to be sources of physiologically beneficial and nontoxic medicines. 5 Also, many insect-pathogenic and insect-associated fungi are recognized as powerful sources of medicinally valuable bioactive compounds. ...
... After surface sterilization with 0.1% mercuric chloride solution for 30 sec, the basidiomata were thoroughly rinsed with sterilized water three times. Under aseptic conditions such basidiomata were dissected out and small (ca. 2 mm × 1 mm) pieces of pileal and stipe context were cut and transferred to Petri dishes containing sterilized potato dextrose agar (PDA) and incubated for 30 d. 4. Young actively growing basidiomata with the basal tubercle removed were surface sterilized A coccid dissected out from the tubercle. ...
Auriculoscypha anacardiicola, an obligate insect-associate and a potential medicinal fungus, is isolated and studied in vitro. Suitable methods for isolation and cultivation of the fungus have been developed. Incubating spore deposits made to fall from basidiomata on tap water agar seems to be the best method for developing cultures. Successful isolations were also accomplished from infected coccids. Cultures could not be developed from single basidiospores and from tissues of the basidiomata. Although production of ballistospores and blastospores as well as germ tube formation were observed at the time of germination of basidiospores, budding blastospores alone produced mycelial cultures. Observations such as the inability of single basidiospores to germinate, emergence of mycelium from a spore deposit, and the apparent conjugation of yeast cells indicate that dikaryotization resulting from fusion of compatible yeast cells is essential for development of mycelium in A. anacardiicola. The fungus grew well on all complex media tested. It seems that a purely synthetic medium devoid of any growth factors cannot support the growth of A. anacardiicola and yeast extract seems to provide the required growth factors.
... as the only known species. 4 Several Basidiomycetes are known to be sourc- es of physiologically beneficial and nontoxic medi- cines. 5 Also, many insect-pathogenic and insect-as- sociated fungi are recognized as powerful sources of medicinally valuable bioactive compounds. ...
... 4. Young actively growing basidiomata with the basal tubercle removed were surface sterilized culture revealed that it was indeed a culture of a fungus belonging to the Septobasidiales. 4 ...
Auriculoscypha anacardiicola, an obligate insect-associate and a potential medicinal fungus, is isolated and studied in vitro. Suitable methods for isolation and cultivation of the fungus have been developed. Incubating spore deposits made to fall from basidiomata on tap water agar seems to be the best method for developing cultures. Successful isolations were also accomplished from infected coccids. Cultures could not be developed from single basidiospores and from tissues of the basidiomata. Although production of ballistospores and blastospores as well as germ tube formation were observed at the time of germination of basidiospores, budding blastospores alone produced mycelial cultures. Observations such as the inability of single basidiospores to germinate, emergence of mycelium from a spore deposit, and the apparent conjugation of yeast cells indicate that dikaryotization resulting from fusion of compatible yeast cells is essential for development of mycelium in A. anacardiicola. The fungus grew well on all complex media tested. It seems that a purely synthetic medium devoid of any growth factors cannot support the growth ofA. anacardiicola and yeast extract seems to provide the required growth factors.
... We measured T. protuberans pores in Smith's fig- ure 1B and C to be 1.4 9 1.2 and 2.8 9 1.2 lm, respectively. The single, central pores in Ascomycota and Basidiomycota range from c. 0.04 to 0.25 lm in diameter (Ellis, Rogers & Mims, 1972;Berndt & Oberwinkler, 1992;Momany et al., 2002;Kumar et al., 2007Kumar et al., , 2012). Members of Kickxellomycotina (see Fig. 1 for phylogenetic position) and the related subphylum Entomophthoromycotina possess single septal pores that reach c. 0.5 lm in diameter, although in contrast with T. protuberans, the sur- rounding walls appear lens-shaped in cross-section (Benny & Aldrich, 1975;Jeffries & Young, 1979;Saikawa, 1989;Saikawa, Oguchi & Ruiz, 1997). ...
In this commentary, we discuss evidence for the phylogenetic affiliations of Tortotubus protuberans, the subject of Martin Smith's 2016 paper in the Botanical Journal of the Linnean Society entitled, ‘Cord-forming Palaeozoic fungi in terrestrial assemblages’. We agree that the fossilized, branching, somatic filaments probably represent fungal hyphae. We were not convinced by Smith's proposal that T. protuberans represents Dikarya, the clade of fungi that includes most modern moulds, yeasts and mushrooms. To justify classification, Smith relied on structures that are analogous between T. protuberans and modern fungi, and argued ‘that Dikarya can produce the range of morphologies expressed by T. protuberans’. We review available information about homologies of the characteristics of T. protuberans, including mycelial cords, retrograde growth, septal pores and ornamented hyphae. Retrograde growth in T. protuberans is intriguing from an evolutionary developmental point of view, but it differs sufficiently in fine detail when compared with growth patterns in croziers or clamp connections of Dikarya, so that homologies are unclear. Tortotubus protuberans is an important fossil form, but we suggest taking a step back and relating it to the distribution of character evolution through the fungal phylogeny rather than to derived characters of modern taxa.
... Malasseziaceae is a monotypic family. The current placement of the Malasseziaceae in the system of the ustilaginomycetous fungi and associated yeasts is based on results obtained from molecular phylogenetic analyses (Begerow et al. 2000, 2007, Fell et al. 2000, Sugita et al. 2002, Sampaio 2004, Weiss et al. 2004, Kumar et al. 2007; see also Hibbet et al. 2007 ...
Names of two families in the Exobasidiomycetes, Malasseziaceae and Ceraceosoraceae, are validated.
... The processing of one week old hyphal samples of W. sebi (grown on 2% ME and 20% dextrose agar) for septal ultrastructural studies followed Kumar et al. (Kumar et al., 2007), except that the samples were dehydrated in a graded ethanol series, and infiltrated using Spurr's low viscosity resin (Spurr, 1969) prepared using the modified formulation of Ellis (Ellis, 2006). ...
Wallemia (Wallemiales, Wallemiomycetes) is a genus of xerophilic Fungi of uncertain phylogenetic position within Basidiomycota. Most commonly found as food contaminants, species of Wallemia have also been isolated from hypersaline environments. The ability to tolerate environments with reduced water activity is rare in Basidiomycota. We sequenced the genome of W. sebi in order to understand its adaptations for surviving in osmotically challenging environments, and we performed phylogenomic and ultrastructural analyses to address its systematic placement and reproductive biology. W. sebi has a compact genome (9.8 Mb), with few repeats and the largest fraction of genes with functional domains compared with other Basidiomycota. We applied several approaches to searching for osmotic stress-related proteins. In silico analyses identified 93 putative osmotic stress proteins; homology searches showed the HOG (High Osmolarity Glycerol) pathway to be mostly conserved. Despite the seemingly reduced genome, several gene family expansions and a high number of transporters (549) were found that also provide clues to the ability of W. sebi to colonize harsh environments. Phylogenetic analyses of a 71-protein dataset support the position of Wallemia as the earliest diverging lineage of Agaricomycotina, which is confirmed by septal pore ultrastructure that shows the septal pore apparatus as a variant of the Tremella-type. Mating type gene homologs were identified although we found no evidence of meiosis during conidiogenesis, suggesting there may be aspects of the life cycle of W. sebi that remain cryptic.
Wallemia (Wallemiales, Wallemiomycetes) is a genus of xerophilic Fungi
that is of uncertain phylogenetic position within the Basidiomycota. Most commonly
found as food contaminants, species of Wallemia have also been isolated
from hypersaline environments. The ability to tolerate low water environments
is rare in the Basidiomycota and suggests several adaptations involving
osmoregulation. We sequenced the genome of Wallemia sebi in order to test
whether it possesses any unique genomic adaptations for surviving osmotically
challenging environments, and coupled genome and phylogenetic analyses with
ultrastructural studies aimed at addressing other questions about its biology.
Compared with other Basidiomycota, W. sebi has a compact genome (9.8 Mb),
with few repeats and the largest fraction of genes with functional domains. In
silico analyses identified >90 putative osmotic stress proteins, the domains of
which possess a relatively high number of transporters that may be correlated
with the ability to live under osmotic stress. Despite the seemingly reduced
genome, several gene family expansions were revealed that also provide cluesto the ability of W. sebi to colonize harsh environments. Phylogenetic analyses
of 71 single protein datasets support the position of Wallemia as the earliest diverging
lineage of Agaricomycotina, which is confirmed by septal pore ultrastructure.
Mating type gene homologs were identified although we found no evidence
of meiosis during conidiogenesis, suggesting there may be aspects of the
life cycle of W. sebi that remain cryptic.
A field survey of ballistosporic yeasts in a Neotropical forest yielded a new species isolated from a fern leaf. The isolate is a cream-colored butyrous yeast that reproduces by budding. Budding occurs at both the apical and basal cell poles; occasionally multiple budding events co-occur, giving rise to rosette-like clusters of cells at both poles of the yeast mother cell. DNA sequences of large and small subunit and the internal transcribed spacer regions of the nuclear ribosomal DNA cistron indicated an affinity to Microbotryomycetes, Pucciniomycotina. A new genus, Meredithblackwellia, is proposed to accommodate the new species, M. eburnea (type strain MCA4105). Based on phylogenetic analyses, Meredithblackwellia is related to Kriegeria eriophori, a sedge parasite, to an aquatic fungus Camptobasidium hydrophilum and to several recently described anamorphic yeasts that have been isolated from plant material or psychrophilic environments. Morphological and ultrastructural studies confirm the relatedness of M. eburnea to these taxa and prompted the re-evaluation of higher-level classification within Microbotryomycetes. We propose here a new order, Kriegeriales, and place two families, Kriegeriaceae fam. nov. and Camptobasidiaceae R.T. Moore, within it. Our study re-emphasizes the need for systematic revision of species described in Rhodotorula.
This is the one essential handbook for all who work with or are interested in fungi (including lichens, slime moulds, yeasts and fungal analogues).This new edition, with more than 21,000 entries, provides the most complete listing available of generic names of fungi, their families and orders, their attributes and descriptive terms. For each genus, the authority, the date of publication, status, systematic position, number of accepted species, distribution, and key references are given. Diagnoses of families and details of orders and higher categories are included for all groups of fungi. In addition, there are biographic notes, information on well-known metabolites and mycotoxins, and concise accounts of almost all pure and applied aspects of the subject (including citations of important literature).All information has been updated as necessary since the publication of the ninth edition in 2001. In addition the tenth edition has the following new features: a completely new classification of the Kingdom Fungi based on recent multi-gene phylogenetics research; a major revision of the classification of the Basidiomycota and substantial modification of the "basal" groups; further integration of anamorphic and teleomorphic genera in the classification; enhanced distinctions between the true fungi and unrelated groups traditionally studied by mycologists; and, improved information on references to publications.
The simple-septate basidiomycetes comprise more than 8,000 species that show a high morphological and ecological heterogeneity. To gain insight in the phylogenetic relationships within this group, we compared several ultrastructural features such as septal pore apparatus, form, and behavior of the spindle pole bodies, types of host–parasite interaction, presence or absence of colacosomes, symplechosomes, atractosomes, and cystosomes as well as nuclear rDNA sequences coding for small- and large-subunit rRNA. Based on our integrated analysis, we propose a new classification system for the simple-septate basidiomycetes with the subphylum Pucciniomycotina and the classes Agaricostilbomycetes, Atractiellomycetes, Classiculomycetes, Cryptomycocolacomycetes, Cystobasidiomycetes, Microbotryomycetes, Mixiomycetes, and Pucciniomycetes. We also propose the pucciniomycotinous taxa Cystobasidiales, Erythrobasidiales, Helicobasidiales, Mixiales, Naohideales, Pachnocybales, Spiculogloeales, and Kondoaceae and the new subphyla Agaricomycotina (equivalent to the current Hymenomycetes) and Ustilaginomycotina (equivalent to the current Ustilaginomycetes).
The subclass Microbotryomycetidae (Basidiomycota, Urediniomycetes) comprises a remarkably diverse assemblage of fungi. This
group includes phytoparasites, mycoparasites and probably also saprobes that show a wide range of ecological preferences.
In order to study the phylogenetic relationships within the Microbotryomycetidae, and to develop a more natural classification
system, mitosporic and meiosporic taxa were investigated using an integrated approach. Sequence data of 26S rDNA D1/D2 domains
were analyzed using several procedures, including the Bayesian Markov chain Monte Carlo method of phylogenetic inference.
Ultrastructural markers such as the type of septal pore and presence / absence of colacosomes were investigated and micromorphological
and nutritional properties were compared. In this study the current concept of the genus Leucosporidium and its apparent polyphyletic nature were addressed, as well as the relationships of this genus with the Microbotryales and
Mastigobasidium. The classification of the anamorphic species closely related to Leucosporidium, and the concepts of the order Sporidiales and family Sporidiobolaceae were also reviewed.
Sequence data from nuclear large subunit ribosomal DNA was used to infer phylogenetic relationships of selected genera of the Uredinales. We investigated 52 rust fungi representing nine families and three outgroup species. Neighbor joining analysis and a Bayesian method of phylogenetic inference using Monte Carlo Markov chains confirm the rust fungi as a natural group and indicate that Puccinia, Uromyces, Endophyllum, and Cumminsiella have a common origin. The autoecious Rosaceae-rusts Phragmidium, Kuehneola, Triphragmium, and Trachyspora are a monophyletic group. The gasteroid genus Ochropsora is closely related to Tranzschelia. While the Pucciniastreae sensu Dietel (1938) is recognized as a monophyletic group in neighbor joining analysis, the Pucciniaceae s.l. (Dietel 1928) is supported by Bayesian analysis. The following genera appear to be monophyletic: Chrysomyxa, Coleosporium, Cronartium, Gymnosporangium, Melampsora, Phragmidium, and Tranzschelia, whereas the genera Puccinia, Pucciniastrum, Thekopsora, and Uromyces are not.
The sequence of the small subunit nuclear ribosomal DNA (18S rDNA) was determined for seven selected species of the teliospore-forming yeasts and Filobasidiaceae in the basidiomycetous yeasts. A phylogenetic tree, including published reference sequences, was inferred from 1623 sites which could be unambiguously aligned. The molecular phylogeny, using a chytridiomycete as an outgroup, divided the eight basidiomycetous yeasts into two groups which correlated well with both septal ultrastructure (simple pore or dolipore) and cellular xylose (present or absent). The first group included the teliospore-forming yeasts Rhodosporidium toruloides and Leucosporidium scottii, and Erythrobasidium hasegawianum, which is currently a member of the Filobasidiaceae. The second group was formed by the filobasidiaceous yeasts, comprising a Cystofilobasidium capitatum/Leucosporidium lari-marini/Mrakia frigida branch, and a Filobasidium floriforme/Filobasidiella neoformans branch. Our molecular data support the principal chemotaxonomic and ultrastructural evidence, which indicates a very close affinity between C. capitatum and L. lari-marini.
As a discipline, phylogenetics is becoming transformed by a flood of molecular data. These data allow broad questions to be asked about the history of life, but also present difficult statistical and computational problems. Bayesian inference of phylogeny brings a new perspective to a number of outstanding issues in evolutionary biology, including the analysis of large phylogenetic trees and complex evolutionary models and the detection of the footprint of natural selection in DNA sequences.
MrBayes 3 performs Bayesian phylogenetic analysis combining information from different data partitions or subsets evolving under different stochastic evolutionary models. This allows the user to analyze heterogeneous data sets consisting of different data types—e.g. morphological, nucleotide, and protein—and to explore a wide variety of structured models mixing partition-unique and shared parameters. The program employs MPI to parallelize Metropolis coupling on Macintosh or UNIX clusters.
Availability: http://morphbank.ebc.uu.se/mrbayes
Contact: fredrik.ronquist@ebc.uu.se
*
To whom correspondence should be addressed.
In this study we provide a phylogenetically based introduction to the classes and orders of Pucciniomycotina (= Urediniomycetes), one of three subphyla of Basidiomycota. More than 8000 species of Pucciniomycotina have been described including putative saprotrophs and parasites of plants, animals and fungi. The overwhelming majority of these (approximately 90%) belong to a single order of obligate plant pathogens, the Pucciniales (= Uredinales), or rust fungi. We have assembled a dataset of previously published and newly generated sequence data from two nuclear rDNA genes (large subunit and small subunit) including exemplars from all known major groups in order to test hypotheses about evolutionary relationships among the Pucciniomycotina. The utility of combining nuc-lsu sequences spanning the entire D1-D3 region with complete nuc-ssu sequences for resolution and support of nodes is discussed. Our study confirms Pucciniomycotina as a monophyletic group of Basidiomycota. In total our results support eight major clades ranked as classes (Agaricostilbomycetes, Atractiellomycetes, Classiculomycetes, Cryptomycocolacomycetes, Cystobasidiomycetes, Microbotryomycetes, Mixiomycetes and Pucciniomycetes) and 18 orders.
DNA sequences of the nuclear encoded small subunit (SSU) 18S ribosomal RNA gene and characters from a morphological/physiological dataset were analyzed by maximum parsimony to investigate phylogenetic relationships among and within two putative sister orders of Zygomycota that produce regularly septate hyphae with ultrastructurally similar plugged, flared septal pores, the Harpellales and Kickxellales. The most parsimonious trees (MPT) inferred from the molecular data, using Chytridium-Spizellomyces and Rhopalomyces as outgroups in separate analyses, were largely concordant. Cladistic analysis of nucleotide characters identified three strongly supported monophyletic lineages that formed an unresolved trichotomy in the 18S rDNA gene trees: the Harpellales, Kickxellales and a Spiromyces clade. Trees inferred from the nonmolecular data were largely unresolved and generally poorly supported by bootstrap and decay analyses. However, results of a partition-homogeneity test detected no positive conflict between the morphological and molecular datasets, indicating that they could be combined. Morphological evolution was investigated by optimizing nonmolecular characters on the 18S rDNA gene tree. Systematic results support the recognition of a Spiromyces clade independent of the Kickxellales where it is currently classified and the apparent paraphyly of Coemansia with Spirodactylon derived within it. A formal diagnosis of Spiromyces aspiralis sp. nov. is provided together with an emendation of the genus.
Until recently the sedge parasite Kriegeria eriophori has never been satisfactorily classified above the genus level. Its classification has been problematic in that a number of characters at the gross micromorphological and ultrastructural levels appeared to be autapomorphic. The use of nucleotide sequence data has proven to be a powerful means of addressing difficult systematic problems in which standard morphological approaches have failed to provide sufficient information. During this study, some of these ultrastructural characters were discovered to be synapomorphies for a group containing K. eriophori and Microbotryum violaceum, a urediniomycete smut species. Ribosomal RNA gene sequences and nucleus-associated ultrastructural characters were analysed separately and combined. These characters serve to define the new subclass Microbotryomycetidae. Inclusion of the sequences in the analysis provided an independent data set by which the ultrastructural character states were polarized. Gross micromorphological characters proved to be homoplasious, and of less utility in determining phylogenetic groups than were ultrastructural and nucleotide characters. Cladistic analysis of ultrastructural and biochemical characters provides a powerful means of identifying monophyletic groups, the prerequisite for creating a phylogenetic classification. A phylogenetic diagnosis is provided for the Microbotryomycetidae, which deviates from the pattern used in typical diagnoses.
— We studied sequence variation in 16S rDNA in 204 individuals from 37 populations of the land snail Candidula unifasciata (Poiret 1801) across the core species range in France, Switzerland, and Germany. Phylogeographic, nested clade, and coalescence analyses were used to elucidate the species evolutionary history. The study revealed the presence of two major evolutionary lineages that evolved in separate refuges in southeast France as result of previous fragmentation during the Pleistocene. Applying a recent extension of the nested clade analysis (Templeton 2001), we inferred that range expansions along river valleys in independent corridors to the north led eventually to a secondary contact zone of the major clades around the Geneva Basin. There is evidence supporting the idea that the formation of the secondary contact zone and the colonization of Germany might be postglacial events. The phylogeographic history inferred for C. unifasciata differs from general biogeographic patterns of postglacial colonization previously identified for other taxa, and it might represent a common model for species with restricted dispersal.
The nucleotide sequences of the 5′ end of the nuclear large subunit ribosomal RNA were obtained for 15 basidiomycetes representing a diversity of orders, including Agaricostilbales, Atractiellales, Auriculariales sensu lato, Boletales, Septobasidiales, Tilletiales, Uredinales and Ustilaginales. These sequences were analyzed together with additional GenBank sequences for three basidiomycetes, five ascomycetes, and a zygomycete, using methods of maximum likelihood, maximum parsimony and minimum-evolution. Using Mucor racemosus as an outgroup, the basidiomycetes analyzed under models of maximum likelihood and maximum parsimony formed nearly identical trees each composed of seven distinct clades: the smuts and bunts, a complex-septate group, two separate groups of auriculariaceous simple-septate species, the rusts and two unique branches each comprising a single taxon Agaricostilbum pulcherrimum and Cryptococcus neoformans. The minimum-evolution topology was different from the former two trees in the placement of the smuts and bunts with respect to the complex septate taxa, the position of Agaricostilbum pulcherrimum and the localized structure of the clade containing the taxa Eocronartium muscicola, Pachnocybe ferruginea and Septobasidium carestianum. Relative rate tests of nucleotide substitutions based on maximum likelihood estimations suggest significant evolutionary rate differences among the orders. To statistically differentiate among the different topologies, all trees were subjected to a likelihood variance test. Under assumptions of maximum likelihood, the test rejected the minimum-evolution tree but did not discriminate against any of the topologies generated by either maximum likelihood or maximum parsimony. These results suggest that the 5′ end of the nuclear large subunit ribosomal DNA is useful for intergeneric comparisons of basidiomycetous fungi. The sequence data clarify the relationships among the genera of the Auriculariales sensu lato and support the hypothesis that the Uredinales are a derived group.
The redisposition of Auriculoscypha from the Auriculariales to the Septobasidiales is suggested based on the observation that the fungus is closely associated with a coccid.
Various patterns of basidiospore germination in Auriculoscypha anacardiicola are described and compared with those of related genera.
Auriculoscypha gen.nov. is described with A. anacardiicola sp. nov. as the type species.
The recently-developed statistical method known as the "bootstrap" can be used to place confidence intervals on phylogenies. It involves resampling points from one's own data, with replacement, to create a series of bootstrap samples of the same size as the original data. Each of these is analyzed, and the variation among the resulting estimates taken to indicate the size of the error involved in making estimates from the original data, In the case of phylogenies, it is argued that the proper method of resampling is to keep all of the original species while sampling characters with replacement, under the assumption that the characters have been independently drawn by the systematist and have evolved independently. Majority-rule consensus trees can be used to construct a phylogeny showing all of the inferred monophyletic groups that occurred in a majority of the bootstrap samples. If a group shows up 95% of the time or more, the evidence for it is taken to be statistically significant. Existing computer programs can be used to analyze different bootstrap samples by using weights on the characters, the weight of a character being how many times it was drawn in bootstrap sampling. When all characters are perfectly compatible, as envisioned by Hennig, bootstrap sampling becomes unnecessary; the bootstrap method would show significant evidence for a group if it is defined by three or more characters.
We have designed two taxon-selective primers for the internal transcribed spacer (ITS) region in the nuclear ribosomal repeat unit. These primers, ITS1-F and ITS4-B, were intended to be specific to fungi and basidiomycetes, respectively. We have tested the specificity of these primers against 13 species of ascomycetes, 14 of basidiomycetes, and 15 of plants. Our results showed that ITS4-B, when paired with either a ‘universal’ primer ITS1 or the fungal-specific primer ITS1-F, efficiently amplified DNA from all basidiomycetes and discriminated against ascomycete DNAs. The results with plants were not as clearcut. The ITS1-F/ITS4-B primer pair produced a small amount of PCR product for certain plant species, but the quantity was in most cases less than that produced by the ‘universal’ ITS primers. However, under conditions where both plant and fungal DNAs were present, the fungal DNA was amplified to the apparent exclusion of plant DNA. ITS1-F/ITS4-B preferential amplification was shown to be particularly useful for detection and analysis of the basidiomycete component in ectomycorrhizae and in rust-infected tissues. These primers can be used to study the structure of ectomycorrhizal communities or the distribution of rusts on alternate hosts.
Rusts (Basidiomycetes: Uredinales) are a large, diverse group of obligate biotrophic fungi that include many important plant
pathogens. Phylogenetic relationships within the group are poorly understood, with between two and 14 families being recognised
on morphological and host taxonomic grounds. We report the first analysis of broad-scale relationships within the Uredinales
using DNA sequence data from the small subunit gene of the rRNA operon. A major split is obvious between genera that have
aecial stages on gymnosperm hosts and those with this stage on angiosperms. This finding is in accord with the early taxonomic
two-family treatments of the Uredinales. Within the major clades, there is evidence of in congruency with morphologically
defined families. Racospermyces, Maravalia, Hemileia and Caeoma are basal to the major phylogenetic sub-division and probably represent the most ancient rust lineages. Using these sequence
data, we estimate that the rusts diverged from their closest relatives around 150 million years ago.
Additional keywords:molecular phylogeny
Small subunit (18S) rRNA gene sequences from 38 species of fungi (three ascomycetes, 35 basidiomycetes) were analysed using parsimony. The resulting phylogeny supported a hypothesis of three major lineages within the basidiomycetes. The lineages are recognized as classes, the Urediniomycetes, Ustilaginomycetes and Hymenomycetes. Each lineage contained some yeasts or yeast producing taxa. The systematic relationships of basidiomycetous yeasts are discussed with reference to the three class taxonomic system.
This paper describes a simple technique for consistent production of clean, unwrinkled, flat thin (500-1000 A) sections for TEM and thick (1/2-1 micron) sections for HVEM mounted on Formvar-covered slot grids for use in conventional and high voltage electron microscopy. The technique centers around use of a Formvar-covered aluminum supporting rack onto which slot grids that contain sections suspended in water are placed.
Detailed restriction analyses of many samples often require substantial amounts of time and effort for DNA extraction, restriction digests, Southern blotting, and hybridization. We describe a novel approach that uses the polymerase chain reaction (PCR) for rapid simplified restriction typing and mapping of DNA from many different isolates. DNA fragments up to 2 kilobase pairs in length were efficiently amplified from crude DNA samples of several pathogenic Cryptococcus species, including C. neoformans, C. albidus, C. laurentii, and C. uniguttulatus. Digestion and electrophoresis of the PCR products by using frequent-cutting restriction enzymes produced complex restriction phenotypes (fingerprints) that were often unique for each strain or species. We used the PCR to amplify and analyze restriction pattern variation within three major portions of the ribosomal DNA (rDNA) repeats from these fungi. Detailed mapping of many restriction sites within the rDNA locus was determined by fingerprint analysis of progressively larger PCR fragments sharing a common primer site at one end. As judged by PCR fingerprints, the rDNA of 19 C. neoformans isolates showed no variation for four restriction enzymes that we surveyed. Other Cryptococcus spp. showed varying levels of restriction pattern variation within their rDNAs and were shown to be genetically distinct from C. neoformans. The PCR primers used in this study have also been successfully applied for amplification of rDNAs from other pathogenic and nonpathogenic fungi, including Candida spp., and ought to have wide applicability for clinical detection and other studies.
This article reports on the use of high pressure freezing followed by freeze substitution (HPF/FS) to study ultrastructural details of host-pathogen interactions in fungal diseases of plants. The specific host-pathogen systems discussed here include a powdery mildew infection of poinsettia and rust infections of daylily and Indian strawberry. The three pathogens considered here all attack the leaves of their hosts and produce specialized hyphal branches known as haustoria that invade individual host cells without killing them. We found that HPF/FS provided excellent preservation of both haustoria and host cells for all three host-pathogen systems. Preservation of fungal and host cell membranes was particularly good and greatly facilitated the detailed study of host-pathogen interfaces. In some instances, HPF/FS provided information that was not available in samples prepared for study using conventional chemical fixation. On the other hand, we did encounter various problems associated with the use of HPF/FS. Examples included freeze damage of samples, inconsistency of fixation in different samples, separation of plant cell cytoplasm from cell walls, breakage of cell walls and membranes, and splitting of thin sections. However, we believe that the outstanding preservation of ultrastructural details afforded by HPF/FS significantly outweighs these problems and we highly recommend the use of this fixation protocol for future studies of fungal host-plant interactions.
Urediniomycetes The Mycota. Volume II Part B. Systematics and Evolution
- Ec Swann
- Em Frieders
- Dj Mclaughlin
Swann EC, Frieders EM, McLaughlin DJ, 2001. Urediniomycetes. In:
McLaughlin DJ, McLaughlin EG, Lemke PA (eds), The Mycota.
Volume II Part B. Systematics and Evolution. Springer-Verlag,
Berlin, pp. 37–56.
Systematics and Evolution
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- B Part
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