Fig. 2

Targeted gene replacement of MoXYL1A compromised turgor-mediated appressorium integrity and impaired the virulence of M. oryzae. a Showed hyphae-mediated virulence characteristics of the individual strains on intact and injured leaves of one-week-old, barley seedlings.induction of blast lesion was assessed at 7-dpi. Images are representative of three independent assays, each assay with three replicates. b Virulence bioassay conducted on 21-days-old rice seedlings using spore-drop inoculation method. Conidial suspensions were prepared as 1 × 10⁵ conidia mL⁻¹ in 0.2% Tween 20, for both the mutants and wild-type 20 µL. c Conidia-mediated pathogenicity/virulence characteristics ΔMoxyl1A, ΔMoxyl1B, and the wild-type on 21-days-old rice seedlings, through spray-inoculation with conidial suspensions (1 × 10⁵ conidia mL⁻¹ in 0.2% Tween 20). d The micrograph portrays the inability of ΔMoxyl1A and double deletion mutants to invade and colonize barley tissue at 48hpi. The absence of invasive hyphae were evident in barley cells inoculated with ΔMoxyl1A or the double deletion strain. In contrast, numerous invasive hyphae were seen in the leaves inoculated with either ΔMoxyl1B or Guy11. Images are representative of n = 2 independent biological replicates. Scale bar, 20 μm. e Appressoria were produced artificially on Thermo-fisher hydrophobic coverslips and observed at 8 hpi. Images show non-functional appressoria lacking melanin lining for the ΔMoxyl1A mutant. f Showed results from incipient cytorrhysis assays performed to evaluate the turgidity of appressorium form by conidia from the ΔMoxyl1A, ΔMoxyl1B, ΔMoxyl1A_Com., ΔMoxyl1B_Com., and the wild-type hydrophobic coverslips for 8-h appressorium formed were treated with 2 M glycerol solutions. Collapse appressorium were counted using the Olympus DP80 light microscope. Scale bar = 20 μm. g The bar graph showed results from the statistical evaluation of the proportion of collapsed appressorium recorded in the ΔMoxyl1A, ΔMoxyl1B, ΔMoxyl1A_Com., ΔMoxyl1B_Com., and the wild-type strains on hydrophobic coverslips for 8-h and treated independently with varying concentrations (1 M, 2 M, 3 M, and 4 M) of glycerol solutions during incipient cytorrhysis assays. Consistent results from three independent biological experiments with each consisting of five technical replicates were used for statistical analyses. For each independent biological experiment, 100 appressoria were counted (n = 100*3). treatments yielding significant difference (P ≤ 0.05) are denoted with asterisks “*”