Contamination of edible produce leaves with human bacterial pathogens has been associated with serious disease outbreaks and has become a major public health concern affecting all aspects of the market, from farmers to consumers. While pathogen populations residing on the surface of ready-to-eat produce can be potentially removed through thorough washing, there is no disinfection technology available that effectively eliminates internal bacterial populations. By screening 303 multi-gene deletion (MGD) mutants of Salmonella enterica serovar Typhimurium (STm) 14028s, we were able to identify ten genomic regions that play a role in opening the stomatal pore of lettuce leaves. The major metabolic functions of the deleted regions are associated with sensing the environment, bacterium movement, transport through the bacterial membrane, and biosynthesis of surface appendages. Interestingly, at 21 days post inoculation, seven of these mutants showed increased population titers inside the leaf, two mutants showed similar titers as the wild type bacterium, whereas one mutant with a large deletion that includes the Salmonella pathogenicity island 2 (SPI-2) showed significantly impaired persistence in the leaf apoplast. These findings suggest that not all the genomic regions required for initiation of leaf colonization (i.e., epiphytic behavior and tissue penetration) are essential for continuing bacterial survival as an endophyte. We also observed that mutants lacking either SPI-1 (Mut3) or SPI-2 (Mut9) induce callose deposition levels comparable to those of the wild type STm 14028s; therefore, these islands do not seem to affect this lettuce defense mechanism. However, the growth of Mut9, but not Mut3, was significantly impaired in the leaf apoplastic wash fluid (AWF) suggesting that the STm persistence in the apoplast may be linked to nutrient acquisition capabilities or overall bacterial fitness in this niche, which are dependent on the gene(s) deleted in the Mut9 strain. The genetic basis of STm colonization of leaves investigated in this study provides a foundation from which to develop mitigation tactics to enhance food safety.

中文翻译：

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肠沙门氏菌鼠伤寒血清型 14028s 生菜叶定植所需的基因组区域。

人类细菌病原体对食用农产品叶子的污染与严重疾病的爆发有关，并已成为影响从农民到消费者的市场各个方面的主要公共卫生问题。虽然可以通过彻底清洗去除即食产品表面的病原体种群，但没有可用的消毒技术可以有效消除内部细菌种群。通过筛选鼠伤寒沙门氏菌血清型 (STm) 14028s 的 303 个多基因缺失 (MGD) 突变体，我们能够鉴定出 10 个在打开生菜叶片气孔中发挥作用的基因组区域。缺失区域的主要代谢功能与环境感知、细菌运动、通过细菌膜的运输以及表面附属物的生物合成有关。有趣的是，在接种后 21 天，其中 7 个突变体在叶内显示出群体滴度增加，两个突变体显示出与野生型细菌相似的滴度，而一个突变体具有大缺失，包括沙门氏菌致病性岛 2 (SPI-2)叶质外体的持久性显着受损。这些发现表明，并非叶子定植开始所需的所有基因组区域（即附生行为和组织渗透）对于细菌作为内生菌的持续生存都是必需的。我们还观察到，缺乏 SPI-1 (Mut3) 或 SPI-2 (Mut9) 的突变体诱导的胼胝质沉积水平与野生型 STm 14028s 相当；因此，这些岛屿似乎并没有影响生菜的这种防御机制。然而，Mut9（而非 Mut3）的生长在叶质外体洗涤液 (AWF) 中显着受损，这表明质外体中的 STm 持久性可能与该生态位中的营养获取能力或整体细菌适应性有关，这取决于Mut9 菌株中删除的基因。本研究调查的叶子上 STm 定植的遗传基础为制定缓解策略以增强食品安全奠定了基础。