Gabay J E, Horwitz M A
J Exp Med. 1985 Feb 1;161(2):409-22. doi: 10.1084/jem.161.2.409.
Legionella pneumophila, the etiologic agent of Legionnaires' disease, is phagocytized in an unusual way and multiplies in human mononuclear phagocytes in a novel phagosome. As a first step toward understanding these L. pneumophila-phagocyte interactions, we have studied the envelope of L. pneumophila Philadelphia 1 strain. We isolated cell envelopes by treating whole bacterial cells with lysozyme and EDTA to convert them to spheroplasts, then lysing the spheroplasts osmotically or sonically. We resolved the cell envelopes into two membrane fractions by isopycnic centrifugation. We localized NADH oxidase to the fraction of buoyant density 1.145, which we designated cytoplasmic membrane, and lipopolysaccharide (LPS) to the fraction of density 1.222, which we designated outer membrane. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed that the L. pneumophila outer membrane contains a single major protein species migrating at 28,000 mol wt; this is the major protein of the bacterium. The cytoplasmic membrane also contains a single major protein species migrating at 65,000 mol wt. Surface iodination of the bacteria and agglutination and immunofluorescence studies with rabbit antibody produced against the purified major outer membrane protein (MOMP) revealed that this protein is exposed at the cell surface. We isolated LPS from L. pneumophila membranes by SDS-EDTA treatment. The pattern obtained by subjecting the LPS to SDS-PAGE and staining the gel with silver nitrate suggests that L. pneumophila LPS might be atypical. We studied patient serologic responses to cell envelope components of L. pneumophila Philadelphia 1, a serogroup 1 organism. Sera from patients with evidence of infection with serogroup 1 L. pneumophila contained large amounts of antibody to this strain. Few of these antibodies recognized the MOMP of L. pneumophila. In contrast, greater than 98% of these antibodies were directed against the LPS. This indicates that LPS is the dominant serogroup antigen and the major antigen responsible for the reactivity of patient sera in the indirect fluorescent antibody assay, currently the principal diagnostic assay for Legionella infection.
嗜肺军团菌是军团病的病原体，它以一种不同寻常的方式被吞噬，并在一种新型吞噬体中的人类单核吞噬细胞内繁殖。作为了解这些嗜肺军团菌 - 吞噬细胞相互作用的第一步，我们研究了嗜肺军团菌费城1菌株的包膜。我们通过用溶菌酶和乙二胺四乙酸（EDTA）处理完整细菌细胞使其转化为原生质球，然后通过渗透压或超声处理裂解原生质球来分离细胞膜。我们通过等密度离心将细胞膜分离为两个膜组分。我们将NADH氧化酶定位到浮力密度为1.145的组分，我们将其指定为细胞质膜，将脂多糖（LPS）定位到密度为1.222的组分，我们将其指定为外膜。十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳（SDS - PAGE）显示，嗜肺军团菌外膜含有一种主要的蛋白质，其分子量为28,000；这是该细菌的主要蛋白质。细胞质膜也含有一种主要的蛋白质，其分子量为65,000。对细菌进行表面碘化以及用针对纯化的主要外膜蛋白（MOMP）产生的兔抗体进行凝集和免疫荧光研究表明，该蛋白暴露于细胞表面。我们通过SDS - EDTA处理从嗜肺军团菌膜中分离出LPS。对LPS进行SDS - PAGE并用硝酸银染色凝胶得到的图谱表明，嗜肺军团菌LPS可能是非典型的。我们研究了患者对嗜肺军团菌费城1（血清群1菌株）细胞膜成分的血清学反应。有血清群1嗜肺军团菌感染证据的患者血清中含有大量针对该菌株的抗体。这些抗体中很少有能识别嗜肺军团菌的MOMP。相比之下，超过98%的这些抗体针对的是LPS。这表明LPS是主要的血清群抗原，也是目前军团菌感染主要诊断检测方法——间接荧光抗体检测中患者血清反应性的主要抗原。
