Ribosomal DNA Repeat Research Articles

Phylogenetic Analysis of Subgenus Vigna Species Using Nuclear Ribosomal RNA ITS: Evidence of Hybridization among Vigna unguiculata Subspecies

Molecular phylogeny among species belonging to subgenus Vigna (genus Vigna) was inferred based on internal transcribed spacer (ITS) sequences of 18S-5.8S-26S ribosomal RNA gene unit. Analysis showed a total of 356 polymorphic sites of which approximately 80% were parsimony informative. Phylogenetic reconstruction by neighbor joining and maximum parsimony methods placed the 57 Vigna accessions (belonging to 15 species) into 5 major clades. Five species viz. Vigna heterophylla, Vigna pubigera, Vigna parkeri, Vigna laurentii, and Vigna gracilis whose position in the subgenus was previously not known were placed in the section Vigna. A single accession (Vigna unguiculata ssp. tenuis, NI 1637) harbored 2 intragenomic ITS variants, indicative of 2 different types of ribosomal DNA (rDNA) repeat units. ITS variant type-I was close to ITS from V. unguiculata ssp. pubescens, whereas type-II was close to V. unguiculata ssp. tenuis. Transcript analysis clearly demonstrates that in accession NI 1637, rDNA repeat units with only type-II ITS variants are transcriptionally active. Evidence from sequence analysis (of 5.8S, ITS1, and ITS2) and secondary structure analysis (of ITS1 and ITS2) indicates that the type-I ITS variant probably does not belong to the pseudogenic rDNA repeat units. The results from phylogenetic and transcript analysis suggest that the rDNA units with the type-I ITS may have introgressed as a result of hybridization (between ssp. tenuis and ssp. pubescens); however, it has been epigenetically silenced. The results also demonstrate differential evolution of ITS sequence among wild and cultivated forms of V. unguiculata.

Nuclear DNA assay in solving issues related to ancestry of the domesticated diploid safflower (Carthamus tinctorius L.) and the polyploid (Carthamus) taxa, and phylogenetic and genomic relationships in the genus Carthamus L. (Asteraceae)

The multipronged nuclear DNA assay by random amplified polymorphic DNA (RAPD) fingerprinting, ribosomal DNA repeat unit length polymorphism, internal transcribed sequence (ITS) RFLP, and comparative sequence analysis of ITS and external transcribed sequence (ETS) regions of the 29 accessions belonging to 18 Carthamus taxa including five unverified species was undertaken to obtain new information on (1) interrelationships among botanical varieties of cultivated safflower, C. tinctorius, and phylogenetic relationships (2) among the safflower and its close relatives and (3) that of Carthamus species and subspecies. The root tip cells of the 12 accessions contained 24 chromosomes followed by 64, 44 and 20 chromosomes in 9, 6 and 2 accessions, respectively. Barring C. lanatus, the accessions within each taxon had the same zygotic number. The present results strongly support the view that the wild C. palaestinus (2n=24) and the cultivated C. tinctorius (2n=24) are closely related. With few exceptions, all DNA based dendrograms support three lineages within the genus. One lineage is constituted by C. arborescens (2n=24) alone. The present data indicates that because of unique composition of its nuclear genome vis-à-vis other Carthamus taxa, C. arborescens should be placed in a separate subgenus. The two remaining lineages, constituted by the taxa with 2n=24, and the taxa with 2n=20, 2n=44 and 2n=64, respectively should be given the rank of two taxonomic sections in the other subgenus. The present study also demonstrates that none of the present taxa with 2n=24 have contributed to the origin of polyploid taxa. Carthamus boisserii (2n=20) and C. glaucus ssp. anatolicus (2n=20) are more likely to be one of the diploid progenitor of C. lanatus ssp. creticus (2n=64), C. lanatus (2n=44), C. lanatus ssp. lanatus (2n=44) and C. lanatus ssp. montanus (2n=44), and C. lanatus ssp. turkestanicus (2n=64), respectively. Within Lanatus species complex, constituted by C. lanatus, C. lanatus ssp. lanatus, C. lanatus ssp. montanus, C. lanatus ssp. turkestanicus and C. lanatus ssp. creticus, high proportion of autapomorphic characters and low number of synapomorphies in the ITS and ETS sequences suggest a relatively recent diversification of the taxa within the species complex. Carthamus lanatus ssp. creticus (2n=64) and C. lanatus ssp. turkestanicus (2n=64) within the complex deserve species rank. This analysis provided evolutionary relatedness of the five unverified taxa with the known Carthamus taxa.

교잡된 Cordyceps militalis 균주의 RAPD 분석 및 생리활성물질인 cordycepin 함량 측정

KACC에서 분양받은 C. militaris 4균주와 야생에서 채집된 6균주 그리고 교배 육성한 4균주를 대상으로 DNA분석에 의한 분류동정을 시행하였다. 표준균주 10균주와 교잡균주 4균주에서 조사된 전 부위(ITS1, 5.8S rDNA, ITS2)에서 많은 염기서열 차이를 보이고 있으나 같은 종으로 확인된 표준균주들과 수집균주들과 비교한 바 같은 균으로 분류된 것을 확인할 수 있었다. 교잡에 쓰여 졌던 표준 균주에서 교잡된 교잡균주 중 가장 우수한 교잡균주 mat1과 교잡에 사용되었던 표준균주(MPNU10306, MPNU10366)를 선택하여 RAPD 분석을 하였으며, RAPD에서 나타난 각 균주간의 다양한 유전적 변이가 관찰되었다. 이는 새로운 C. militaris종 균주로 판단되어지며, HPLC 분석에서도 교잡에 쓰여 졌던 표준균주보다 교잡한 C. militaris 균주가 cordycepin 생산량이 우수하게 나타났으며, 이는 교잡균주가 cordycepin 함량을 보다 더 높일 수 있음을 의미하고 상업적 가치가 있을 것이라 판단되어지며, 지속적인 연구가 필요할 것으로 생각된다. The C. militaris belongs to entomopathogenic fungi, which have their specific sequences in internal transcribed spacer regions (ITS1 and ITS2) depending on species. In this study, to identify the phylogenetic relationship of the strain hybrided by mating of C. militaris, we compared DNA sequences of ITS regions and 5.8S ribosomal DNA (rDNA) repeat unit of hybrid strain and its parental strains. The result revealed that hybrid strains are C. militaris species. In addition, cordycepins produced by hybrid strains and other strains of C. militaris were analyzed by HPLC with 20mM <TEX>$KH_2PO_4$</TEX> of mobile phase and C-18 columns. The result indicated that the strain hybrided by mating produce higher concentration of phytochemical cordycepin than other C. militaris strains.

Distribution and evolution of variable group-I introns in the small ribosomal subunit of North Atlantic Porphyra (Bangiales, Rhodophyta)

Within the Bangialean red algae the multi-copy small subunit nuclear ribosomal gene (SSU) has been shown to contain group-I introns at positions corresponding to 516 and 1506. Using the polymerase chain reaction (PCR) with primers flanking the 1506 intron we found that this intron was present in some copies of the rDNA in several North Atlantic Porphyra species. However, when the combination of a flanking and internal primer was used to amplify across the intron the percentages of individuals containing the intron increased. The likely explanation for these results was that, when flanking primers were used for PCR, template competition favoured the amplification of the intronless rDNA fragments. A combination of internal and flanking primers used to screen for the 516 intron amplified the intron in every accession of five taxa (Porphyra dioica, P. leucosticta, P. suborbiculata, P. umbilicalis and P. yezoensis) and in most samples of P. amplissima, P. linearis, and P. purpurea. When PCR primers were optimized for the 1506 intron and SSU of P. umbilicalis, the intron was detected in 28/28 samples. Such observations suggest that the 1506 intron is ubiquitous in Porphyra and is present in some, but not all, copies of the ribosomal DNA (rDNA) repeat. Multiple size 1506 introns were detected in individuals of P. umbilicalis. Four of the larger size variants were shown to encode a putative His–Cys box that was associated with mobility of other group-I introns. Our observations suggest that the 1506 group-I intron may still be mobile within the Porphyra lineage.

Comparative genomics and molecular dynamics of DNA repeats in eukaryotes.

Repeated elements can be widely abundant in eukaryotic genomes, composing more than 50% of the human genome, for example. It is possible to classify repeated sequences into two large families, "tandem repeats" and "dispersed repeats." Each of these two families can be itself divided into subfamilies. Dispersed repeats contain transposons, tRNA genes, and gene paralogues, whereas tandem repeats contain gene tandems, ribosomal DNA repeat arrays, and satellite DNA, itself subdivided into satellites, minisatellites, and microsatellites. Remarkably, the molecular mechanisms that create and propagate dispersed and tandem repeats are specific to each class and usually do not overlap. In the present review, we have chosen in the first section to describe the nature and distribution of dispersed and tandem repeats in eukaryotic genomes in the light of complete (or nearly complete) available genome sequences. In the second part, we focus on the molecular mechanisms responsible for the fast evolution of two specific classes of tandem repeats: minisatellites and microsatellites. Given that a growing number of human neurological disorders involve the expansion of a particular class of microsatellites, called trinucleotide repeats, a large part of the recent experimental work on microsatellites has focused on these particular repeats, and thus we also review the current knowledge in this area. Finally, we propose a unified definition for mini- and microsatellites that takes into account their biological properties and try to point out new directions that should be explored in a near future on our road to understanding the genetics of repeated sequences.

The leukemogenic t(8;21) fusion protein AML1-ETO controls rRNA genes and associates with nucleolar-organizing regions at mitotic chromosomes

RUNX1/AML1 is required for definitive hematopoiesis and is frequently targeted by chromosomal translocations in acute myeloid leukemia (AML). The t(8;21)-related AML1-ETO fusion protein blocks differentiation of myeloid progenitors. Here, we show by immunofluorescence microscopy that during interphase, endogenous AML1-ETO localizes to nuclear microenvironments distinct from those containing native RUNX1/AML1 protein. At mitosis, we clearly detect binding of AML1-ETO to nucleolar-organizing regions in AML-derived Kasumi-1 cells and binding of RUNX1/AML1 to the same regions in Jurkat cells. Both RUNX1/AML1 and AML1-ETO occupy ribosomal DNA repeats during interphase, as well as interact with the endogenous RNA Pol I transcription factor UBF1. Promoter cytosine methylation analysis indicates that RUNX1/AML1 binds to rDNA repeats that are more highly CpG methylated than those bound by AML1-ETO. Downregulation by RNA interference reveals that RUNX1/AML1 negatively regulates rDNA transcription, whereas AML1-ETO is a positive regulator in Kasumi-1 cells. Taken together, our findings identify a novel role for the leukemia-related AML1-ETO protein in epigenetic control of cell growth through upregulation of ribosomal gene transcription mediated by RNA Pol I, consistent with the hyper-proliferative phenotype of myeloid cells in AML patients.

Sequence characterization and phylogenetic analysis of the 5S ribosomal DNA in some scallops (Bivalvia: Pectinidae)

This study was designed to characterize the 5S ribosomal DNA repeat unit and to evaluate its phylogenetic informativeness in Mimachlamys varia, Hinnites distortus, Aequipecten opercularis and Pecten maximus, scallops of the bivalve family Pectinidae. The repeat unit was PCR amplified and several products cloned and sequenced. The deduced coding region covered 120 bp, showed 52-53% GC content and an identifiable internal promoter. The spacer region was 313-345 bp in length with 30-40% GC and the ends contained elements showing similarity with upstream and downstream sequences involved in the transcription. The sequences of P. maximus were identical and those of M. varia and H. distortus differed only at 2-3 positions. However, the sequences of A. opercularis showed a percentage of differences of 0.69-9.15%, distinguishing two types of units differing in sequence and length of the spacer region. All scallops displayed an identical gene sequence, except M. varia that differed by one nucleotide substitution, but a highly variable spacer. In the phylogenetic trees the sequences grouped by species with two well supported clades, one containing the sequences of M. varia and H. distortus and the other those of P. maximus and A. opercularis, the latter grouped according to the unit type. The topology obtained was in accordance with scallop evolutionary relationships inferred from previous phylogenetic studies.

Genetic Linkage Map of the Nucleolus Organizer Region in the Soybean

Simple polymorphisms in ribosomal DNA repeats in the nucleolus organizer region (NOR) permitted the development of markers for the genetic mapping of the soybean NOR. The markers map to the top end of soybean linkage group F, one of either telomeric end predicted in the cytogenetic and primary trisomic studies.

Peripheral blood serum from healthy donors contains antibodies against the fragment of transcribed region of ribosomal repeat

Enzyme immunoassay showed that blood serum from healthy donors contains specific high-affinity antibodies (apparent association constant > or = 5 x 10(9) M(-1)) against a fragment of transcribed region of ribosomal DNA repeat of human serum, which are present in a free form or are bound to extracellular DNA. Preheating of the serum at 55 degrees C and high ionic strength (1.5 M NaCl) had no effect on the interaction of antibodies with this fragment. Competitive binding assay showed that these antibodies recognize DNA epitopes, which differ from the epitopes recognized by most anti-DNA antibodies in systemic lupus erythematosus.

Tissue culture triggers chromosome alterations, amplification, and transposition of repeat sequences in Allium fistulosum

Structural alterations in nuclei and chromosomes of cells derived from callus culture of Allium fistulosum have been studied with fluorescent in situ hybridization (FISH) using 5S ribosomal DNA (rDNA), 45S rDNA, and 375-bp repeat probes. A high frequency of chromosome abnormalities was found to be caused by the loss of telomere-located 375-bp repeats, chromosome fusion, and subsequent breakage-fusion-bridge cycles. Products of chromosome fusions and monocentric and regularly shaped chromosomes showed additional 375-bp repeat and 45S rDNA clusters at unusual sites, suggesting dynamic copy-number changes and transposition of these repeats. Southern hybridization revealed no differences in the 375-bp repeat and 45S rDNA repeat array order or the degree of methylation between DNA isolated from leaves or tissue-culture cells. In addition, protruding, spike-like structures positive for 375-bp repeats were identified on the surface of different-sized nuclei. Transmission electron microscopy analysis revealed the accumulation of densely packed chromatin within spike-like structures. Because root calyptra cells showed similar structures, it is likely that heterochromatic spike-like structures are a feature of nondividing cells at the onset of programmed cell death.

Assessment of genetic diversity, and phylogenetic relationships based on ribosomal DNA repeat unit length variation and Internal Transcribed Spacer (ITS) sequences in chickpea (Cicer arietinum) cultivars and its wild species

Repeat unit length variation and internal transcribed spacer (ITS) sequences of nuclear ribosomal DNA were used to assess genetic diversity, and phylogenetic relationships in chickpea (C. arietinum) cultivars, and its related wild species. Total genomic DNAs of 76 accessions of 10 Cicer species, belonging to three sections of the genus, were restricted with seven enzymes and the restriction fragments were hybridized to heterologous ribosomal clones of wheat pTa71 and Vicia faba probes Ver 6-5 and Ver18-6. A single repeat unit length class of 11.4 kb or 10.5 kb was recognized across Cicer accessions with pTa71. The intraspecific variation was negligible in those species where more than one accession was studied, except the four C. judaicum accessions, which were different from the rest. EcoRI and DraI digests gave two and one-two fragments, respectively. All the accessions produced three and three-five bands with BamHI and SacI, respectively. Both the accessions of C. yamashitae differed in their rDNA repeat unit length as well as restriction site variation. Maximum likelihood tree with rDNA RFLP recognized five clades which were more or less congruent with the previous data. Length of ITS-1 region was more variable (235–239 bp) than the ITS-2 region (212–213 bp). Cladistic analysis of ITS data revealed two major clades, clade I consisting of C. arietinum, C. reticulatum and C. echinospermum, and clade II comprised of C. judaicum, C. chorassanicum, C. bijugum and C. cuneatum. C. microphyllum grouped with the above four species. C. pinnatifidum was present as a separate branch. C. yamashitae emerged as the most distinct species.

Highly efficient concerted evolution in the ribosomal DNA repeats: Total rDNA repeat variation revealed by whole-genome shotgun sequence data

Repeat families within genomes are often maintained with similar sequences. Traditionally, this has been explained by concerted evolution, where repeats in an array evolve "in concert" with the same sequence via continual turnover of repeats by recombination. Another form of evolution, birth-and-death evolution, can also explain this pattern, although in this case selection is the critical force maintaining the repeats. The level of intragenomic variation is the key difference between these two forms of evolution. The prohibitive size and repetitive nature of large repeat arrays have made determination of the absolute level of intragenomic repeat variability difficult, thus there is little evidence to support concerted evolution over birth-and-death evolution for many large repeat arrays. Here we use whole-genome shotgun sequence data from the genome projects of five fungal species to reveal absolute levels of sequence variation within the ribosomal RNA gene repeats (rDNA). The level of sequence variation is remarkably low. Furthermore, the polymorphisms that are detected are not functionally constrained and seem to exist beneath the level of selection. These results suggest the rDNA is evolving via concerted evolution. Comparisons with a repeat array undergoing birth-and-death evolution provide a clear contrast in the level of repeat array variation between these two forms of evolution, confirming that the rDNA indeed does evolve via concerted evolution. These low levels of intra-genomic variation are consistent with a model of concerted evolution in which homogenization is very rapid and efficiently maintains highly similar repeat arrays.

Characterisation of the subtelomeric regions of Giardia lamblia genome isolate WBC6

Giardia trophozoites are polyploid and have five chromosomes. The chromosome homologues demonstrate considerable size heterogeneity due to variation in the subtelomeric regions. We used clones from the genome project with telomeric sequence at one end to identify six subtelomeric regions in addition to previously identified subtelomeric regions, to study the telomeric arrangement of the chromosomes. The subtelomeric regions included two retroposons, one retroposon pseudogene, and two vsp genes, in addition to the previously identified subtelomeric regions that include ribosomal DNA repeats. The presence of vsp genes in a subtelomeric region suggests that telomeric rearrangements may contribute to the generation of vsp diversity. These studies of the subtelomeric regions of Giardia may contribute to our understanding of the factors that maintain stability, while allowing diversity in chromosome structure.

Cloning and sequencing of the complicated rDNA gene family of Bos taurus

The rDNA genes coding for ribosomal RNA (rRNA) in animals are repeat sequences with high GC content and complicated structure. Based on the sequences of human ribosomal DNA repeat unit and transcription unit and the long and accurate PCR method with LA Taq DNA polymerase and GC buffer, we were able to amplify the complicated repeat sequences of rDNA genes in Bos taurus. Three rDNA genes and 2 internal transcribed spacer (ITS) fragments were cloned and confirmed by sequencing. The conditions for the cloning of complicated DNA sequences such as special rules of primer design, improvement of the reaction system, selection of DNA polymerase and adjustment of cycle parameters were discussed. &amp;nbsp;

Novel Nucleomorph Genome Architecture in the Cryptomonad Genus Hemiselmis

Cryptomonads are ubiquitous aquatic unicellular eukaryotes that acquired photosynthesis through the uptake and retention of a red algal endosymbiont. The nuclear genome of the red alga persists in a highly reduced form termed a nucleomorph. The nucleomorph genome of the model cryptomonad Guillardia theta has been completely sequenced and is a mere 551 kilobases (kb) in size, spread over three chromosomes. The presence of three chromosomes appears to be a universal characteristic of nucleomorph genomes in cryptomonad algae as well as in the chlorarachniophytes, an unrelated algal lineage with a nucleomorph and plastid genome derived from a green algal endosymbiont. Another feature of nucleomorph genomes in all cryptomonads and chlorarachniophytes examined thus far is the presence of subtelomeric ribosomal DNA (rDNA) repeats at the ends of each chromosome. Here we describe the first exception to this canonical nucleomorph genome architecture in the cryptomonad Hemiselmis rufescens CCMP644. Using pulsed-field gel electrophoresis (PFGE), we estimate the size of the H. rufescens nucleomorph genome to be approximately 580 kb, slightly larger than the G. theta genome. Unlike the situation in G. theta and all other known cryptomonads, sub-telomeric repeats of the rDNA cistron appear to be absent on both ends of the second largest chromosome in H. rufescens and two other members of this genus. Southern hybridizations using a variety of nucleomorph protein gene probes against PFGE-separated H. rufescens chromosomes indicate that recombination has been a major factor in shaping the karyotype and genomic structure of cryptomonad nucleomorphs.

Inhibition of homologous recombination by a cohesin-associated clamp complex recruited to the rDNA recombination enhancer

Silencing within the yeast ribosomal DNA (rDNA) repeats protects the integrity of this highly repetitive array by inhibiting hyperrecombination and repressing transcription from foreign promoters. Using affinity purification combined with highly sensitive mixture mass spectrometry, we have analyzed the protein interaction network involved in suppressing homologous recombination within the rDNA locus. We show that the Net1 and Sir2 subunits of the RENT (regulator of nucleolar silencing and telophase exit) silencing complex, and Fob1, which recruits RENT to the nontranscribed spacer I (NTS1) region of rDNA, are physically associated with Tof2. In addition to RENT components and Fob1, Tof2 copurified with a two-subunit complex composed of Lrs4 and Csm1. Tof2, Lrs4, and Csm1 are recruited to the NTS1 region by Fob1 and are specifically required for silencing at this rDNA region. Moreover, Lrs4 and Csm1 act synergistically with Sir2 to suppress unequal crossover at the rDNA and are released from the nucleolus during anaphase. Together with previous observations showing that Csm1 physically associates with cohesin, these findings suggest a possible model in which RENT, Tof2, and Lrs4/Csm1 physically clamp rDNA to the cohesin ring, thereby restricting the movement of rDNA sister chromatids relative to each other to inhibit unequal exchange.

RNA polymerase I-specific subunit CAST/hPAF49 has a role in the activation of transcription by upstream binding factor.

Eukaryotic RNA polymerases are large complexes, 12 subunits of which are structurally or functionally homologous across the three polymerase classes. Each class has a set of specific subunits, likely targets of their cognate transcription factors. We have identified and characterized a human RNA polymerase I (Pol I)-specific subunit, previously identified as ASE-1 (antisense of ERCC1) and as CD3epsilon-associated signal transducer (CAST), and here termed CAST or human Pol I-associated factor of 49 kDa (hPAF49), after mouse orthologue PAF49. We provide evidence for growth-regulated Tyr phosphorylation of CAST/hPAF49, specifically in initiation-competent Pol Ibeta complexes in HeLa cells, at a conserved residue also known to be important for signaling during T-cell activation. CAST/hPAF49 can interact with activator upstream binding factor (UBF) and, weakly, with selectivity factor 1 (SL1) at the rDNA (ribosomal DNA repeat sequence encoding the 18S, 5.8S, and 28S rRNA genes) promoter. CAST/hPAF49-specific antibodies and excess CAST/hPAF49 protein, which have no effect on basal Pol I transcription, inhibit UBF-activated transcription following functional SL1-Pol I-rDNA complex assembly and disrupt the interaction of UBF with CAST/hPAF49, suggesting that interaction of this Pol I-specific subunit with UBF is crucial for activation. Drawing on parallels between mammalian and Saccharomyces cerevisiae Pol I transcription machineries, we advance one model for CAST/hPAF49 function in which the network of interactions of Pol I-specific subunits with UBF facilitates conformational changes of the polymerase, leading to stabilization of the Pol I-template complex and, thereby, activation of transcription.

Phylogenetic relationships among Puccinia hemerocallidis, P. funkiae, and P. patriniae (Uredinales) inferred from ITS sequence data

Puccinia hemerocallidis and P. funkiae resemble each other morphologically; however, they are biologically and taxonomically distinct, with telia of the former being restricted to species of Hemerocallis and the latter to Hosta species. However, both fungi share a macrocyclic and heteroecious life cycle with Patrinia villosa as the spermogonial and aecial host. An additional microcyclic rust fungus, P. patriniae, is also known on P. villosa. This microcyclic fungus is similar to the two macrocyclic fungi in its telial structure and teliospore morphology. These similarities in morphology and host relationships suggest the three fungi may also share a close evolutionary relationship. To determine the phylogenetic relationships of the three species, a portion of the nuclear ribosomal DNA repeat encoding the ITS and 5.8S subunit regions was amplified by PCR, sequenced, and analyzed. The resulting phylogenetic trees showed that P. hemerocallidis and P. funkiae share a recent common ancestor and that P. patriniae is closely allied with P. hemerocallidis. The results suggest a possible evolutionary derivation of microcyclic P. patriniae from macrocyclic heteroecious P. hemerocallidis, which fits the evolutionary interpretation of correlated species known as Tranzschel's law.

Metschnikowia noctiluminum sp. nov., Metschnikowia corniflorae sp. nov., and Candida chrysomelidarum sp. nov., isolated from green lacewings and beetles

Fourteen yeast isolates belonging to the Metschnikowia clade were isolated from the digestive tracts of lacewings ( Neuroptera: Chrysopidae), soldier beetles and leaf beetles ( Coleoptera: Cantharidae and Chrysomelidae), and a caddisfly ( Trichoptera: Hydropsychidae). The insect hosts were associated with sugary substances of plants, a typical habitat for yeasts in this clade. Based on DNA sequence comparisons and phenetic characters, the yeasts were identified as Candida picachoensis, Candida pimensis, and four undescribed taxa. Among the undescribed taxa, three yeasts were distinguished from one another and from other described taxa by nucleotide differences in the ribosomal DNA repeat, which were sufficient to consider them as new species. Two of the novel yeast species are described as Metschnikowia noctiluminum (NRRL Y-27753 T) and M. corniflorae spp. nov. (NRRL Y-27750 T) based in part on production of needle-shaped ascospores, which are found in most Metschnikowia species. Sexual reproduction was not observed in the third new yeast, Candida chrysomelidarum sp. nov. (NRRL Y-27749 T). A fourth isolate, NRRL Y-27752, was not significantly distinct from Metschnikowia viticola and Candida kofuensis to be described as a new species. Phylogenetic analysis of the D1/D2 loop sequences placed M. noctiluminum within the M. viticola clade, while C. chrysomelidarum was a sister taxon of Candida rancensis. Metschnikowia corniflorae was phylogenetically distinct from other new species and fell outside of the large-spored Metschnikowia group.

Aurapex penicillata gen. sp. nov. from native Miconia theaezans and Tibouchina spp. in Colombia

Conidiomata of a fungus resembling Chrysoporthe cubensis, a serious canker pathogen of Eucalyptus spp. (Myrtaceae, Myrtales) in tropical and subtropical parts of the world, was found on Eucalyptus grandis in Colombia. Fruiting structures of the fungus could be distinguished from those of C. cubensis by their distinctly orange conidiomatal necks. This fungus also was found on several plant species native to Colombia including Tibouchina urvilleana, T. lepidota and Miconia theaezans (Melastomataceae, Myrtales). Morphological comparisons, as well as those based on sequences of the ITS1/ITS2 region of the ribosomal DNA repeat and the β-tubulin gene, were used to characterize this fungus. Its pathogenicity was assessed on various plants from which it has been collected, either in field or greenhouse trials. Phylogenetic analyses showed that isolates reside in a clade distinct from the four clades accommodating Chrysoporthe, Cryphonectria, Endothia and Rostraureum. Members of this clade are distinguished by the presence of orange conidiomatal necks with black bases and a unique internal stromatal structure. No teleomorph has been found for this fungus, for which we have provided the name Aurapex penicillata gen. sp. nov. A. penicillata produced only small lesions after inoculation on young T. urvilleana, M. theaezans and E. grandis trees and appears not to be a serious pathogen.