Abstract：
Opportunistic pathogens (OPs) are of concern in drinking water distribution systems because they persist despite disinfectant residuals. While many OPs garner protection from disinfectants via a biofilm lifestyle, Legionella pneumophila (Lp) also gains disinfection resistance by being harbored within free-living amoebae (FLA). It has been long established, but poorly understood, that Lp grown within FLA show increased infectivity toward subsequent FLA or human cells (i.e., macrophage), via a process we previously coined \"protozoan-priming\". The objectives of this study are (i) to identify in Lp a key genetic determinant of how protozoan-priming increases its infectivity, (ii) to determine the chemical stimulus within FLA to which Lp responds during protozoan-priming, and (iii) to determine if more infectious forms of Lp also exhibit enhanced disinfectant resistance. Using Acanthamoeba castellanii as a FLA host, the priming effect was isolated to Lp\'s sidGV locus, which is activated upon sensing elevated magnesium concentrations. Supplementing growth medium with 8 mM magnesium is sufficient to produce Lp grown in vitro with an infectivity equivalent to that of Lp grown via the protozoan-primed route. Both Lp forms with increased infectivity (FLA-grown and Mg2+-supplemented) exhibit greater monochloramine resistance than Lp grown in standard media, indicating that passage through FLA not only increases Lp\'s infectivity but also enhances its monochloramine resistance. Therefore, laboratory-based testing of disinfection strategies should employ conditions that simulate or replicate intracellular growth to accurately assess disinfectant resistance.
摘要：
机会病原体（OP）在饮用水分配系统中受到关注，因为尽管有消毒剂残留，它们仍会持续存在。虽然许多OPs通过生物膜生活方式获得消毒剂的保护，嗜肺军团菌（Lp）也通过在自由生活的变形虫（FLA）中携带而获得消毒抗性。它早已建立,但却知之甚少,在FLA中生长的Lp对随后的FLA或人类细胞显示出增加的感染性(即，巨噬细胞)，通过我们以前创造的“原生动物启动”的过程。这项研究的目的是（i）在Lp中确定原生动物启动如何增加其感染性的关键遗传决定因素，（ii）确定在原生动物引发期间Lp响应的FLA内的化学刺激，和(iii)确定更多感染性形式的Lp是否也表现出增强的消毒剂抗性。使用棘阿米巴作为FLA宿主，启动效应分离到Lp的sidGV基因座，它在感应到升高的镁浓度时被激活。补充8mM镁的生长培养基足以产生体外生长的Lp，其感染性等同于通过原生动物引发的途径生长的Lp。与标准培养基中生长的Lp相比，感染性增加的两种Lp形式（FLA生长和补充Mg2的）均表现出较高的一氯胺抗性。表明通过FLA不仅增加了Lp的感染性，而且增强了其对一氯胺的抗性。因此,基于实验室的消毒策略测试应采用模拟或复制细胞内生长的条件，以准确评估消毒剂抗性。
