Evolutionary and genomic comparisons of hybrid uninucleate and nonhybrid Rhizoctonia fungi

The basidiomycetous fungal genus, Rhizoctonia, can cause severe damage to many plants and is composed of multinucleate, binucleate, and uninucleate species differing in pathogenicity. Here we generated chromosome-scale genome assemblies of the three nuclear types of Rhizoctonia isolates. The genomic comparisons revealed that the uninucleate JN strain likely arose by somatic hybridization of two binucleate isolates, and maintained a diploid nucleus. Homeolog gene pairs in the JN genome have experienced both decelerated or accelerated evolution. Homeolog expression dominance occurred between JN subgenomes, in which differentially expressed genes show potentially less evolutionary constraint than the genes without. Analysis of mating-type genes suggested that Rhizoctonia maintains the ancestral tetrapolarity of the Basidiomycota. Long terminal repeat-retrotransposons displayed a reciprocal correlation with the chromosomal GC content in the three chromosome-scale genomes. The more aggressive multinucleate XN strain had more genes encoding enzymes for host cell wall decomposition. These findings demonstrate some evolutionary changes of a recently derived hybrid and in multiple nuclear types of Rhizoctonia.


Supplementary Figures
Supplementary Figure 1 Rhizoctonia strains JN, LY and XN.Mycelial growth of (a) JN, (b) LY, and (c) XN on PDA medium for 96 h and photographed from top (left) and underside (right) of each petri dish.(d-f) DAPI staining of nuclei in hyphae of JN (uninucleate), LY (binucleate) and XN (multinucleate).

2 Supplementary Figure 3 4 5 6
The 17-mer depth distribution of whole-genome Illumina reads of Rhizoctonia spp..The bimodal distributions in isolates RW and XN suggested high heterozygous genomes.Circos-plot characterizations of Rhizoctonia LY genome.I, the frequency distributions of read coverage (bar 0-250×).II, the syntenic gene links with 17 intragenomic syntenic fragments of 496 genes.III-V, the frequency distribution of repetitive density (bar 0-100%), LTR density (bar 0-100%) and heterozygous rate (bar 0-10‰), respectively.VI, the chromosome karyotype.All statistics were based on nonoverlapping windows (window size = 25 kb).Circos-plot characterization of XN genomic features.I, the frequency distributions of read coverage (bar 0-300×).II, the syntenic gene links, having eight intragenomic syntenic fragments of 464 genes.III-V, the frequency distribution of repetitive density (bar 0-100%), LTR density (bar 0-100%) and heterozygous rate (bar 0-10‰), respectively.VI, the chromosome karyotype.All statistics were based on nonoverlapping windows (window size = 25 kb).Copy number distribution of orthologous genes in Rhizoctonia spp.. Gene number is presented as a percentage of the total genes.Distribution of syntenic genes between JN subgenomes and LY genome.(a) Amino acid (aa) sequence identity between the syntenic gene pairs of LY and each JN subgenome (JNa and JNb).(b) Boxplots of the distribution of Ks between the syntenic gene pairs.(c) Links of syntenic genes between JN ( relationship between Basidiomycota Rhizoctonia and selected Ascomycota fungi using RAxML.Single-copy orthologous genes (637) were obtained for construction of the phylogenetic tree.Selected Basidiomycota Rhizoctonia: hybrid uninucleate JN (divided into JNa and JNb subgenomes) and SM (divided into SMa and SMb subgenomes), binucleate LY and YR, and multinucleate XN and GD118 PRJNA51401, and the reference Ascomycota: C. glabrata PRJNA374542, L. waltii PRJNA10734, S. cerevisiae, reads.(a) The LTR-RT reads of genome sequencing were compared with the number of full-length LTR-RTs (FL-TRs).The number at the top of each bar represented the ratio of FL-TR to LTR-RT reads, normalized to XN.The bottom showed the GC content of LTR-RTs, genome and exons.(b) Linear relationship of transcriptional reads with the numbers of FL-RTs or FL-RTs with insertion age of

Supplementary Figure 9
Relationship of syntenic FL-RTs among LY and JN subgenomes.(a) between LY and JNa, (b) between LY and JNb, and (c) among JNa, JNb and LY.The FL-RTs at the same horizontal ordinate indicating synteny and the FL-RTs overlap indicating the same insertion time.
Ti/Tv for the ratio of transitions to transversions.
of GC contents of 4508 orthologous genes among JNa, JNb, LY, and XN.(a-b) Correlation between the frequency of GC3 and GC3*, reflecting the AT to GC substitution rates of JN subgenomes (JNa and JNb) in relation to XN orthologs at the third codon positions.(c) GC3 SNP mutation patterns and ratios of transition (Ti) to transversion (Tv) in the data sets.

Supplementary Figure 21 Comparison of genome-wide GC contents among JNa, JNb
, LY, and XN.P-values shown in corresponding comparisons.

Table 2
Description of Rhizoctonia genome assembled using Illumina reads 1Representing the Longest or Average length of scaffold (bp)2Divided into JNa and JNb subgenomes; 175 scaffolds(2,178,459 bp, 494 genes)of JN could not be assigned to the JNa and JNb subgenoemes.3DividedintoSMaandSMb subgenomes; 211 scaffolds (1,725,342 bp, 333 genes) of SM could not be assigned to the SMa and SMb subgenoemes.4DividedintoYRa and YRb subgenomes; 97 scaffolds (396,674 bp, 71 genes) of YR could not be assigned to the YRa and YRb subgenoemes.Supplementary

Table 6
Characteristics of LY chromosomes Characteristics of XN chromosomes tRNA characteristic constituents of Rhizoctonia spp.Prediction of secreted and effector proteins in Rhizoctonia spp.