Intercellular communication is required for trap formation in the nematode-trapping fungus Duddingtonia flagrans

Nematode-trapping fungi (NTF) are a large and diverse group of fungi, which may switch from a saprotrophic to a predatory lifestyle if nematodes are present. Different fungi have developed different trapping devices, ranging from adhesive cells to constricting rings. After trapping, fungal hyphae penetrate the worm, secrete lytic enzymes and form a hyphal network inside the body. We sequenced the genome of Duddingtonia flagrans, a biotechnologically important NTF used to control nematode populations in fields. The 36.64 Mb genome encodes 9,927 putative proteins, among which are more than 638 predicted secreted proteins. Most secreted proteins are lytic enzymes, but more than 200 were classified as small secreted proteins (< 300 amino acids). 117 putative effector proteins were predicted, suggesting interkingdom communication during the colonization. As a first step to analyze the function of such proteins or other phenomena at the molecular level, we developed a transformation system, established the fluorescent proteins GFP and mCherry, adapted an assay to monitor protein secretion, and established gene-deletion protocols using homologous recombination or CRISPR/Cas9. One putative virulence effector protein, PefB, was transcriptionally induced during the interaction. We show that the mature protein is able to be imported into nuclei in Caenorhabditis elegans cells. In addition, we studied trap formation and show that cell-to-cell communication is required for ring closure. The availability of the genome sequence and the establishment of many molecular tools will open new avenues to studying this biotechnologically relevant nematode-trapping fungus

Relative abundance of integronase genes (A) and plasmids (B).

<p>Relative abundance of integronase genes (A) and plasmids (B).</p

Influence of a Non-Hospital Medical Care Facility on Antimicrobial Resistance in Wastewater

<div><p>The global widespread use of antimicrobials and accompanying increase in resistant bacterial strains is of major public health concern. Wastewater systems and wastewater treatment plants are considered a niche for antibiotic resistance genes (ARGs), with diverse microbial communities facilitating ARG transfer via mobile genetic element (MGE). In contrast to hospital sewage, wastewater from other health care facilities is still poorly investigated. At the instance of a nursing home located in south-west Germany, in the present study, shotgun metagenomics was used to investigate the impact on wastewater of samples collected up- and down-stream in different seasons. Microbial composition, ARGs and MGEs were analyzed using different annotation approaches with various databases, including Antibiotic Resistance Ontologies (ARO), integrons and plasmids. Our analysis identified seasonal differences in microbial communities and abundance of ARG and MGE between samples from different seasons. However, no obvious differences were detected between up- and downstream samples. The results suggest that, in contrast to hospitals, sewage from the nursing home does not have a major impact on ARG or MGE in wastewater, presumably due to much less intense antimicrobial usage. Possible limitations of metagenomic studies using high-throughput sequencing for detection of genes that seemingly confer antibiotic resistance are discussed.</p></div

Characterization and Phylogenetic Analysis of the Mitochondrial Genome of <i>Glarea lozoyensis</i> Indicates High Diversity within the Order Helotiales

<div><p>Background</p><p><i>Glarea lozoyensis</i> is a filamentous fungus used for the industrial production of non-ribosomal peptide pneumocandin B₀. In the scope of a whole genome sequencing the complete mitochondrial genome of the fungus has been assembled and annotated. It is the first one of the large polyphyletic Helotiaceae family. A phylogenetic analysis was performed based on conserved proteins of the oxidative phosphorylation system in mitochondrial genomes.</p><p>Results</p><p>The total size of the mitochondrial genome is 45,038 bp. It contains the expected 14 genes coding for proteins related to oxidative phosphorylation,two rRNA genes, six hypothetical proteins, three intronic genes of which two are homing endonucleases and a ribosomal protein rps3. Additionally there is a set of 33 tRNA genes. All genes are located on the same strand. Phylogenetic analyses based on concatenated mitochondrial protein sequences confirmed that <i>G. lozoyensis</i> belongs to the order of Helotiales and that it is most closely related to <i>Phialocephala subalpina</i>. However, a comparison with the three other mitochondrial genomes known from Helotialean species revealed remarkable differences in size, gene content and sequence. Moreover, it was found that the gene order found in <i>P. subalpina</i> and <i>Sclerotinia sclerotiorum</i> is not conserved in <i>G. lozoyensis.</i></p><p>Conclusion</p><p>The arrangement of genes and other differences found between the mitochondrial genome of <i>G. lozoyensis</i> and those of other Helotiales indicates a broad genetic diversity within this large order. Further mitochondrial genomes are required in order to determine whether there is a continuous transition between the different forms of mitochondri<i>a</i>l genomes or <i>G. lozoyensis</i> belongs to a distinct subgroup within Helotiales.</p></div

Abundance of the 12 most frequent ARGs types with the highest blast hits number (Relative abundance in any sample ≥10 ppm).

<p>Abundance of the 12 most frequent ARGs types with the highest blast hits number (Relative abundance in any sample ≥10 ppm).</p

Effect of the ferredoxin electron donor on sunflower (Helianthus annuus) desaturases

Ferredoxins are proteins that participate in photosynthesis and in other processes that require reducing equivalents, such as the reduction of nitrogen or fatty acid desaturation. Two classes of ferredoxins have been described in plants: light-regulated photosynthetic ferredoxins and heterotrophic ferredoxins whose activity is not influenced by light. Genes encoding the two forms of ferredoxin have been cloned and characterized in developing sunflower cotyledons. Here, these genes were overexpressed in Escherichia coli and they were purified by ion exchange and size exclusion chromatography to study their capacity to supply electrons to two different sunflower desaturases: soluble stearoyl-ACP desaturase from sunflower cotyledons, and membrane bound desaturase FAD7 expressed in yeast. In both cases photosynthetic ferredoxin was the form that promoted the strongest desaturase activity.This work was supported by MEC and FEDER project AGL 2005-00100.Peer reviewe

Selected fungus species with published mt genomes.

1<p><i>Botrytis cinerea</i> Sequencing Project.</p>2<p><i>Sclerotinia sclerotiorum</i> Sequencing Project.</p>3<p><i>Neurospora crassa</i> Sequencing Project, (<a href="http://www.broadinstitute.org/" target="_blank">http://www.broadinstitute.org/</a>).</p

Codon usage of protein-coding genes in <i>G. lozoyensis</i> mt genome.

<p>The overall percentage of codon usage in the protein coding genes atp6, atp8, atp9, cob, cox1, cox2, cox3, nad1, nad2, nad3, nad4, nad4L, nad5, nad6, orf 1, orf 2, orf 3, orf 4, orf 5, orf 6, rps3, intron protein (IA), intron protein (IB) and intron protein (IE) is depicted.</p