Gene discovery in EST sequences from the wheat leaf rust fungus Puccinia triticina sexual spores, asexual spores and haustoria, compared to other rust and corn smut fungi

© 2011 Xu et al.; licensee BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.DOI: 10.1186/1471-2164-12-161Background.Rust fungi are biotrophic basidiomycete plant pathogens that cause major diseases on plants and trees world-wide, affecting agriculture and forestry. Their biotrophic nature precludes many established molecular genetic manipulations and lines of research. The generation of genomic resources for these microbes is leading to novel insights into biology such as interactions with the hosts and guiding directions for breakthrough research in plant pathology. Results. To support gene discovery and gene model verification in the genome of the wheat leaf rust fungus, Puccinia triticina (Pt), we have generated Expressed Sequence Tags (ESTs) by sampling several life cycle stages. We focused on several spore stages and isolated haustorial structures from infected wheat, generating 17,684 ESTs. We produced sequences from both the sexual (pycniospores, aeciospores and teliospores) and asexual (germinated urediniospores) stages of the life cycle. From pycniospores and aeciospores, produced by infecting the alternate host, meadow rue (Thalictrum speciosissimum), 4,869 and 1,292 reads were generated, respectively. We generated 3,703 ESTs from teliospores produced on the senescent primary wheat host. Finally, we generated 6,817 reads from haustoria isolated from infected wheat as well as 1,003 sequences from germinated urediniospores. Along with 25,558 previously generated ESTs, we compiled a database of 13,328 non-redundant sequences (4,506 singlets and 8,822 contigs). Fungal genes were predicted using the EST version of the self-training GeneMarkS algorithm. To refine the EST database, we compared EST sequences by BLASTN to a set of 454 pyrosequencing-generated contigs and Sanger BAC-end sequences derived both from the Pt genome, and to ESTs and genome reads from wheat. A collection of 6,308 fungal genes was identified and compared to sequences of the cereal rusts, Puccinia graminis f. sp. tritici (Pgt) and stripe rust, P. striiformis f. sp. tritici (Pst), and poplar leaf rust Melampsora species, and the corn smut fungus, Ustilago maydis (Um). While extensive homologies were found, many genes appeared novel and species-specific; over 40% of genes did not match any known sequence in existing databases. Focusing on spore stages, direct comparison to Um identified potential functional homologs, possibly allowing heterologous functional analysis in that model fungus. Many potentially secreted protein genes were identified by similarity searches against genes and proteins of Pgt and Melampsora spp., revealing apparent orthologs. Conclusions. The current set of Pt unigenes contributes to gene discovery in this major cereal pathogen and will be invaluable for gene model verification in the genome sequence

Accurate Inference of Subtle Population Structure (and Other Genetic Discontinuities) Using Principal Coordinates

Accurate inference of genetic discontinuities between populations is an essential component of intraspecific biodiversity and evolution studies, as well as associative genetics. The most widely-used methods to infer population structure are model-based, Bayesian MCMC procedures that minimize Hardy-Weinberg and linkage disequilibrium within subpopulations. These methods are useful, but suffer from large computational requirements and a dependence on modeling assumptions that may not be met in real data sets. Here we describe the development of a new approach, PCO-MC, which couples principal coordinate analysis to a clustering procedure for the inference of population structure from multilocus genotype data.PCO-MC uses data from all principal coordinate axes simultaneously to calculate a multidimensional "density landscape", from which the number of subpopulations, and the membership within subpopulations, is determined using a valley-seeking algorithm. Using extensive simulations, we show that this approach outperforms a Bayesian MCMC procedure when many loci (e.g. 100) are sampled, but that the Bayesian procedure is marginally superior with few loci (e.g. 10). When presented with sufficient data, PCO-MC accurately delineated subpopulations with population F(st) values as low as 0.03 (G'(st)>0.2), whereas the limit of resolution of the Bayesian approach was F(st) = 0.05 (G'(st)>0.35).We draw a distinction between population structure inference for describing biodiversity as opposed to Type I error control in associative genetics. We suggest that discrete assignments, like those produced by PCO-MC, are appropriate for circumscribing units of biodiversity whereas expression of population structure as a continuous variable is more useful for case-control correction in structured association studies

Evidence for maintenance of sex determinants but not of sexual stages in red yeasts, a group of early diverged basidiomycetes

<p>Abstract</p> <p>Background</p> <p>The red yeasts are an early diverged group of basidiomycetes comprising sexual and asexual species. Sexuality is based on two compatible mating types and sexual identity is determined by <it>MAT </it>loci that encode homeodomain transcription factors, peptide pheromones and their receptors. The objective of the present study was to investigate the presence and integrity of <it>MAT </it>genes throughout the phylogenetic diversity of red yeasts belonging to the order Sporidiobolales.</p> <p>Results</p> <p>We surveyed 18 sexual heterothallic and self-fertile species and 16 asexual species. Functional pheromone receptor homologues (<it>STE3.A1 </it>and <it>STE3.A2</it>) were found in multiple isolates of most of the sexual and asexual species. For each of the two mating types, sequence comparisons with whole-genome data indicated that synteny tended to be conserved along the pheromone receptor region. For the homeodomain transcription factor, likelihood methods suggested that diversifying selection acting on the self/non-self recognition region promotes diversity in sexual species, while rapid evolution seems to be due to relaxed selection in asexual strains.</p> <p>Conclusions</p> <p>The majority of both sexual and asexual species of red yeasts have functional pheromone receptors and homeodomain homologues. This and the frequent existence of asexual strains within sexual species, makes the separation between sexual and asexual species imprecise. Events of loss of sexuality seem to be recent and frequent, but not uniformly distributed within the Sporidiobolales. Loss of sex could promote speciation by fostering the emergence of asexual lineages from an ancestral sexual stock, but does not seem to contribute to the generation of exclusively asexual lineages that persist for a long time.</p

Contrasted Patterns of Molecular Evolution in Dominant and Recessive Self-Incompatibility Haplotypes in Arabidopsis

Self-incompatibility has been considered by geneticists a model system for reproductive biology and balancing selection, but our understanding of the genetic basis and evolution of this molecular lock-and-key system has remained limited by the extreme level of sequence divergence among haplotypes, resulting in a lack of appropriate genomic sequences. In this study, we report and analyze the full sequence of eleven distinct haplotypes of the self-incompatibility locus (S-locus) in two closely related Arabidopsis species, obtained from individual BAC libraries. We use this extensive dataset to highlight sharply contrasted patterns of molecular evolution of each of the two genes controlling self-incompatibility themselves, as well as of the genomic region surrounding them. We find strong collinearity of the flanking regions among haplotypes on each side of the S-locus together with high levels of sequence similarity. In contrast, the S-locus region itself shows spectacularly deep gene genealogies, high variability in size and gene organization, as well as complete absence of sequence similarity in intergenic sequences and striking accumulation of transposable elements. Of particular interest, we demonstrate that dominant and recessive S-haplotypes experience sharply contrasted patterns of molecular evolution. Indeed, dominant haplotypes exhibit larger size and a much higher density of transposable elements, being matched only by that in the centromere. Overall, these properties highlight that the S-locus presents many striking similarities with other regions involved in the determination of mating-types, such as sex chromosomes in animals or in plants, or the mating-type locus in fungi and green algae

Urinary excretion of 2-methyl-2,3-butanediol and 2,3-pentanediol in patients with disorders of propionate and methylmalonate metabolism

Urine samples from patients with propionic acidemia and from a patient with methylmalonic acidemia contained unknown non-acidic metabolites by gas chromatography/mass spectrometry after ethyl acetate extraction. It could be demonstrated by mass spectrometric studies and by synthesis of reference compounds that the major metabolite was 2-methyl-2,3-butanediol, while smaller amounts of 2,3-pentanediol were also present. These diols were present in abnormal amounts in these patients during attacks of metabolic decompensation

Using transcription of six Puccinia triticina races to identify the effective secretome during infection of wheat

Citation: Bruce, M., . . . & Fellers, J. (2014). Using transcription of six Puccinia triticina races to identify the effective secretome during infection of wheat. Frontiers in Plant Science, 4(1), 520. https://doi.org/10.3389/fpls.2013.00520Wheat leaf rust, caused by the basidiomycete Puccinia triticina, can cause yield losses of up to 20% in wheat producing regions. During infection, the fungus forms haustoria that secrete proteins into the plant cell and effect changes in plant transcription, metabolism, and defense. It is hypothesized that new races emerge as a result of overcoming plant resistance via changes in the secreted effector proteins. To understand gene expression during infection and find genetic differences associated with races, RNA from wheat leaves infected with six different rust races, at 6 days post inoculation, was sequenced using Illumina. As P. triticina is an obligate biotroph, RNA from both the host and fungi were present and separated by alignment to the P. triticina genome and a wheat EST reference. A total of 222,571 rust contigs were assembled from 165 million reads. An examination of the resulting contigs revealed 532 predicted secreted proteins among the transcripts. Of these, 456 were found in all races. Fifteen genes were found with amino acid changes, corresponding to putative avirulence effectors potentially recognized by 11 different leaf rust resistance (Lr) genes. Twelve of the potential avirulence effectors have no homology to known genes. One gene had significant similarity to cerato-platanin, a known fungal elicitor, and another showed similarity to fungal tyrosinase, an enzyme involved in melanin synthesis. Temporal expression profiles were developed for these genes by qRT-PCR and show that the genes expression patterns were consistent between races from infection initiation to just prior to spore eruption