Biomedical subjects

R Vilgalys

Publications and source records attributed to R Vilgalys.

8 recordsLinked to original sources

PubMed1996

Molecular markers reveal that population structure of the human pathogen Candida albicans exhibits both clonality and recombination.

The life history of Candida albicans presents an enigma: this species is thought to be exclusively asexual, yet strains show extensive phenotypic variation. To address the population genetics of C. albicans, we developed a genetic typing method for codominant single-locus markers by screening randomly amplified DNA for single-strand conformation polymorphisms. DNA fragments amplified by arbitrary primers were initially screened for single-strand conformation polymorphisms and later sequenced using locus-specific primers. A total of 12 single base mutations and insertions were detected from six out of eight PCR fragments. Patterns of sequence-level polymorphism observed for individual strains detected considerable heterozygosity at the DNA sequence level, supporting the view that most C. albicans strains are diploid. Population genetic analyses of 52 natural isolates from Duke University Medical Center provide evidence for both clonality and recombination in C. albicans. Evidence for clonality is supported by the presence of several overrepresented genotypes, as well as by deviation of genotypic frequencies from random (Hardy-Weinberg) expectations. However, tests for nonrandom association of alleles across loci reveal less evidence for linkage disequilibrium than expected for strictly clonal populations. Although C. albicans populations are primarily clonal, evidence for recombination suggests that sexual reproduction or some other form of genetic exchange occurs in this species.

Candida albicans↗

PubMed1996

Ancient single origin for Malagasy primates.

We report new evidence that bears decisively on a long-standing controversy in primate systematics. DNA sequence data for the complete cytochrome b gene, combined with an expanded morphological data set, confirm the results of a previous study and again indicate that all extant Malagasy lemurs originated from a single common ancestor. These results, as well as those from other genetic studies, call for a revision of primate classifications in which the dwarf and mouse lemurs are placed within the Afro-Asian lorisiforms. The phylogenetic results, in agreement with paleocontinental data, indicate an African origin for the common ancestor of lemurs and lorises (the Strepsirrhini). The molecular data further suggest the surprising conclusion that lemurs began evolving independently by the early Eocene at the latest. This indicates that the Malagasy primate lineage is more ancient than generally thought and places the split between the two strepsirrhine lineages well before the appearance of known Eocene fossil primates. We conclude that primate origins were marked by rapid speciation and diversification sometime before the late Paleocene.

Animals↗

PubMed1996

Molecular evolutionary dynamics of cytochrome b in strepsirrhine primates: the phylogenetic significance of third-position transversions.

DNA sequences of the complete cytochrome b gene are shown to contain robust phylogenetic signal for the strepsirrhine primates (i.e., lemurs and lorises). The phylogeny derived from these data conforms to other molecular studies of strepsirrhine relationships despite the fact that uncorrected nucleotide distances are high for nearly all intrastrepsirrhine comparisons, with most in the 15%-20% range. Cytochrome b sequences support the hypothesis that Malagasy lemuriforms and Afro-Asian lorisiforms each comprise clades that share a sister-group relationship. A study (Adkins and Honeycutt 1994) of the cytochrome c oxidase subunit II (COII) gene placed one Malagasy primate (Daubentonia) at the base of the strepsirrhine clade, thereby suggesting a diphyletic Lemuriformes. The reanalysis of COII third-position transversions, either alone or in combination with cytochrome b third-position transversions, however, yields a tree that is congruent with phylogenetic hypotheses derived from cytochrome b and other genetic data sets.

Animals↗

PubMed1995

Analysis of genes coding for small-subunit rRNA sequences in studying phylogenetics of dematiaceous fungal pathogens.

Because of their ability to display yeast-like growth forms in various environmental conditions, dematiaceous (melanized) hyphomycetes of the form-genera Exophiala, Rhinocladiella, and Wangiella have been informally termed "black yeasts." Cladistic analysis of 1,050 bp of the genes coding for small-subunit rRNA (SSU rDNA) supported a close relationship among species of these black yeasts with other dematiaceous hyphomycetes in the form-genera Fonsecaea, Phialophora, and Ramichloridium. The conventional categories of these fungi based on asexual states are not supported by phylogenetic analysis of SSU rDNA sequences. Isolates exhibiting annellidic modes of blastic conidiogenesis (e.g., Exophiala spp.) were not monophyletic and were placed as sister taxa to isolates that produce phialides or sympodulae. The results indicated very close relationships between isolates of Wangiella dermatitidis and Exophiala mansonii and between Rhinocladiella aquaspersa and Exophiala jeanselmei. This clade of dematiaceous hyphomycetes was a sister group to a clade comprising members of two orders of cleistothecial ascomycetes, Eurotiales and Onygenales. The etiological agents of chromoblastomycosis were found to be a closely related group (clade), while the agents of phaeohyphomycosis displayed a broader distribution on the SSU rDNA tree.

Ascomycota↗

PubMed1994

Ancient and recent patterns of geographic speciation in the oyster mushroom Pleurotus revealed by phylogenetic analysis of ribosomal DNA sequences.

Evidence from molecular systematic studies suggests that many mushroom species may be quite ancient. Gene phylogenies were developed to examine the relationship between reproductive isolation, genetic divergence, and biogeography in oyster mushrooms (Pleurotus). Sequence data were obtained for two regions of DNA from populations belonging to eight intersterility groups (biological species). Phylogenetic analysis of sequences from the 5' portion of the nuclear encoded large subunit rDNA demonstrates an ancient origin for four intersterility groups of broad geographic distribution (world-wide), with a more recent radiation of several intersterility groups that are restricted to the Northern Hemisphere. An expanded analysis using sequence data from the more variable rDNA internal transcribed spacer region also reveals a phylogenetically based pattern of genetic divergence associated with allopatric speciation among populations from different continents in the Northern Hemisphere. The ability of rDNA sequences to resolve phylogenetic relationships among geographically isolated populations within intersterility groups illustrates the importance of biogeography for understanding speciation in Pleurotus. Patterns of geographic distribution among intersterility groups suggest that several species lineages evolved quite early, with recently evolved groups restricted to the Northern Hemisphere and older lineages occurring throughout the world. Based on phylogenetic evidence, analysis of historical biogeography using area cladograms shows that multiple dispersal and vicariance events are responsible for patterns of speciation observed.

Base Sequence↗

PubMed1993

Hybridization probes for conventional DNA fingerprinting used as single primers in the polymerase chain reaction to distinguish strains of Cryptococcus neoformans.

In conventional DNA fingerprinting, hypervariable and repetitive sequences (minisatellite or microsatellite DNA) are detected with hybridization probes. As demonstrated here, these probes can be used as single primers in the polymerase chain reaction (PCR) to generate individual fingerprints. Several conventional DNA fingerprinting probes were used to prime the PCR, yielding distinctive, hypervariable multifragment profiles for different strains of Cryptococcus neoformans. PCR fingerprinting with the oligonucleotide primers (GTG)5, (GACA)4, and the phage M13 core sequence (GAGGGTGGXGGXTCT), but not with (CA)8 or (CT)8, generated DNA polymorphisms with all 42 strains of C. neoformans investigated. PCR fingerprints produced by priming with (GTG)5, (GACA)4, or the M13 core sequence differentiated the two varieties of C. neoformans, C. neoformans var. neoformans (serotypes A and D) and C. neoformans var. gattii (serotypes B and C). Furthermore, strains of serotypes A, D, and B or C could be distinguished from each other by specific PCR fingerprint patterns. These primers, which also successfully amplified hypervariable DNA segments from other species, provide a convenient method of identification at the species or individual level. Amplification of polymorphic DNA patterns by PCR with these primers offers several advantages over classical DNA fingerprinting techniques, appears to be more reliable than other PCR-based methods for detecting polymorphic DNA, such as analysis of random-amplified polymorphic DNA, and should be applicable to many other organisms.

Base Sequence↗

PubMed1992

Evolutionary relationships within the fungi: analyses of nuclear small subunit rRNA sequences.

Nucleotide sequences of the small subunit ribosomal RNA (18S) gene were used to investigate evolutionary relationships within the Fungi. The inferred tree topologies are in general agreement with traditional classifications in the following ways: (1) the Chytridiomycota and Zygomycota appear to be basal groups within the Fungi. (2) The Ascomycota and Basidiomycota are a derived monophyletic group. (3) Relationships within the Ascomycota are concordant with traditional orders and divide the hemi- and euascomycetes into distinct lineages. (4) The Basidiomycota is divided between the holobasidiomycetes and phragmobasidiomycetes. Conflicts with traditional classification were limited to weakly supported branches of the tree. Strongly supported relationships were robust to minor changes in alignment, method of analysis, and various weighting schemes. Weighting, either of transversions or by site, did not convincingly improve the status of poorly supported portions of the tree. The rate of variation at particular sites does not appear to be independent of lineage, suggesting that covariation of sites may be an important phenomenon in these genes.

Animals↗

PubMed1990

Rapid genetic identification and mapping of enzymatically amplified ribosomal DNA from several Cryptococcus species.

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.

Base Sequence↗