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首页> 外文期刊>Infection and immunity >Intracellular Growth of Legionella pneumophila Gives Rise to a Differentiated Form Dissimilar to Stationary-Phase Forms
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Intracellular Growth of Legionella pneumophila Gives Rise to a Differentiated Form Dissimilar to Stationary-Phase Forms

机译：嗜肺军团菌的细胞内生长使与静止阶段形式不同的分化形式上升

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When Legionella pneumophila grows in HeLa cells, it alternates between a replicative form and a morphologically distinct “cyst-like” form termed MIF (mature intracellular form). MIFs are also formed in natural amoebic hosts and to a lesser extent in macrophages, but they do not develop in vitro. Since MIFs accumulate at the end of each growth cycle, we investigated the possibility that they are in vivo equivalents of stationary-phase (SP) bacteria, which are enriched for virulence traits. By electron microscopy, MIFs appeared as short, stubby rods with an electron-dense, laminar outer membrane layer and a cytoplasm largely occupied by inclusions of poly-β-hydroxybutyrate and laminations of internal membranes originating from the cytoplasmic membrane. These features may be responsible for the bright red appearance of MIFs by light microscopy following staining with the phenolic Giménez stain. In contrast, SP bacteria appeared as dull red rods after Giménez staining and displayed a typical gram-negative cell wall ultrastructure. Outer membranes from MIFs and SP bacteria were equivalent in terms of the content of the peptidoglycan-bound and disulfide bond cross-linked OmpS porin, although additional proteins, including Hsp60 (which acts as an invasin for HeLa cells), were detected only in preparations from MIFs. Proteomic analysis revealed differences between MIFs and SP forms; in particular, MIFs were enriched for an ～20-kDa protein, a potential marker of development. Compared with SP bacteria, MIFs were 10-fold more infectious by plaque assay, displayed increased resistance to rifampin (3- to 5-fold) and gentamicin (10- to 1,000-fold), resisted detergent-mediated lysis, and tolerated high pH. Finally, MIFs had a very low respiration rate, consistent with a decreased metabolic activity. Collectively, these results suggest that intracellular L. pneumophila differentiates into a cyst-like, environmentally resilient, highly infectious, post-SP form that is distinct from in vitro SP bacteria. Therefore, MIFs may represent the transmissible environmental forms associated with Legionnaires' disease.
机译：当嗜肺军团菌在HeLa细胞中生长时，它在复制形式和形态独特的“囊状”形式（称为MIF（成熟的细胞内形式））之间交替变化。 MIFs也在自然的阿米巴宿主中形成，并在较小程度上在巨噬细胞中形成，但它们不在体外发育。由于MIF在每个生长周期结束时积累，因此我们研究了它们是体内等同于固定相（SP）细菌的可能性，这些细菌具有丰富的毒力特征。通过电子显微镜观察，MIF表现为短而短的棒状，具有电子致密的层状外膜层和胞质，其主要被聚-β-羟基丁酸酯的内含物和源自胞质膜的内膜层合占据。这些特征可能是酚醛Giménez染色后通过光学显微镜观察MIF的鲜红色外观的原因。相反，SP细菌在Giménez染色后显示为暗红色杆，并显示出典型的革兰氏阴性细胞壁超微结构。 MIF和SP细菌的外膜在肽聚糖结合和二硫键交联的OmpS孔蛋白的含量方面是相同的，尽管仅在制剂中检测到了其他蛋白，包括Hsp60（对HeLa细胞的侵袭素）来自MIF。蛋白质组学分析揭示了MIF和SP形式之间的差异。特别是，MIF富含〜20 kDa的蛋白质，这是潜在的发育标志。与SP细菌相比，通过噬菌斑分析，MIF的感染力增加了10倍，显示出对利福平（3至5倍）和庆大霉素（10至1,000倍）的耐药性增加，抵抗去污剂介导的裂解，并耐受高pH 。最后，MIF的呼吸频率非常低，与代谢活性降低相一致。总的来说，这些结果表明细胞内 L。肺炎可分化为一种不同于体外SP细菌的类囊肿，对环境具有抵抗力，高度感染性的SP后形式。因此，MIF可能代表与退伍军人症相关的可传播环境形式。

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