The AmyR regulon in Erwinia amylovora. Erwinia amylovora

Microarray analysis was used to identify genes that were controlled by AmyR in minimal and on immature pear fruits. Consistent with amylovoran production, an inverse correlation was observed between amyR expression and the expression level of amylovoran biosynthetic genes in liquid media. Interestingly, over-expression of AmyR suppressed the expression of type III secretion system genes including hrpA, hrpN and dspEF after pear fruit infection. Consistent with levan production and swarming motility, levasucrase and flagellar genes were both down-regulated both in the amyR mutant and over-expression strains in liquid media. Together, our results suggest that AmyR plays an important role in regulating bacterial exopolysaccharide production and virulence in E. amylovora. Overall design: A total of 14 samples were analyzed in two conditions: For the in vitro condition (MBMA medium + 1% sorbitol), Erwinia amylovora wild type strain (2 replicates); E. amylovora amyR mutant strain (3 replicates); E. amylovora amyR over-expression strain (3 replicates); For the in vivo condition (on wounded immature pear fruits), Erwinia amylovora wild type strain (2 replicates); E. amylovora amyR mutant strain (2 replicates); E. amylovora amyR over-expression strain (2 replicates).

本研究采用微阵列（Microarray）分析技术，筛选出在基本培养基及未成熟梨果实上受AmyR调控的基因。与淀粉样多糖（amylovoran）的合成规律一致，在液体培养基中，amyR的表达量与淀粉样多糖生物合成基因的表达水平呈负相关。值得注意的是，在梨果实侵染实验中，AmyR过表达会抑制III型分泌系统（type III secretion system）相关基因（包括hrpA、hrpN及dspEF）的表达。与果聚糖（levan）合成及群集运动特性相符，在液体培养基中，amyR敲除突变株与AmyR过表达菌株内的果聚糖蔗糖酶（levasucrase）编码基因及鞭毛相关基因均出现表达下调。综上，本研究结果表明，AmyR在解淀粉欧文氏菌（Erwinia amylovora, E. amylovora）的胞外多糖合成与毒力调控过程中发挥重要作用。
实验设计概述：本研究共分析14份样本，分为两种实验条件：
1. 体外培养条件（MBMA培养基+1%山梨醇）：解淀粉欧文氏菌野生型菌株（2个生物学重复）、解淀粉欧文氏菌amyR敲除突变株（3个生物学重复）、解淀粉欧文氏菌AmyR过表达菌株（3个生物学重复）；
2. 体内侵染条件（接种于损伤的未成熟梨果实）：解淀粉欧文氏菌野生型菌株（2个生物学重复）、解淀粉欧文氏菌amyR敲除突变株（2个生物学重复）、解淀粉欧文氏菌AmyR过表达菌株（2个生物学重复）。