Biofilm formation is a significant cause for the environmental persistence of foodborne pathogens. This phenomenon remains misunderstood in Shigella flexneri whose pathogenicity is mainly associated with the virulence plasmid pWR100. Sequence analysis of the latter predicts a putative lipopolysaccharides (LPS) glycosyltransferase (Gtr) encoded by Sfgtr4, which is the second gene of the SfpgdA-orf186-virK-msbB2 locus. We demonstrated here that purified SfGtr4 exhibited a Gtr activity in vitro by transferring glucose to lipid A. To establish the role of SfGtr4 in virulence, we generated a Sfgtr4 mutant and assessed its phenotype in vitro. Sfgtr4 mutant significantly reduced HeLa cells invasion without impairing type III effectors secretion, increased susceptibility to lysozyme degradation, and enhanced bacterial killing by polymorphonuclear neutrophils (PMNs). SfGtr4 is related to proteins required in biofilm formation. We established conditions whereby wild-type Shigella formed biofilm and revealed that its appearance was accelerated by the Sfgtr4 mutant. Additional phenotypical analysis revealed that single SfpdgA and double SfpgdA-Sfgtr4 mutants behaved similarly to Sfgtr4 mutant. Furthermore, a molecular interaction between SfGtr4 and SfPgdA was identified. In summary, the dual contribution of SfGtr4 and SfPgdA to the pathogenicity and the regulation biofilm formation by S. flexneri was demonstrated here.

中文翻译：

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弗氏志贺氏菌sfgtr4基因的失活诱导生物膜形成并影响细菌致病性。

生物膜形成是食源性病原体在环境中持久存在的重要原因。这种现象在弗氏志 贺氏菌中仍被误解，其致病性主要与毒力质粒pWR100有关。后者的序列分析预测了Sfgtr4编码的推定脂多糖（LPS）糖基转移酶（Gtr），Sfgtr4是SfpgdA-orf186-virK-msbB2基因座的第二个基因。我们在这里证明了纯化的SfGtr4通过将葡萄糖转移至脂质A在体外表现出Gtr活性。为了确定SfGtr4在毒力中的作用，我们生成了Sfgtr4突变体并在体外评估了其表型。sfgtr4突变体显着减少了HeLa细胞的侵袭，而不会损害III型效应子的分泌，增加了对溶菌酶降解的敏感性，并增强了多形核中性粒细胞（PMN）对细菌的杀伤力。SfGtr4与生物膜形成所需的蛋白质有关。我们建立了野生型志贺氏菌形成生物膜的条件，并揭示了Sfgtr4突变体加速了它的出现。进一步的表型分析表明，单个SfpdgA和双SfpgdA - Sfgtr4突变体的行为与Sfgtr4相似突变体。此外，确定了SfGtr4和SfPgdA之间的分子相互作用。总之，SfGtr4和SfPgdA对致病性的双重贡献，并通过调控形成生物膜志贺氏菌在这里展示。