Introduction to the \u3cem\u3eToxins\u3c/em\u3e Special Issue on Ergot Alkaloids

Ergot alkaloids are among the most relevant natural products in the history of toxins and pharmaceuticals. Until the late 20th century, human and livestock exposure to ergot alkaloids was primarily through ingestion of “ergots,” which are spur-shaped or seed-like resting structures (sclerotia) of ergot fungi, the Claviceps species. Because ergots have similar density to grains, traditional threshing techniques generally failed to remove them, and outbreaks of ergot typically led to mass poisonings. [...

A study of exothermic chemical reaction in a porous catalyst wedge

Temperature and composition were measured at various locations in a system where ethylene is hydrogenated to ethane on a 1/4 inch porous catalyst wedge made of nickel supported on alumina. \u27When the wedge of catalyst was bathed in hydrogen, experimental results indicated a high temperature rise from the feed temperature to the catalyst which is implied by the diffusion controlled regime for solid catalyzed, highly exothermic reactions. This high activity was reversibly reduced by bathing the catalyst wedge in nitrogen for extended time periods (greater than 24 hours). This nitrogen soaking changed the experimental conditions from those of diffusion controlled kinetics to those typically exhibited by systems in the kinetic regime. The experimental system was simulated numerically for a variety of boundary conditions using reasonable assumptions and physical property data for this reaction system. Heat and mass transfer coefficients were allowed to vary along the wedge according to boundary layer theory results. For the internal wedge temperatures predicted by the model to agree within 1% of those measured experimentally, it was necessary to consider finite heat transfer at the stagnation point which is contrary to the classical boundary layer theory commonly applied to flat plate and wedge flows. It was also necessary to allow modest (~6% or less) heat loss from the back edge of the wedge --Abstract, page ii

Endophyte Ergot Alkaloid Synthetic Compounds, Compounds Which Encode Therefor and Related Methods

The present invention provides, inter alia, dmaW nucleic acid sequences and the proteins for which they encode. Also provided are methods for the utilization of knockout mutants of the sequences which are useful for engineering ergot alkaloid-deficient fungal symbionts (endophytes) of plants. Other methods and materials related to these sequences are also provided

Statistical Phylogenetic Tree Analysis Using Differences of Means

We propose a statistical method to test whether two phylogenetic trees with given alignments are significantly incongruent. Our method compares the two distributions of phylogenetic trees given by the input alignments, instead of comparing point estimations of trees. This statistical approach can be applied to gene tree analysis for example, detecting unusual events in genome evolution such as horizontal gene transfer and reshuffling. Our method uses difference of means to compare two distributions of trees, after embedding trees in a vector space. Bootstrapping alignment columns can then be applied to obtain p-values. To compute distances between means, we employ a "kernel trick" which speeds up distance calculations when trees are embedded in a high-dimensional feature space, e.g. splits or quartets feature space. In this pilot study, first we test our statistical method's ability to distinguish between sets of gene trees generated under coalescence models with species trees of varying dissimilarity. We follow our simulation results with applications to various data sets of gophers and lice, grasses and their endophytes, and different fungal genes from the same genome. A companion toolkit, {\tt Phylotree}, is provided to facilitate computational experiments.Comment: 17 pages, 6 figure

Non-Transgenic CRISPR-Mediated Knockout of Entire Ergot Alkaloid Gene Clusters in Slow-Growing Asexual Polyploid Fungi

The Epichloë species of fungi include seed-borne symbionts (endophytes) of cool-season grasses that enhance plant fitness, although some also produce alkaloids that are toxic to livestock. Selected or mutated toxin-free endophytes can be introduced into forage cultivars for improved livestock performance. Long-read genome sequencing revealed clusters of ergot alkaloid biosynthesis (EAS) genes in Epichloë coenophiala strain e19 from tall fescue (Lolium arundinaceum) and Epichloë hybrida Lp1 from perennial ryegrass (Lolium perenne). The two homeologous clusters in E. coenophiala—a triploid hybrid species—were 196 kb (EAS1) and 75 kb (EAS2), and the E. hybrida EAS cluster was 83 kb. As a CRISPR-based approach to target these clusters, the fungi were transformed with ribonucleoprotein (RNP) complexes of modified Cas9 nuclease (Cas9-2NLS) and pairs of single guide RNAs (sgRNAs), plus a transiently selected plasmid. In E. coenophiala, the procedure generated deletions of EAS1 and EAS2 separately, as well as both clusters simultaneously. The technique also gave deletions of the EAS cluster in E. hybrida and of individual alkaloid biosynthesis genes (dmaW and lolC) that had previously proved difficult to delete in E. coenophiala. Thus, this facile CRISPR RNP approach readily generates non-transgenic endophytes without toxin genes for use in research and forage cultivar improvement

Experimenting with database segmentation size vs time performance for mpiBLAST on an IBM HS21 blade cluster

Large-scale genomic projects such as the Epichloë festucae Genome Project require regular use of bioinformatic tools. When using BLAST in conjunction with larger databases, processing complex sequences often uses substantial computation time. Parallelization is considered a standard method of curbing extensive computing requirements and parallel implementations of BLAST, such as mpiBLAST, are freely available

Phylogenetic Analyses Reveal Monophyletic Origin of the Ergot Alkaloid Gene \u3cem\u3edmaW\u3c/em\u3e in Fungi

Ergot alkaloids are indole-derived mycotoxins that are important in agriculture and medicine. Ergot alkaloids are produced by a few representatives of two distantly related fungal lineages, the Clavicipitaceae and the Trichocomaceae. Comparison of the ergot alkaloid gene clusters from these two lineages revealed differences in the relative positions and orientations of several genes. The question arose: is ergot alkaloid biosynthetic capability from a common origin? We used a molecular phylogenetic approach to gain insights into the evolution of ergot alkaloid biosynthesis. The 4-γ,γ-dimethylallyltryptophan synthase gene, dmaW, encodes the first step in the pathway. Amino acid sequences deduced from dmaW and homologs were submitted to phylogenetic analysis, and the results indicated that dmaW of Aspergillus fumigatus (mitosporic Trichocomaceae) has the same origin as corresponding genes from clavicipitaceous fungi. Relationships of authentic dmaW genes suggest that they originated from multiple gene duplications with subsequent losses of original or duplicate versions in some lineages