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

Abstract

BackgroundRust 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.ResultsTo 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.ConclusionsThe 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.

Transcriptome analysis of enriched Golovinomyces orontii haustoria by deep 454 pyrosequencing

“…Previously, haustorial transcriptomes of obligate biotrophic rust fungi have been shown to contain many transcripts involved in metabolism and protein synthesis as well as virulence (Catanzariti et al, 2006;Hahn and Mendgen, 1997;Xu et al, 2011;Yin et al, 2009). This is in line with our results, as we identified elements of gene expression and protein turnover to be the most abundant transcripts (Fig.…”

“…A comparatively low number of genes with homologs is also within the range of reports from rust (Puccinia striiformis f.sp. tritici) haustorial cDNA libraries (Xu et al, 2011;Yin et al, 2009). The genome of Bgh contains 5854 curated genes, a low number in comparison to other fungi (Schmidt and Panstruga, 2011;Spanu et al, 2010).…”

Uromycladium acaciae, the cause of a sudden, severe disease epidemic on Acacia mearnsii in South Africa

“…This large genome size is in part explained by transcriptome analyses that showed unique genes are expressed by different stages of the life cycle (Hacquard et al 2013;Xu et al 2011). Microcyclic rust fungi can lose their aecial and uredinial stages in conditions favourable for continuous growth and reproduction (Ono 2002); the unique genes associated with these stages may remain in the genome, but are not expressed.…”

Advancing Knowledge on Biology of Rust Fungi Through Genomics

“…A larger number of genes specifically expressed in telia were found, most of which encode unknown functions. Similarly, a large number of specific ESTs with unknown functions were identified in P. triticina teliospores (Xu et al, 2011) highlighting that the biological processes associated with this type of rust spores remain mostly uncharacterized. In M. larici-populina, telia and uredinia have the most similar expression profiles when compared to fungal hyphae undergoing biotrophic growth in poplar leaves, indicating shared components in the genetic programs ongoing in these spore-forming structures.…”

“…These gene expression studies particularly showed that the haustorium is a crucial structure both for nutrients uptake (Mendgen & Hahn, 2002;Voegele, Hahn, & Mendgen, 2009;Voegele, Struck, Hahn, & Mendgen, 2001) and effector release (Catanzariti, Dodds, Lawrence, Ayliffe, & Ellis, 2006;Hahn & Mendgen, 1997;Joly, Feau, Tanguay, & Hamelin, 2010;Link & Voegele, 2008). A few studies explored other stages of the rust fungi biology, providing insights into relevant functions expressed at various developmental steps of the rust life cycle (Warren & Covert, 2004;Xu et al, 2011). Most expression studies were focused on resting rust uredospores collected outside the host plant and during in planta biotrophic growth by directly isolating RNA from infected host tissues.…”