Haplotype‐phased and chromosome‐level genome assembly of Puccinia polysora, a giga‐scale fungal pathogen causing southern corn rust

Abstract

Rust fungi are characterized by large genomes with high repeat content and have two haploid nuclei in most life stages, which makes achieving high-quality genome assemblies challenging. Here, we described a pipeline using HiFi reads and Hi-C data to assemble a gigabase-sized fungal pathogen, Puccinia polysora f.sp. zeae, to haplotype-phased and chromosome-scale. The final assembled genome is 1.71 Gbp, with ~850 Mbp and 18 chromosomes in each haplotype, being currently one of the two giga-scale fungi assembled to chromosome level. Transcript-based annotation identified 47,512 genes for the dikaryotic genome with a similar number for each haplotype. A high level of interhaplotype variation was found with 10% haplotypespecific BUSCO genes, 5.8 SNPs/kbp, and structural variation accounting for 3% of the genome size. The P. polysora genome displayed over 85% repeat contents, with genome-size expansion and copy number increasing of species-specific orthogroups.Interestingly, these features did not affect overall synteny with other Puccinia species having smaller genomes. Fine-time-point transcriptomics revealed seven clusters of coexpressed secreted proteins that are conserved between two haplotypes. The fact that candidate effectors interspersed with all genes indicated the absence of a "two-speed genome" evolution in P. polysora. Genome resequencing of 79 additional isolates revealed a clonal population structure of P. polysora in China with low geographic differentiation. Nevertheless, a minor population differentiated from the major population by having mutations on secreted proteins including AvrRppC, indicating the ongoing virulence to evade recognition by RppC, a major resistance gene in Chinese corn cultivars. The high-quality assembly provides valuable genomic resources for future studies on disease management and the evolution of P. polysora.

Resolving the haplotypes of arbuscular mycorrhizal fungi highlights the role of two nuclear populations in host interactions

“…On average, only 54 short contigs totaling 2.7 Mb (L50: 61 Kb) remained un-scaffolded per strain. Consistent with phasing of fungal dikaryons based on Hi-C data 22–24 , the haplotypes of AMF heterokaryons exhibit a strong dikaryotic phasing signal, with ∼90% of inter-chromosome ( trans ) Hi-C contacts contained within nuclear haplotypes ( Supplemental Figures 1-4) . In our assemblies, one part of the chromosome 28 previously identified in AMF homokaryons 25 maps to chromosome 2 and the other to chromosome 17, but inspection of Hi-C contact maps and HiFi read coverage did not provide conclusive evidence that these chromosomes should be broken.…”

“…On average, only 54 short contigs totaling 2.7 Mb (L50: 61 Kb) remained un-scaffolded per strain. Consistent with phasing of fungal dikaryons based on Hi-C data 22–24 , the haplotypes of AMF heterokaryons exhibit a strong dikaryotic phasing signal, with ~90% of inter-chromosome ( trans ) Hi-C contacts contained within nuclear haplotypes ( Supplemental Figures 1-4) . In support of molecular analyses of single spores 17 , the MAT-2 and MAT-5 haplotypes have higher coverage (relative abundance) in the isolates A4 and G1, respectively (~9.2% and 63% higher) but are closer to balance in A5 and SL1 in extraradical mycelium after propagation with Daucus carota cv P68 (Carrot; 3.5% average difference) ( Table 1 ).…”

“…High-molecular-weight DNA was extracted for all strains using a protocol proposed by Schwessinger & McDonald 43 , and over 10 micrograms of DNA was subjected to high fidelity (HiFi) PacBio sequencing at Discovery Life Science (Huntsville, AL). PacBio sequencing produced an average 24,647,320,191 base pairs (bp) of HiFi reads with read quality of Q33 and median read length of 13,138.…”

Genome biology and evolution of mating type loci in four cereal rust fungi

“…polysora f. sp. zeae STE3.2-1 as outgroup for rooting the gene tree (63). The two STE3.2-1 alleles identified in each isolate were almost completely identical and grouped by species identity.…”

“…MAT genes including the HD and PRA genes previously identified in P. triticina (Table 2) (18) were used as blastp and blastn (89) query to identify orthologs in P. graminis f.sp tritici Ug99 and 21-0 isolates (30), P. striiformis f.sp tritici 134E (47), 104E (48) and DK0911 (49), P. triticina 76 (43), 19NSW04 , 20QLD87 (32) and 15 (55), P. polysora GD1913 (63), P. coronate f.sp avenae 203 (39), 12NC29 and 12SD80 (38) isolates. Missed STE3.2-3 in Pst 104E was fixed by mapping short reads to reference genome Pst 134E with bwa-mem2 (90).…”