Intracellular survival of Burkholderia cenocepacia in macrophages is associated with a delay in the maturation of bacteria-containing vacuoles
Intracellular survival of Burkholderia cenocepacia in macrophages is associated with a delay in the maturation of bacteria-containing vacuoles
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DOI：
10.1111/j.1462-5822.2006.00766.x
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发表时间：
2007-01-01
影响因子：
3.4
通讯作者：
Valvano, Miguel A.
中科院分区：
文献类型：
--
作者：
Lamothe, Julie;Huynh, Kassidy K.;Valvano, Miguel A.
洋葱伯克霍尔德菌复合体 (Bcc) 菌株是机会性细菌，可导致囊性纤维化和慢性肉芽肿病患者危及生命的感染。先前的工作表明，Bcc 分离株可以在阿米巴和巨噬细胞内的膜结合液泡中持续存在，而无需细菌复制，但细菌持续存在的详细机制尚不清楚。在这项研究中，我们研究了新洋葱伯克霍尔德菌菌株 J2315 在 RAW264.7 小鼠巨噬细胞中的存活情况。 J2315 菌株是广泛传播的 ET12 克隆的原型分离株。与内化后不久到达溶酶体的热灭活细菌不同，含有活新洋葱伯克霍尔德杆菌 J2315 的液泡积累晚期内体/溶酶体标记物 LAMP-1，并仅在内化后 6 小时后开始与溶酶体区室融合。使用荧光液相探针，我们还证明了含有新洋葱伯克霍尔德杆菌的液泡继续与新形成的内体相互作用，并维持管腔 pH 值 6.4 +/- 0.12。相比之下，含有热灭活细菌的液泡的平均 pH 值为 4.8 +/- 0.03，并迅速与溶酶体融合。使用液泡 H+-ATP 酶的特异性抑制剂刀那霉素 A 进行的其他实验表明，含有活细菌的液泡并不排除 H+-ATP 酶。这种细菌生存模式不需要III型分泌，因为野生型和III型分泌突变株之间没有发现差异。总的来说，我们的结果表明，细胞内新洋葱伯克霍尔德杆菌会导致吞噬体成熟延迟，这可能有助于细菌逃离宿主细胞的杀菌活性。
Strains of the Burkholderia cepacia complex (Bcc) are opportunistic bacteria that can cause life-threatening infections in patients with cystic fibrosis and chronic granulomatous disease. Previous work has shown that Bcc isolates can persist in membrane-bound vacuoles within amoeba and macrophages without bacterial replication, but the detailed mechanism of bacterial persistence is unknown. In this study, we have investigated the survival of the Burkholderia cenocepacia strain J2315 within RAW264.7 murine macrophages. Strain J2315 is a prototypic isolate of the widespread and transmissible ET12 clone. Unlike heat-inactivated bacteria, which reach lysosomes shortly after internalization, vacuoles containing live B. cenocepacia J2315 accumulate the late endosome/lysosome marker LAMP-1 and start fusing with lysosomal compartments only after 6 h post internalization. Using fluorescent fluid-phase probes, we also demonstrated that B. cenocepacia-containing vacuoles continued to interact with newly formed endosomes, and maintained a luminal pH of 6.4 +/- 0.12. In contrast, vacuoles containing heat-inactivated bacteria had an average pH of 4.8 +/- 0.03 and rapidly merged with lysosomes. Additional experiments using concanamycin A, a specific inhibitor of the vacuolar H+-ATPase, revealed that vacuoles containing live bacteria did not exclude the H+-ATPase. This mode of bacterial survival did not require type III secretion, as no differences were found between wild type and a type III secretion mutant strain. Collectively, our results suggest that intracellular B. cenocepacia cause a delay in the maturation of the phagosome, which may contribute to facilitate bacterial escape from the microbicidal activities of the host cell.