Koczan Jessica M, McGrath Molly J, Zhao Youfu, Sundin George W
Department of Plant Biology, Michigan State University, East Lansing 48824, USA.
Phytopathology. 2009 Nov;99(11):1237-44. doi: 10.1094/PHYTO-99-11-1237.
Erwinia amylovora is a highly virulent, necrogenic, vascular pathogen of rosaceous species that produces the exopolysaccharide amylovoran, a known pathogenicity factor, and levan, a virulence factor. An in vitro crystal violet staining and a bright-field microscopy method were used to demonstrate that E. amylovora is capable of forming a biofilm on solid surfaces. Amylovoran and levan production deletion mutants were used to determine that amylovoran was required for biofilm formation and that levan contributed to biofilm formation. In vitro flow cell and confocal microscopy were used to further reveal the architectural detail of a mature biofilm and differences in biofilm formation between E. amylovora wild-type (WT), Deltaams, and Deltalsc mutant cells labeled with green fluorescent protein or yellow fluorescent protein. Scanning electron microscopy analysis of E. amylovora WT cells following experimental inoculation in apple indicated that extensive biofilm formation occurs in xylem vessels. However, Deltaams mutant cells were nonpathogenic and died rapidly following inoculation, and Deltalsc mutant cells were not detected in xylem vessels and were reduced in movement into apple shoots. These results demonstrate that biofilm formation plays a critical role in the pathogenesis of E. amylovora.
梨火疫病菌是一种对蔷薇科植物具有高度毒性、致坏死性的维管束病原体，它能产生胞外多糖淀粉果聚糖（一种已知的致病因子）和果聚糖（一种毒力因子）。采用体外结晶紫染色和明场显微镜方法证明梨火疫病菌能够在固体表面形成生物膜。利用淀粉果聚糖和果聚糖产生缺失突变体来确定淀粉果聚糖是生物膜形成所必需的，而果聚糖有助于生物膜形成。体外流动小室和共聚焦显微镜用于进一步揭示成熟生物膜的结构细节以及梨火疫病菌野生型（WT）、Δams和Δlsc突变体细胞（用绿色荧光蛋白或黄色荧光蛋白标记）之间生物膜形成的差异。对苹果进行实验接种后，对梨火疫病菌WT细胞进行扫描电子显微镜分析表明，木质部导管中会大量形成生物膜。然而，Δams突变体细胞无致病性，接种后迅速死亡，并且在木质部导管中未检测到Δlsc突变体细胞，其向苹果嫩枝中的移动也减少。这些结果表明生物膜形成在梨火疫病菌的致病过程中起关键作用。
