Nanopore and Illumina Genome Sequencing of Fusarium oxysporum f. sp. lini Strains of Different Virulence.

Abstract

Fusarium oxysporum f. sp. lini is the one of notoriously known pathogens of flax (Linum usitatissimum L.), causing wilt. In the case of young seedlings, the disease may lead to complete yield losses (Kommedahl et al., 1970). At the same time, flax is an important crop used for manufacturing oil of high nutritional value, food supplements, industrial products, and fiber (Jhala and Hall, 2010; Czemplik et al., 2011; Kezimana et al., 2018; Parikh and Pierce, 2019). However, flax production is dependent on the resistance of flax varieties to phytopathogens, whereas F. oxysporum demonstrates considerable genetic diversity (Edel et al., 2001; Michielse and Rep, 2009). At present, phytopathogenic strains of F. oxysporum are classified into numerous formae speciales according to their ability to colonize different hosts (Armstrong and Armstrong, 1981). Unfortunately, this system cannot provide researchers with enough information on a type of the pathogen and the severity of the infection it causes (Edel-Hermann and Lecomte, 2019). Like other representatives of the species, forma specialis lini is morphologically and genetically heterogeneous. Strains of the pathogen differ in such characteristics as the type of sporulation, conidia formation, pigment production, and the rate of growth on different media. Moreover, the degree of pathogenicity varies within the forma and depends on the infected crop variety (Kommedahl et al., 1970). For molecular classification of F. oxysporum, numerous markers were used (Baayen et al., 2000; Lievens et al., 2008; Baysal et al., 2010; Sharma et al., 2014; van Dam et al., 2018; Srinivas et al., 2019; Sasseron et al., 2020), and genes associated with virulence were considered as targets for molecular discrimination of strains of the fungus (Lievens et al., 2008; van Dam et al., 2018). It was shown that secreted in xylem (SIX) genes are associated with pathogenicity of F. oxysporum, and the majority of them are distributed within the sequence of one chromosome (Rep et al., 2004; Houterman et al., 2007; Kashiwa et al., 2017; Carvalhais et al., 2019). Importantly, the combination of these genes differs between and within formae (Lievens et al., 2009). Secreted in xylem are also responsible for the resistance of cultivars to certain pathogen races due to gene-for-gene interaction between SIX genes of a pathogen and R genes (resistance genes) of a plant. Breaking the resistance of plant lineages to wild-type F. oxysporum could be accomplished by deletion of a certain SIX gene, which is recognized by the immune system of a plant (Houterman et al., 2008). However, in the case of F. oxysporum f. sp. lini, the connection between the degree of pathogenicity of a strain and the set of its SIX genes is not investigated deeply, and gaining insights into the mechanisms of pathogenicity offers an opportunity for effective disease control. Besides, F. oxysporum representatives differ not only in the content of genes and their sequences but also in the number of chromosomes due to chromosomal rearrangements and the mobility of lineage-specific chromosomes (Daviere et al., 2001; Ma et al., 2010; Schmidt et al., 2013; Vlaardingerbroek et al., 2016; Wang et al., 2020). Having learned the statistics of deposited F. oxysporum assemblies, one may conclude that genome sizes vary from about 50 to 70 Mb and differ between many formae (data of the NCBI Genome database, https://www.ncbi.nlm.nih.gov/genome/browse/#!/eukaryotes/707/). Summing up, F. oxysporum f. sp. lini appears to be heterogeneous. However, there is a lack of next-generation sequencing data concerning the genome structure of the flax pathogen and the molecular basis of pathogenicity in relation to diverse virulence of the strains. In this work, we have chosen six strains of F. oxysporum f. sp. lini of low, medium, and high virulence (two strains per each degree of pathogenicity), performed genome sequencing on Oxford Nanopore Technologies (ONT) and Illumina platforms, and obtained de novo assemblies of the sequenced strains.