Posts Tagged ‘pathogen’

Genome of the Honey Mushroom Unearthed

May 23rd, 2013

Collins C, Keane TM, Turner DJ, O’Keeffe G, Fitzpatrick DA, Doyle S (2013) Genomic and Proteomic Dissection of the Ubiquitous Plant Pathogen, Armillaria mellea: Towards a New Infection Model System. J Proteome Research, DOI: 10.1021/pr301131t

Abstract

Armillaria mellea is a major plant pathogen. Yet, no large-scale ‘-omic’ data are available to enable new studies, and limited experimental models are available to investigate basidiomycete pathogenicity. Here we reveal that the A. mellea genome comprises 58.35 Mb, contains 14,473 gene models, of average length 1575 bp (4.72 introns/gene). Tandem mass spectrometry identified 921 mycelial (n = 629 unique) and secreted (n = 183 unique) proteins. Almost 100 mycelial proteins were either species-specific or previously unidentified at the protein level. A number of proteins (n = 111) were detected in both mycelia and culture supernatant extracts. Signal sequence occurrence was fourfold greater for secreted (50.2%) compared to mycelial (12%) proteins. Analyses revealed a rich reservoir of carbohydrate degrading enzymes, laccases and lignin peroxidases in the A. mellea proteome, reminiscent of both basidiomycete and ascomycete glycodegradative arsenals. We discovered that A. mellea exhibits a specific killing effect against Candida albicans, during co-culture. Proteomic investigation of this interaction revealed the unique expression of defensive and potentially offensive A. mellea proteins (n = 30). Overall, our data reveal new insights into the origin of basidiomycete virulence and we present a new model system for further studies aimed at deciphering fungal pathogenic mechanisms.]

Photo: Fruiting body of Armillaria mellea © F Martin

Genomics of Fungal Drug Producers

March 2nd, 2013

In a breakthrough paper, Schardl’s group and collaborators have published 15 genomes of diverse species of Clavicipitaceae plant endophytes and parasites in the last issue of PloS Genetics. The Clavicipitaceae (PezizomycotinaSordariomycetes, Hypocreales) includes “ergot” fungi that parasitize ears of cereals and produce  the toxic ergoline derivatives; ergot fungi have historically caused epidemics of gangrenous poisonings, the ergotism, also known as the Saint Anthony’s Fire. The ascomycetous family also includes plant endophytic symbionts that produce several psychoactive and bioprotective alkaloids. The family includes grass symbionts in the epichloae clade (Epichloë and Neotyphodium species), which are extraordinarily diverse both in their host interactions and in their alkaloid profiles. They synthesize alkaloids with chemical similarities to biogenic amines that deter insects, livestock, and wildlife from feeding on the fungus or plant. Thanks to this chemical warfare, Epichloae protect their hosts from cattle grazing. The lysergic acid diethylamide (LSD), a semisynthetic ergot alkaloid originally developed as an antidepressant, is the most potent known hallucinogen.

In this study, they sequenced the genomes of 10 epichloae, three ergot fungi (Claviceps species), a morning-glory symbiont (Periglandula ipomoeae), and the bamboo witch’s broom pathogen (Aciculosporium take), profiled the alkaloids in these species and compared the gene clusters for four classes of alkaloids. The genomes were primarily sequenced by shotgun 454 pyrosequencing, but paired-end and mate-pair reads were used to scaffold several assemblies. Size of the assembled genome among the sequenced strains varied 2-fold from 29.2 to 58.7 Mb, with wide ranges even within the genera Claviceps (31–52.3 Mb) and Epichloë (29.2–49.3 Mb). This genome size variation is mainly resulting from the abundance of repeated elements, which ranged from 4.7 to 56.9%. Annotated genome sequences have been posted at www.endophyte.uky.edu.

In the epichloae, the clusters of genes coding for enzymes of alkaloid biosynthesis contain very large blocks of repetitive elements which promote gene losses, mutations, and even the evolution of new genes. Two striking features emerged from the detailed analysis of alkaloid biosynthesis gene clusters. Firstly, in most alkaloid loci in most species, the periphery of each cluster was enriched in genes that by virtue of their presence, absence, or sequence variations determined the diversity of alkaloids within the respective chemical class. Second, alkaloid gene loci of the epichloae had extraordinarily large and pervasive blocks of AT- rich repeats derived from retroelements, DNA transposons, and MITEs. This finding suggests that these plant-interacting fungi are under selection for alkaloid diversification.

In their conclusions, the authors suggest that this selection of chemotypes is related to the variable life histories of the epichloae, their protective roles as symbionts, and their associations with the ecologically diverse cool-season grasses.

Schardl CL, Young CA, Hesse U, Amyotte SG, Andreeva K, et al. (2013) Plant-Symbiotic Fungi as Chemical Engineers: Multi-Genome Analysis of the Clavicipitaceae Reveals Dynamics of Alkaloid Loci. PLoS Genet 9(2): e1003323. doi:10.1371/journal.pgen.1003323

Image: Claviceps purpurea -Franz Eugen Köhler, Köhler’s Medizinal-Pflanzen (Wikimedia Commons).

Genome sequence of the insect pathogenic fungus Cordyceps militaris

March 4th, 2012


Species in the ascomycete fungal genus Cordyceps have been proposed to be the teleomorphs of Metarhizium species. The latter have been widely used as insect biocontrol agents. Cordyceps species are highly prized for use in traditional Chinese medicines, but the genes responsible for biosynthesis of bioactive components, insect pathogenicity and the control of sexuality and fruiting have not been determined.

Chengshu Wang’s group from the Shanghai Institutes for Biological Sciences report the genome sequence of the type species Cordyceps militaries in the last issue of Genome Biology. Phylogenomic analysis suggests that different species in the Cordyceps/Metarhizium genera have evolved into insect pathogens independently of each other, and that their similar large secretomes and gene family expansions are due to convergent evolution. However, relative to other fungi, including Metarhizium spp., many protein families are reduced in C. militaris, which suggests a more restricted ecology. Consistent with its long track record of safe usage as a medicine, the Cordyceps genome does not contain genes for known human mycotoxins. This study shows that C. militaris is sexually heterothallic but, very unusually, fruiting can occur without an opposite mating-type partner. Transcriptional profiling indicates that fruiting involves induction of the Zn2Cys6-type transcription factors and MAPK pathway; unlike other fungi, however, the PKA pathway is not activated.

The data offer a better understanding of Cordyceps biology and will facilitate the exploitation of medicinal compounds produced by the fungus.

Zheng et al. (2011) Genome Biology 12: R116

Photo: Chinese Tussah silkmoth pupae colonized by C. militaris (© Zheng et al.)

Genomes of Skin Invaders

February 27th, 2011

hyphaecolonizinghairPathogenic fungi colonizing human skin, so-called dermatophytes, are a pleague for millions of humans. There are highly specialized filamentous fungi exclusively infecting keratinized animal structures causing mycoses.  Studies of dermatophyte pathogenic interactions have been hampered by a lack of full genome sequences. To provide broad insights into the molecular basis of the pathogenicity-associated traits, Burmester et al. published in the last issue of Genome Biology (2011/12/1/R7)  the first genome sequences of two closely phylogenetically related dermatophytes, Arthroderma benhamiae and Trichophyton verrucosum, both of which induce highly inflammatory infections in humans.

The genomes of A. benhamiae and T. verrucosum were sequenced by a whole-genome shotgun hybrid approach (Sanger, 454 FLX). These genomes are smaller than those of phylogenetically related ascomycetes. The assembly of A. benhamiae spans 22.3 Mb and that of T. verrucosum comprises 22.6 Mb. The genomes of A. benhamiae and T. verrucosum are compact and only contain 7,980 and 8,024 predicted protein-encoding genes, respectively. Most of these genes lie in collinear regions and are shared between the two fungi suggesting a very recent speciation.

As expected from their peculiar ecological niche, i.e. the animal skin, the two dermatophytes belong to the most protease-rich fungal species. They contain 235 protease-encoding genes, 87 of the predicted proteins having a secretion signal. A comprehensive analysis of the secretome during keratin degradation was carried out by combining 2D-PAGE and MALDI-TOF/TOF. RNA-Seq transcriptome profiling of A. benhamiae growing on human keratinocytes was performed to investigate the the entire process of infection. Subtilisin-like serine proteases, fungalysine-type metalloproteases, leucine aminopeptidases and dipeptidyl-peptidases are secreted during growth of A. benhamiae on keratin and keratinocytes. In addition, these dermatophytes can efficiently assimilate lipids, major constituents of the skin, thanks to the presence of 16 lipase genes. As discussed by the authors, the presence of large protease gene families in dermatophytes and their striking induction on keratin and human keratinocytes likely reflects selection during evolution and the ability of these fungi to adapt to different environmental conditions during infection and saprophytic growth. It appears that secondary metabolites also play a crucial role during keratinocyte infection.

Burmester et al. (2011) Comparative and functional genomics provide insights into the pathogenicity of dermatophytic fungi. Genome Biology 12:R7

Photo: A. benhamiae on human hair © Burmester et al.

Friend or foe? … a Sweet Story

August 16th, 2010

Sans titreThe sky is overcast with clouds and the rain is ceaseless‘ in Gitanjali by Rabindranath Tagore.

Mid-Summer: Across Lorraine, this is generally the hottest month of the year, and when most everyone is headed to a beach or the mountains to look for a little respite. No scorching mid-Summer heat this year. Over the week-end, the sky was pouring with rain and the sun never shone from dawn to dusk. It poured with rain the entire night.

Well, I take this opportunity to get back to outstanding papers. The study of GH32 invertases in fungi from Jerri Parent, together with Tim James and Andy Taylor, was worth re-reading. In this BMC Evolutionary Biology paper, they tested for occurrence of the glycosyl hydrolase family 32 (GH32) genes in all available fungal genomes and an additional 149 species representing a broad phylogenetic and ecological range of biotrophic fungi. This GH32 family, containing mostly invertases, is crucial for plant-interacting fungi. Sucrose is the primary metabolite used by most plants to translocate carbon throughout their tissues, and its abundance within plants makes it a valuable carbon source for the many fungi that are obligate plant associates. To acquire the host sucrose, colonizing fungi must possess the necessary enzymes, such as extracellular invertase(s), to split sucrose into its constituent monosaccharides, glucose and fructose.

Ancestral state reconstruction of GH32 gene abundance showed a strong correlation with nutritional mode (saprobic, endophytic, mutualist, pathogenic). Expansion of gene families was observed in several clades of pathogenic filamentous Ascomycota species. GH32 gene number was negatively correlated with animal pathogenicity and positively correlated with plant biotrophy (e.g. Puccinia graminis), with the notable exception of mycorrhizal taxa (e.g. Laccaria bicolor). Few mycorrhizal species were found to have GH32 genes as compared to other guilds of plant-associated fungi, such as pathogens, endophytes and lichen-forming fungi. GH32 genes were also more prevalent in the Ascomycota than in the Basidiomycota.

We noticed in our Nature paper the lack of invertase in the ectomycorrhizal L. bicolor suggesting that this symbiont depends on its host plant to provide glucose in exchange for nitrogen. I have checked the presence of the extracellular invertase in our draft genome sequences of L. amethystina and Glomus intraradices, and the 454 transcripts of Lactarius quietus and Pisolithus microcarpus. None of these ECM fungi have a gene coding for this enzyme, whereas the poplar rust, Melampsora larici-populina has two sequences similar to the wheat rust Puccinia graminis GH32s. Intriguingly, the genome of the ectomycorrhizal ascomycete, Tuber melanosporum — the Black Truffe of Perigord — contains a gene encoding a GH32 enzyme, suggesting that the truffle may act as a scavenger instead of being a true mutualist. However, the corresponding transcript is lowly expressed in free-living mycelium, fruiting body and ectomycorrhiza according to our NimbleGen oligoarray and RNA-Seq transcript profilings.

As stressed by Parent et al. “Reliance on plant GH32 enzyme activity for C acquisition in these [ECM] symbionts supports earlier predictions of a general absence of invertase in mycorrhizal fungi, and a highly evolved mutualistic relationship between plants and mycorrhizal fungi, a remarkable scenario in light of the high degree of phylogenetic diversity spanned by mycorrhizal fungal taxa. Whether the plant host is able to detect fungal invertase activity and use such a signal to differentiate antagonistic from mutualistic biotrophic symbionts is a completely speculative, though plausible hypothesis.”

Parrent et al. (2009) Friend or foe? Evolutionary history of glycoside hydrolase family 32 genes encoding for sucrolytic activity in fungi and its implications for plant-fungal symbioses. BMC Evolutionary Biology 9:148 doi:10.1186/1471-2148-9-148.

Photo: Lorraine Big Sky © F Martin