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

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% haplotype-specific 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.

Clonal Expansion and Dispersal Pathways of Puccinia polysora in China.

Southern corn rust (SCR) caused by Puccinia polysora is one of the most devastating diseases in the world. In recent years, SCR has been upgraded from a minor to a major disease around the world, including in China. However, little is known about its population genetics and structure in China. In this study, we analyzed 288 isolates collected from various localities during 2017 in seven Chinese provinces: Guangxi, Guangdong, Anhui, Hunan, Shandong, Henan, and Shaanxi. The isolates were analyzed using nine microsatellite markers. The population structure, genetic diversity, and reproduction mode of P. polysora were investigated based on genotype data. Strong genotypic diversity was detected and clonal reproduction was dominant. The populations collected from the pathogen's winter-reproductive regions harbored more genotypes than those collected from the pathogen's epidemic regions. The spatial differences in genotypic richness, and evenness among the populations were significant, and showed a decreasing trend from south to north. Most isolates were clustered into two clonal groups. Two high-frequency multilocus genotypes (MLGs), MLG1 and MLG2, were widely distributed in all populations. Our analyses confirmed that P. polysora employed clone dispersal from the pathogen's winter-reproductive regions to the pathogen's epidemic regions, and in addition to the sources from the pathogen's winter-reproductive regions, the pathogen in Anhui and Hunan might also have other sources from areas such as Taiwan, China, or/and Southeast Asia, and the pathogen went through a genetic bottleneck during its dispersal. These findings provide initial insights into the reproduction mode and dispersal pathways of P. polysora in China.

Population Genetic Structures of Puccinia striiformis f. sp. tritici in the Gansu-Ningxia Region and Hubei Province, China.

Wheat stripe rust, caused by the fungal pathogen Puccinia striiformis f. sp. tritici (Pst), is a destructive wheat disease in China. The Gansu-Ningxia region (GN) is a key area for pathogen over-summering in China, and northwestern Hubei (HB) is an important region for pathogen over-wintering, serving as a source of inoculum in spring epidemic regions. The spatiotemporal population genetic structure of Pst in HB and the pathogen population exchanges between GN and HB are important for estimating the risk of interregional epidemics. Here, 567 isolates from GN and HB were sampled from fall 2016 to spring 2018 and were genotyped using simple sequence repeat markers. The genotypic and genetic diversity of Pst subpopulations in HB varied among seasons and locations. Greater genetic diversification levels were found in the spring compared with fall populations using principal coordinate analysis and Bayesian assignments. In total, there were 17 common genotypes among the 208 determined, as shown by a small overlap of genotypes in the principal coordinate analysis and dissimilar Bayesian assignments in both regions, which revealed the limited genotype exchange between the populations of GN and HB.

De novo transcriptome assembly, polymorphic SSR markers development and population genetics analyses for southern corn rust (Puccinia polysora).

Southern corn rust is a destructive maize disease caused by Puccinia polysora Underw that can lead to severe yield losses. However, genomic information and microsatellite markers are currently unavailable for this disease. In this study, we generated a total of 27,295,216 high-quality cDNA sequence reads using Illumina sequencing technology. These reads were assembled into 17,496 unigenes with an average length of 1015 bp. The functional annotation indicated that 8113 (46.37%), 1933 (11.04%) and 5516 (31.52%) unigenes showed significant similarity to known proteins in the NCBI Nr, Nt and Swiss-Prot databases, respectively. In addition, 2921 (16.70%) unigenes were assigned to KEGG database categories; 4218 (24.11%), to KOG database categories; and 6,603 (37.74%), to GO database categories. Furthermore, we identified 8,798 potential SSRs among 6653 unigenes. A total of 9 polymorphic SSR markers were developed to evaluate the genetic diversity and population structure of 96 isolates collected from Guangdong Province in China. Clonal reproduction of P. polysora in Guangdong was dominant. The YJ (Yangjiang) population had the highest genotypic diversity and the greatest number of the multilocus genotypes, followed by the HY (Heyuan), HZ (Huizhou) and XY (Xinyi) populations. These results provide valuable information for the molecular genetic analysis of P. polysora and related species.