Pantoea ananatis is an unusual bacterial pathogen that lacks typical virulence determinants yet causes extensive necrosis in onion foliage and bulb tissues. The onion necrosis phenotype is dependent on the expression of the phosphonate toxin, pantaphos, which is synthesized by putative enzymes encoded by the HiVir (high virulence) gene cluster. The genetic contributions of individual hvr genes in HiVir-mediated onion necrosis remain largely unknown, except for the first gene, hvrA (phosphoenolpyruvate mutase, pepM), whose deletion resulted in the loss of onion pathogenicity. In this study, using gene-deletion mutation and complementation, we report that, of the ten remaining genes, hvrB to hvrF are also strictly required for the HiVir-mediated onion necrosis and in-planta bacterial growth, whereas hvrG to hvrJ partially contributed to these phenotypes. As the HiVir gene cluster is a common genetic feature shared among the onion-pathogenic P. ananatis strains that could serve as a useful diagnostic marker of onion pathogenicity, we sought to understand the genetic basis of HiVir-positive yet phenotypically deviant (non-pathogenic) strains. We identified and genetically characterized inactivating single nucleotide polymorphisms in the essential hvr genes of six phenotypically deviant P. ananatis strains. Finally, inoculation of cell-free spent medium of the isopropylthio-β-galactoside (IPTG)-inducible promoter (P_tac)-driven HiVir strain caused P. ananatis–characteristic red onion scale necrosis as well as cell death symptoms in tobacco. Co-inoculation of the spent medium with essential hvr mutant strains restored in-planta populations of the strains to the wild-type level, suggesting that necrotic tissues are important for the proliferation of P. ananatis in onion.

Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Pantoea ananatis是一种不寻常的细菌病原体，缺乏典型的毒力决定因素，但会导致洋葱叶和球茎组织广泛坏死。洋葱坏死表型取决于磷酸盐毒素 pantaphos 的表达，该毒素是由 HiVir（高毒力）基因簇编码的推定酶合成的。除了第一个基因hvrA（磷酸烯醇丙酮酸变位酶，pepM）之外，单个hvr基因在 HiVir 介导的洋葱坏死中的遗传贡献仍然很大程度上未知，该基因的缺失导致洋葱致病性丧失。在这项研究中，利用基因缺失突变和互补，我们报告说，在剩下的 10 个基因中，hvrB到hvrF也是 HiVir 介导的洋葱坏死和植物内细菌生长所严格需要的，而hvrG到hvrJ部分促成了HiVir 介导的洋葱坏死和植物内细菌生长。这些表型。由于 HiVir 基因簇是洋葱致病性P . ananatis菌株可以作为洋葱致病性的有用诊断标记，我们试图了解 HiVir 阳性但表型异常（非致病性）菌株的遗传基础。我们鉴定了六种表型异常的P的必需hvr基因中的失活单核苷酸多态性并进行了遗传表征。菠萝菌株。最后，接种异丙硫基-β-半乳糖苷 (IPTG) 诱导型启动子 (P _tac ) 驱动的 HiVir 菌株的无细胞废培养基引起P . ananatis – 典型的红洋葱鳞片坏死以及烟草中的细胞死亡症状。用过的培养基与必需的hvr突变菌株共同接种使菌株的植物内种群恢复到野生型水平，这表明坏死组织对于P的增殖很重要。洋葱中的菠萝。

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