Root cortical anatomy is associated with differential pathogenic and symbiotic fungal colonization in maize.

Root anatomical phenotypes vary among maize (Zea mays) cultivars and may have adaptive value by modifying the metabolic cost of soil exploration. However, the microbial trade-offs of these phenotypes are unknown. We hypothesized that nodal roots of maize with contrasting cortical anatomy have different patterns of mutualistic and pathogenic fungal colonization. Arbuscular mycorrhizal colonization in the field and mesocosms, root rots in the field, and Fusarium verticillioides colonization in mesocosms were evaluated in maize genotypes with contrasting root cortical anatomy. Increased aerenchyma and decreased living cortical area were associated with decreased mycorrhizal colonization in mesocosm and field experiments with inbred genotypes. In contrast, mycorrhizal colonization of hybrids increased with larger aerenchyma lacunae; this increase coincided with larger root diameters of hybrid roots. F. verticillioides colonization was inversely correlated with living cortical area in mesocosm-grown inbreds, and no relation was found between root rots and living cortical area or aerenchyma in field-grown hybrids. Root rots were positively correlated with cortical cell file number and inversely correlated with cortical cell size. Mycorrhizae and root rots were inversely correlated in field-grown hybrids. We conclude that root anatomy is associated with differential effects on pathogens and mycorrhizal colonization of nodal roots in maize.

Deoxynivalenol biosynthesis-related gene expression during wheat kernel colonization by Fusarium graminearum.

Deoxynivalenol (DON) is a potent mycotoxin and virulence factor produced by Fusarium graminearum. We examined the expression of the core DON biosynthetic gene Tri5 during wheat head infection of susceptible and resistant cultivars and susceptible cultivars treated with strobilurin fungicides (e.g., azoxystrobin). DON was quantified to correlate expression with toxin accumulation. The highest Tri5 expression relative to housekeeping genes occurred at the infection front. As infection progressed, earliest-infected kernels showed diminished relative Tri5 expression but Tri5 expression never ceased during the 21 days observed. Azoxystrobin treatment showed no significant effect on either relative Tri5 expression or DON quantity. The resistant cultivar 'Alsen' showed minimal spread of the fungus, with no fungus detected by day 21. DON was not detected in significant quantities in Alsen in the later stages sampled. In Wheaten, DON levels were negligible at 8 days postinoculation (dpi), with detectable DON at later-sampled time points. Tri5 was detected even in fully senesced kernels 21 dpi. Our data demonstrate the presence of Tri5 transcripts in a susceptible cultivar over a much longer time period than has been previously documented. This suggests the ability of the fungus to rapidly resume toxin biosynthesis in dried infected grain should conducive environmental conditions be present, and provides a possible mechanism for high DON levels in asymptomatic grain.

Population Genetic Structure and Chemotype Diversity of Fusarium graminearum Populations from Wheat in Canada and North Eastern United States.

Fusarium head blight (FHB) is a major disease in wheat causing severe economic losses globally by reducing yield and contaminating grain with mycotoxins. In Canada, Fusarium graminearum is the principal etiological agent of FHB in wheat, producing mainly the trichothecene mycotoxin, deoxynivalenol (DON) and its acetyl derivatives (15-acetyl deoxynivalenol (15ADON) and 3-acetyl deoxynivalenol (3ADON)). Understanding the population biology of F. graminearum such as the genetic variability, as well as mycotoxin chemotype diversity among isolates is important in developing sustainable disease management tools. In this study, 570 F. graminearum isolates collected from commercial wheat crops in five geographic regions in three provinces in Canada in 2018 and 2019 were analyzed for population diversity and structure using 10 variable number of tandem repeats (VNTR) markers. A subset of isolates collected from the north-eastern United States was also included for comparative analysis. About 75% of the isolates collected in the Canadian provinces of Saskatchewan and Manitoba were 3ADON indicating a 6-fold increase in Saskatchewan and a 2.5-fold increase in Manitoba within the past 15 years. All isolates from Ontario and those collected from the United States were 15ADON and isolates had a similar population structure. There was high gene diversity (H = 0.803-0.893) in the F. graminearum populations in all regions. Gene flow was high between Saskatchewan and Manitoba (Nm = 4.971-21.750), indicating no genetic differentiation between these regions. In contrast, less gene flow was observed among the western provinces and Ontario (Nm = 3.829-9.756) and USA isolates ((Nm = 2.803-6.150). However, Bayesian clustering model analyses of trichothecene chemotype subpopulations divided the populations into two clusters, which was correlated with trichothecene types. Additionally, population cluster analysis revealed there was more admixture of isolates among isolates of the 3ADON chemotypes than among the 15ADON chemotype, an observation that could play a role in the increased virulence of F. graminearum. Understanding the population genetic structure and mycotoxin chemotype variations of the pathogen will assist in developing FHB resistant wheat cultivars and in mycotoxin risk assessment in Canada.

Development of a Real-Time RT-PCR SYBR Green Assay for Tomato ring spot virus in Grape.

Grapevines infected with Tomato ring spot virus (ToRSV) pose an economic risk for growers in the northeastern United States. This study describes a one-step real-time reverse-transcription polymerase chain reaction (RT-PCR) SYBR Green assay for detecting ToRSV in grapevines. Two newly designed primer pairs based on the ToRSV coat protein gene sequence were evaluated for specificity and optimized for a SYBR Green assay. The primer pair ToRSV1f/1r yielded a 130-bp product with strong primer-dimer products, whereas the primer pair ToRSV2f/2r yielded a 330-bp product with weak primer dimer products. Real-time RT-PCR detected ToRSV in more naturally infected grapevines maintained in the greenhouse than did enzyme-linked immunosorbent assay. The nucleotide sequences of the fragments amplified from grapevine growing in Pennsylvania using real-time PCR were divergent from previously published sequences.

FvSO regulates vegetative hyphal fusion, asexual growth, fumonisin B1 production, and virulence in Fusarium verticillioides.

Hyphal anastomosis is a hallmark of filamentous fungi and plays vital roles including cellular homoeostasis, interhyphal communication and nutrient translocation. Here we identify a gene, FvSO, in Fusarium verticillioides, a filamentous ascomycete causing maize ear and stalk rot and producing fumonisin mycotoxins. FvSO, like its Neurospora crassa homologue SO, is required for vegetative hyphal fusion. It is also essential for normal vegetative growth, sporulation, and pathogenesis. FvSO encodes a predicted WW domain protein and shares 70 % protein sequence identity with N. crassa SO. FvSO deletion mutants (ΔFvSO) had abnormal distribution of conidia size, and conidia of ΔFvSO germinated much later and slower than wild type. ΔFvSO was deficient in hyphal anastomosis, had slower radial growth and produced less fungal biomass than wild type. ΔFvSO were unable to perform anastomosis, a key feature of filamentous fungi. Interestingly, production of fumonisin B1 by ΔFvSO was significantly reduced compared to wild type. Additionally, ΔFvSO was nonpathogenic to corn ears, stalks and seedlings, likely due to defective growth and development. In conclusion, FvSO is essential for vegetative hyphal fusion and is required for normal vegetative growth and sporulation, normal levels of fumonisin production and pathogenicity in F. verticillioides. The pleiotropic nature of ΔFvSO phenotypes suggests that FvSO is likely involved in certain signalling pathways that regulate multiple cellular functions.

Induced systemic resistance responses in perennial ryegrass against Magnaporthe oryzae elicited by semi-purified surfactin lipopeptides and live cells of Bacillus amyloliquefaciens.

The suppressive ability of several strains of cyclic lipopeptide-producing Bacillus rhizobacteria to grey leaf spot disease caused by Magnaporthe oryzae has been documented previously; however, the underlying mechanism(s) involved in the induced systemic resistance (ISR) activity in perennial ryegrass (Lolium perenne L.) remains unknown. Root-drench application of solid-phase extraction (SPE)-enriched surfactin and live cells of mutant Bacillus amyloliquefaciens strain FZB42-AK3 (produces surfactin, but not bacillomycin D and fengycin) significantly reduced disease incidence and severity on perennial ryegrass. The application of the treatments revealed a pronounced multilayered ISR defence response activation via timely and enhanced accumulation of hydrogen peroxide (H2O2), elevated cell wall/apoplastic peroxidase activity, and deposition of callose and phenolic/polyphenolic compounds underneath the fungal appressoria in naïve leaves, which was significantly more intense in treated plants than in mock-treated controls. Moreover, a hypersensitive response (HR)-type reaction and enhanced expression of LpPrx (Prx, peroxidase), LpOXO4 (OXO, oxalate oxidase), LpPAL (PAL, phenylalanine ammonia lyase), LpLOXa (LOX, lipoxygenase), LpTHb (putative defensin) and LpDEFa (DEFa, putative defensin) in perennial ryegrass were associated with SPE-enriched surfactin and live AK3 cell treatments, acting as a second layer of defence when pre-invasive defence responses failed. The results indicate that ISR activity following surfactin perception may sensitize H2O2 -mediated defence responses, thereby providing perennial ryegrass with enhanced protection against M. oryzae.