Alternaria alternata can resist high levels of reactive oxygen species (ROS). The protective roles of autophagy or autophagy-mediated degradation of peroxisomes (termed pexophagy) against oxidative stress remain unclear. The present study, using transmission electron microscopy and fluorescence microscopy coupled with a GFP-AaAtg8 proteolysis assay and an mCherry tagging assay with peroxisomal targeting tripeptides, demonstrated that hydrogen peroxide (H 2 O 2 ) and nitrogen depletion induced autophagy and pexophagy. Experimental evidence showed that H 2 O 2 triggered autophagy and the translocation of peroxisomes into the vacuoles. Mutational inactivation of the AaAtg8 gene in A. alternata led to autophagy impairment, resulting in the accumulation of peroxisomes, increased ROS sensitivity, and decreased virulence.Compared to the wild type, ΔAaAtg8 failed to detoxify ROS effectively, leading to ROS accumulation. Deleting AaAtg8 down-regulated the expression of genes encoding an NADPH oxidase and a Yap1 transcription factor, both involved in ROS resistance.Deleting AaAtg8 affected the development of conidia and appressorium-like structures. Deleting AaAtg8 also compromised the integrity of the cell wall. Reintroduction of a functional copy of AaAtg8 in the mutant completely restored all defective phenotypes. Although ΔAaAtg8 produced wild-type toxin levels in axenic culture, the mutant induced a lower level of H 2 O 2 and smaller necrotic lesions on citrus leaves. In addition to H 2 O 2 , nitrogen starvation triggered peroxisome turnover. We concluded that ΔAaAtg8 failed to degrade peroxisomes effectively, leading to the accumulation of peroxisomes and the reduction of the stress response. Autophagy-mediated peroxisome turnover could increase cell adaptability and survival under oxidative stress and starvation conditions.
Dickeya spp. cause severe diseases in many crops. Most previous studies in Taiwan identified these pathogens as a single species (Erwinia chrysanthemi), and little is known about their genetic and phenotypic diversity. This study collected 40 Dickeya strains isolated from different host plants in Taiwan and conducted a series of phylogenetic and phenotypic analyses. Reconstruction of maximum likelihood trees revealed that the isolated strains belonged to D. dadantii, D. chrysanthemi, D. undicola, and D. fangzhongdai. Among the 40 tested strains, 32 collected from Phalaenopsis orchids and Welsh onions were classified as D. fangzhongdai, while those isolated from the other hosts were assigned to D. dadantii, D. chrysanthemi, and D. undicola, suggesting that some of these bacteria exhibit host preferences. Inoculation of representative strains of the four Dickeya species onto potato, Phalaenopsis and African violet showed that the maceration potentials varied inter‐ and intra‐specifically and that the differential infection patterns were host‐dependent. Phenotypic assays also revealed that strain‐level variation in maceration potential was associated (in part) with the pathogens' tolerance to hostile pH and temperature or regulation of indigoidine production. Although comparisons of colony morphology and pigment production were not sufficient to differentiate Dickeya species, Biolog analyses identified several nutrients and chemicals potentially capable of differentiating among different species. Overall, our data revealed the genetic diversity and phenotypic characteristics of Dickeya spp. in Taiwan and provided information useful for their identification. The study also presented the first evidence showing that D. undicola, a recently identified species inhabiting surface waters, could naturally infect plants.
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