Pathogenicity Determinants in Smut Fungi Revealed by Genome Comparison

Biotrophic pathogens, such as the related maize pathogenic fungi Ustilago maydis and Sporisorium reilianum, establish an intimate relationship with their hosts by secreting protein effectors. Because secreted effectors interacting with plant proteins should rapidly evolve, we identified variable genomic regions by sequencing the genome of S. reilianum and comparing it with the U. maydis genome. We detected 43 regions of low sequence conservation in otherwise well-conserved syntenic genomes. These regions primarily encode secreted effectors and include previously identified virulence clusters. By deletion analysis in U. maydis, we demonstrate a role in virulence for four previously unknown diversity regions. This highlights the power of comparative genomics of closely related species for identification of virulence determinants.

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Fungal Pathogen Emergence: Investigations with an Ustilago maydis × Sporisorium reilianum Hybrid

Journal of Fungi ◽

10.3390/jof7080672 ◽

2021 ◽

Vol 7 (8) ◽

pp. 672

Author(s):

Emilee R. M. Storfie ◽

Barry J. Saville

Keyword(s):

Zea Mays ◽

Fungal Pathogen ◽

Fungal Pathogens ◽

Crop Production ◽

Virulence Genes ◽

Ustilago Maydis ◽

Smut Fungi ◽

Transcript Levels ◽

Sporisorium Reilianum ◽

Molecular Aspects

The emergence of new fungal pathogens threatens sustainable crop production worldwide. One mechanism by which new pathogens may arise is hybridization. To investigate hybridization, the related smut fungi, Ustilago maydis and Sporisorium reilianum, were selected because they both infect Zea mays, can hybridize, and tools are available for their analysis. The hybrid dikaryons of these fungi grew as filaments on plates but their colonization and virulence in Z. mays were reduced compared to the parental dikaryons. The anthocyanin induction caused by the hybrid dikaryon infections was distinct, suggesting its interaction with the host was different from that of the parental dikaryons. Selected virulence genes previously characterized in U. maydis and their predicted S. reilianum orthologs had altered transcript levels during hybrid infection of Z. mays. The downregulated U. maydis effectors, tin2, pit2, and cce1, and transcription factors, rbf1, hdp2, and nlt1, were constitutively expressed in the hybrid. Little impact was observed with increased effector expression; however, increased expression of rbf1 and hdp2, which regulate early pathogenic development by U. maydis, increased the hybrid’s capacity to induce symptoms including the rare induction of small leaf tumors. These results establish a base for investigating molecular aspects of smut fungal hybrid pathogen emergence.

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Higher Gene Flow in Sex-Related Chromosomes than in Autosomes during Fungal Divergence

Molecular Biology and Evolution ◽

10.1093/molbev/msz252 ◽

2019 ◽

Vol 37 (3) ◽

pp. 668-682 ◽

Cited By ~ 6

Author(s):

Fanny E Hartmann ◽

Ricardo C Rodríguez de la Vega ◽

Pierre Gladieux ◽

Wen-Juan Ma ◽

Michael E Hood ◽

...

Keyword(s):

Gene Flow ◽

Mating Type ◽

Sex Chromosomes ◽

Smut Fungi ◽

Effective Population ◽

Closely Related Species ◽

Fungal Mating ◽

Anther Smut ◽

Genomic Regions

Abstract Nonrecombining sex chromosomes are widely found to be more differentiated than autosomes among closely related species, due to smaller effective population size and/or to a disproportionally large-X effect in reproductive isolation. Although fungal mating-type chromosomes can also display large nonrecombining regions, their levels of differentiation compared with autosomes have been little studied. Anther-smut fungi from the Microbotryum genus are castrating pathogens of Caryophyllaceae plants with largely nonrecombining mating-type chromosomes. Using whole genome sequences of 40 fungal strains, we quantified genetic differentiation among strains isolated from the geographically overlapping North American species and subspecies of Silene virginica and S. caroliniana. We inferred that gene flow likely occurred at the early stages of divergence and then completely stopped. We identified large autosomal genomic regions with chromosomal inversions, with higher genetic divergence than the rest of the genomes and highly enriched in selective sweeps, supporting a role of rearrangements in preventing gene flow in genomic regions involved in ecological divergence. Unexpectedly, the nonrecombining mating-type chromosomes showed lower divergence than autosomes due to higher gene flow, which may be promoted by adaptive introgressions of less degenerated mating-type chromosomes. The fact that both mating-type chromosomes are always heterozygous and nonrecombining may explain such patterns that oppose to those found for XY or ZW sex chromosomes. The specific features of mating-type chromosomes may also apply to the UV sex chromosomes determining sexes at the haploid stage in algae and bryophytes and may help test general hypotheses on the evolutionary specificities of sex-related chromosomes.

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A cell surface-exposed protein complex with an essential virulence function in Ustilago maydis

Nature Microbiology ◽

10.1038/s41564-021-00896-x ◽

2021 ◽

Author(s):

Nicole Ludwig ◽

Stefanie Reissmann ◽

Kerstin Schipper ◽

Carla Gonzalez ◽

Daniela Assmann ◽

...

Keyword(s):

Membrane Proteins ◽

Protein Complex ◽

Pathogenic Bacteria ◽

Constitutive Expression ◽

Pathogenic Fungi ◽

Ustilago Maydis ◽

Effector Proteins ◽

Host Cells ◽

Smut Fungi ◽

Living Plant

AbstractPlant pathogenic fungi colonizing living plant tissue secrete a cocktail of effector proteins to suppress plant immunity and reprogramme host cells. Although many of these effectors function inside host cells, delivery systems used by pathogenic bacteria to translocate effectors into host cells have not been detected in fungi. Here, we show that five unrelated effectors and two membrane proteins from Ustilago maydis, a biotrophic fungus causing smut disease in corn, form a stable protein complex. All seven genes appear co-regulated and are only expressed during colonization. Single mutants arrest in the epidermal layer, fail to suppress host defence responses and fail to induce non-host resistance, two reactions that likely depend on translocated effectors. The complex is anchored in the fungal membrane, protrudes into host cells and likely contacts channel-forming plant plasma membrane proteins. Constitutive expression of all seven complex members resulted in a surface-exposed form in cultured U. maydis cells. As orthologues of the complex-forming proteins are conserved in smut fungi, the complex may become an interesting fungicide target.

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Mating Type Loci of Sporisorium reilianum: Novel Pattern with Three a and Multiple b Specificities

Eukaryotic Cell ◽

10.1128/ec.4.8.1317-1327.2005 ◽

2005 ◽

Vol 4 (8) ◽

pp. 1317-1327 ◽

Cited By ~ 68

Author(s):

Jan Schirawski ◽

Bernadette Heinze ◽

Martin Wagenknecht ◽

Regine Kahmann

Keyword(s):

Mating Type ◽

Molecular Basis ◽

Plant Invasion ◽

Ustilago Maydis ◽

Smut Fungi ◽

Symptom Development ◽

Receptor System ◽

Mating Partner ◽

Sporisorium Reilianum ◽

High Degree

ABSTRACT Sporisorium reilianum and Ustilago maydis are two closely related smut fungi, which both infect maize but differ fundamentally in their mode of plant invasion and site of symptom development. As a prelude to studying the molecular basis of these differences, we have characterized the mating type loci of S. reilianum. S. reilianum has two unlinked mating type loci, a and b. Genes in both loci and adjacent regions show a high degree of synteny to the corresponding genes of U. maydis. The b locus occurs in at least five alleles and encodes two subunits of a heterodimeric homeodomain transcription factor, while the a locus encodes a pheromone/receptor system. However, in contrast to that of U. maydis, the a locus of S. reilianum exists in three alleles containing two active pheromone genes each. The alleles of the a locus appear to have arisen through recent recombination events within the locus itself. This has created a situation where each pheromone is specific for recognition by only one mating partner.

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Cross-species analysis between the maize smut fungi Ustilago maydis and Sporisorium reilianum highlights the role of transcriptional plasticity of effector orthologs for virulence and disease

10.1101/2020.11.03.366443 ◽

2020 ◽

Cited By ~ 1

Author(s):

Weiliang Zuo ◽

Deepak K Gupta ◽

Jasper RL Depotter ◽

Marco Thines ◽

Gunther Doehlemann

Keyword(s):

Ustilago Maydis ◽

Effector Proteins ◽

Differentially Expressed ◽

Transcriptional Level ◽

Smut Fungi ◽

List Type ◽

Sequencing Analysis ◽

Distinct Disease ◽

Sporisorium Reilianum ◽

Transcriptional Plasticity

SummaryThe constitution and regulation of effector repertoires determines and shapes the outcome of the interaction with the host. Ustilago maydis and Sporisorium reilianum are two closely related smut fungi, which both infect maize, but cause distinct disease symptoms. Understanding how effector orthologs are regulated in these two pathogens can therefore provide insights to pathogen evolution and host adaption.We tracked the infection progress of U. maydis and S. reilianum in maize leaves, characterized two distinct infection stages for cross species RNA-sequencing analysis and identified 207 out of 335 one-to-one effector orthologs being differentially regulated during host colonization, while transcriptional plasticity of the effector orthologs correlated with the distinct disease development strategies.By using CRISPR-Cas9 mediated gene conversion, we identified two differentially expressed effector orthologs with conserved function between two pathogens. Thus, differential expression of functionally conserved genes contributes to species specific adaptation and symptom development. Interestingly, another differentially expressed orthogroup (UMAG_05318/sr1007) showed diverged protein function during speciation, providing a possible case for neofunctionalization.Together, we showed the diversification of effector genes in related pathogens can be caused both by plasticity on the transcriptional level, as well as through neofunctionalization of the encoded effector proteins.

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Cross‐species analysis between the maize smut fungi Ustilago maydis and Sporisorium reilianum highlights the role of transcriptional change of effector orthologs for virulence and disease

New Phytologist ◽

10.1111/nph.17625 ◽

2021 ◽

Author(s):

Weiliang Zuo ◽

Jasper RL Depotter ◽

Deepak K Gupta ◽

Marco Thines ◽

Gunther Doehlemann

Keyword(s):

Ustilago Maydis ◽

Smut Fungi ◽

Transcriptional Change ◽

Species Analysis ◽

Sporisorium Reilianum ◽

Maize Smut

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Development of a loop-mediated isothermal amplification assay for the detection of Tilletia controversa based on genome comparison

Scientific Reports ◽

10.1038/s41598-021-91098-2 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Somayyeh Sedaghatjoo ◽

Monika K. Forster ◽

Ludwig Niessen ◽

Petr Karlovsky ◽

Berta Killermann ◽

...

Keyword(s):

Test Performance ◽

Genomic Dna ◽

Lamp Assay ◽

Fungal Species ◽

Isothermal Amplification ◽

Genome Comparison ◽

Performance Study ◽

Loop Mediated Isothermal Amplification ◽

Tilletia Controversa ◽

Genomic Regions

AbstractTilletia controversa causing dwarf bunt of wheat is a quarantine pathogen in several countries. Therefore, its specific detection is of great phytosanitary importance. Genomic regions routinely used for phylogenetic inferences lack suitable polymorphisms for the development of species-specific markers. We therefore compared 21 genomes of six Tilletia species to identify DNA regions that were unique and conserved in all T. controversa isolates and had no or limited homology to other Tilletia species. A loop-mediated isothermal amplification (LAMP) assay for T. controversa was developed based on one of these DNA regions. The specificity of the assay was verified using 223 fungal samples comprising 43 fungal species including 11 Tilletia species, in particular 39 specimens of T. controversa, 92 of T. caries and 40 of T. laevis, respectively. The assay specifically amplified genomic DNA of T. controversa from pure cultures and teliospores. Only Tilletia trabutii generated false positive signals. The detection limit of the LAMP assay was 5 pg of genomic DNA per reaction. A test performance study that included five laboratories in Germany resulted in 100% sensitivity and 97.7% specificity of the assay. Genomic regions, specific to common bunt (Tilletia caries and Tilletia laevis together) are also provided.

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Chromosomal Map of the Model LegumeLotus japonicus

Genetics ◽

10.1093/genetics/161.4.1661 ◽

2002 ◽

Vol 161 (4) ◽

pp. 1661-1672 ◽

Cited By ~ 7

Author(s):

Andrea Pedrosa ◽

Niels Sandal ◽

Jens Stougaard ◽

Dieter Schweizer ◽

Andreas Bachmair

Keyword(s):

Repetitive Sequences ◽

Lotus Japonicus ◽

Chromosome Complement ◽

Linkage Groups ◽

Closely Related Species ◽

F1 Hybrid ◽

Bac Clones ◽

Genomic Regions ◽

Chromosomal Map

AbstractLotus japonicus is a model plant for the legume family. To facilitate map-based cloning approaches and genome analysis, we performed an extensive characterization of the chromosome complement of the species. A detailed karyotype of L. japonicus Gifu was built and plasmid and BAC clones, corresponding to genetically mapped markers (see the accompanying article by Sandal et al. 2002, this issue), were used for FISH to correlate genetic and chromosomal maps. Hybridization of DNA clones from 32 different genomic regions enabled the assignment of linkage groups to chromosomes, the comparison between genetic and physical distances throughout the genome, and the partial characterization of different repetitive sequences, including telomeric and centromeric repeats. Additional analysis of L. filicaulis and its F1 hybrid with L. japonicus demonstrated the occurrence of inversions between these closely related species, suggesting that these chromosome rearrangements are early events in speciation of this group.

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Ultrastructure of meiosis and the spindle pole body cycle in freeze-substituted basidia of the smut fungi Ustilago maydis and Ustilago avenae

Canadian Journal of Botany ◽

10.1139/b92-081 ◽

1992 ◽

Vol 70 (3) ◽

pp. 629-638 ◽

Cited By ~ 8

Author(s):

Kerry O'Donnell

Keyword(s):

Electron Microscopy ◽

Spindle Pole Body ◽

Ustilago Maydis ◽

Spindle Pole ◽

Meiotic Spindle ◽

Smut Fungi ◽

Late Prophase ◽

Prophase I ◽

Pole Body ◽

Middle Piece

Meiosis in the smut fungi Ustilago maydis and Ustilago avenae (Basidiomycota, Ustilaginales) was studied by electron microscopy of serial-sectioned freeze substituted basidia. At prophase I, a spindle pole body composed of two globular elements connected by a middle piece was attached to the extranuclear surface of each nucleus. Astral and spindle microtubules were initiated at each globular element at late prophase I to prometaphase I. During spindle initiation, the middle piece disappeared and interdigitating half-spindles entered the nucleoplasm, which was surrounded by discontinuous nuclear envelope together with perinuclear endoplasmic reticulum. Kinetochore pairs at metaphase I were analyzed to obtain a karyotype for each species. The meiotic spindle pole body replicational cycle is described. Key words: electron microscopy, freeze-substitution, meiosis, Ustilago, spindle pole body.

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