Genome sequencing of Sporisorium scitamineum provides insights into the pathogenic mechanisms of sugarcane smut

Abstract

Background: Sugarcane smut can cause losses in cane yield and sugar content that range from 30% to total crop failure. Losses tend to increase with the passage of years. Sporisorium scitamineum is the fungus that causes sugarcane smut. This fungus has the potential to infect all sugarcane species unless a species is resistant to biotrophic fungal pathogens. However, it remains unclear how the fungus breaks through the cell walls of sugarcane and causes the formation of black or gray whip-like structures on the sugarcane plants.

Results: Here, we report the first high-quality genome sequence of S. scitamineum assembled de novo with a contig N50 of 41 kb, a scaffold N50 of 884 kb and genome size 19.8 Mb, containing an estimated 6,636 genes. This phytopathogen can utilize a wide range of carbon and nitrogen sources. A reduced set of genes encoding plant cell wall hydrolytic enzymes leads to its biotrophic lifestyle, in which damage to the host should be minimized. As a bipolar mating fungus, a and b loci are linked and the mating-type locus segregates as a single locus. The S. scitamineum genome has only 6 G protein-coupled receptors (GPCRs) grouped into five classes, which are responsible for transducing extracellular signals into intracellular responses, however, the genome is without any PTH11-like GPCR. There are 192 virulence associated genes in the genome of S. scitamineum, among which 31 expressed in all the stages, which mainly encode for energy metabolism and redox of short-chain compound related enzymes. Sixty-eight candidates for secreted effector proteins (CSEPs) were found in the genome of S. scitamineum, and 32 of them expressed in the different stages of sugarcane infection, which are probably involved in infection and/or triggering defense responses. There are two non-ribosomal peptide synthetase (NRPS) gene clusters that are involved in the generation of ferrichrome and ferrichrome A, while the terpenes gene cluster is composed of three unknown function genes and seven biosynthesis related genes.

Conclusions: As a destructive pathogen to sugar industry, the S. scitamineum genome will facilitate future research on the genomic basis and the pathogenic mechanisms of sugarcane smut.

Figure 1

Infection process of Sporisorium scitamineum on sugarcane. 1, Teliospore; 2, Soil or sugarcane plants contaminated with S. scitamineum; 3, Teliospore germination; 4, The formation of infection hyphae; 5, Hypha infection in sugarcane meristem and growth point; 6, The formation of disease panicle.

Figure 2

Overlap among gene families for four smut fungi species. This is the venn diagram used to compare the gene families of four smut fungi, four colors (green, red, yellow and blue) mean four gene family sets, and the overlap parts have the overlapping color. Numbers indicate the gene families in each comparison. S means the gene families specific to S. scitamineum, S-U means common gene families of S. scitamineum and U. hordei, S-U-R means common gene families of S. scitamineum, U. hordei and S. reilianum, S-U-R-M means common gene families of four smut fungi.

Figure 3

Taxonomic placement of Sporisorium scitamineum and genome synteny with S. reilianum , Ustilago maydis, and U. hordei . (a) Evolutionary placing of S. scitamineum in relation to other smut fungi and A. nidulans. Numbers on the branch indicate branch length. Numbers for the internal node indicate bootstrap value; (b) Dot plot of the synteny occurring between the assembled chromosomes of S. reilianum and the assembled scaffolds of S. scitamineum; (c) Dot plot of the synteny occurring between the assembled chromosomes of U. maydis and the assembled scaffolds of S. scitamineum; (d) Dot plot of the synteny occurring between the assembled chromosomes of U. hordei and the assembled scaffolds of S. scitamineum.

Figure 4

Genetic organization of the mating-type loci of smut fungi. Genes are indicated by arrows with the arrow denoting the direction of transcription. Related genes are denoted by the same color and respective gene functions are explained in the lower part of the Figure. *indicates that the relative order and orientation of these genes have not been determined. In the tetrapolar species, U. maydis and S. reilianum, the a and b specific sequences reside on different chromosomes, while they are linked by spacer regions (which are not drawn to scale and whose length is indicated) in the bipolar species U. hordei and S. scitamineum. The black bars on top of the figure indicate the regions of the b locus, which covers the two homeodomain protein genes bE and bW, and the a locus (that expands to different length in the different loci, indicated by a broken line) from the lba gene to the rba gene. Sequence information was obtained from the following Accession Numbers: AF043940, AM118080, AF184070, AF184069, Z18531, AJ884588, AJ884583, AJ884590, AJ884585, AJ884589, AJ884584, U37796, M84182, AACP01000083 and AACP01000013.

Figure 5

The expression pattern of candidate genes for encoding pant cell wall degradation enzymes of Sporisorium scitamineum at six stages in the infection process. These infection stages includes six time points during the course of infection (the 0-, 12-, 24-, 48-, 96-, and 120-hpi). The number on the left side, such as Ss 6317, means gene ID. The color scale indicates relative RT-qPCR expression.

Figure 6

Comparative growth profiling of Sporisorium scitamineum on various carbon substrates. This figure is used to explain that S. scitamineum can utilize all the eight carbon substrates without significant differences.

Figure 7

Comparative growth profiling of Sporisorium scitamineum on various nitrogen substrates. This figure is used to explain that S. scitamineum can uptake a broad range of nitrogen sources, including sodium nitrate, ammonium sulfate, ammonium acetate, and all the twenty kinds of amino acids, without significant differences.

Figure 8

The expression pattern of candidate virulence associated genes of Sporisorium scitamineum at six stages in the infection process. These infection stages includes six time points during the course of infection (the 0-, 12-, 24-, 48-, 96-, and 120-hpi). The number on the left side, such as Ss 3916, means gene ID. The color scale indicates relative RT-qPCR expression.

Figure 9

The expression pattern of genes coding for secreted effector proteins of Sporisorium scitamineum at six stages in the infection process. These infection stages includes six time points during the course of infection (the 0-, 12-, 24-, 48-, 96-, and 120-hpi). The number on the left side, such as Ss 0541, means gene ID. The color scale indicates relative RT-qPCR expression.

Figure 10

Three gene clusters in the genome of Sporisorium scitamineum . (a) Physical map of ferrichrome gene cluster (SmutADNA4_GLEAN_10002720-10002733). Coding regions are indicated with arrows and are putatively involved in ferrichrome biosynthesis (green yellow), siderophore transport (blue), or unknown functions (grey). Location of InterPro domains in NRPS SmutADNA4_GLEAN_10002728 is as bottom. AD, AMP-dependent synthetase/ligase domain (green), IPR00873; AC, acyl carrier domain (purple), IPR009081; CD, condensation domain (red), IPR001242; AB, AMP-binding enzyme C-terminal domain (black), IPR025110. (b) Physical map of ferrichrome A gene cluster (SmutADNA4_GLEAN_10004009-10004020). Coding regions are indicated with arrows and are putatively involved in ferrichrome A biosynthesis (green yellow), siderophore transport (blue), or unknown functions (grey). Location of InterPro domains in NRPS SmutADNA4_GLEAN_10004013 is as bottom. AD, AMP-dependent synthetase/ligase domain (green), IPR00873; AC, acyl carrier domain (purple), IPR009081;CD, condensation domain (red), IPR001242; AB, AMP-binding enzyme C-terminal domain (black), IPR025110. (c) Physical map of terpene gene cluster (SmutADNA4_GLEAN_10000695-10000704). Coding regions are indicated with arrows and are putatively involved in terpene biosynthesis (green yellow) or unknown functions (grey).