Department of Microbiology and Molecular Genetics, Oklahoma State University, Stillwater, Oklahoma 74078, USA
Vector Borne Zoonotic Dis. 2011 Jul;11(7):917-22. doi: 10.1089/vbz.2010.0126. Epub 2011 Jan 22.
Q fever, a zoonotic disease, is caused by a gram-negative intracellular bacterium, Coxiella burnetii. Although normally transmitted during exposure to infectious aerosols, C. burnetii is also found in arthropod vectors. In the environment, ticks are thought to play a crucial role in bacterial maintenance and transmission by infecting various mammalian species. However, the nature of the pathogen-tick relationship is not well defined. To determine C. burnetii's interactions with a cultured tick cell line, we introduced purified C. burnetii NMII into Ixodes scapularis-derived IDE8 cells and assayed for bacterial presence, replication, gene expression, and subsequent infectivity for mammalian cells. Tick cells were harvested at 24 h, 72 h, 7 days, and 11 days postinfection (PI). C. burnetii uptake and subsequent replication was demonstrated by indirect immunofluorescence assay, electron microscopy, and real-time polymerase chain reaction (PCR). When a genome equivalent multiplicity of infection of 30 was used, 30%-40% of exposed cells were seen to have small, rounded, vacuoles at 72 h PI, whereas at 7 and 11 days PI, 60%-70% of cells contained enlarged vacuoles harboring large numbers of bacteria. Quantitative PCR analysis of total genomic DNA confirmed that C. burnetii genome numbers increased significantly from 24 h to 11 days PI. Expression of C. burnetii type four secretion system homologs at 7 days PI was demonstrated by reverse transcriptase PCR. Finally, indirect immunofluorescence assay demonstrated that C. burnetii propagated within IDE8 cells were infectious for mammalian cells. These studies demonstrate the utility of cultured tick cell lines as a model to investigate C. burnetii's molecular interactions with its arthropod vectors.
Q 热是一种人畜共患病，由革兰氏阴性细胞内细菌伯氏考克斯体引起。虽然正常情况下是在接触传染性气溶胶时传播，但伯氏考克斯体也存在于节肢动物载体中。在环境中，蜱虫被认为通过感染各种哺乳动物物种在细菌的维持和传播中起着至关重要的作用。然而，病原体与蜱虫的关系性质尚不清楚。为了确定伯氏考克斯体与培养的蜱细胞系的相互作用，我们将纯化的伯氏考克斯体 NMII 引入到 Ixodes scapularis 衍生的 IDE8 细胞中，并检测细菌的存在、复制、基因表达以及随后对哺乳动物细胞的感染性。在感染后 24 小时、72 小时、7 天和 11 天收获蜱细胞。通过间接免疫荧光法、电子显微镜和实时聚合酶链反应（PCR）证明了伯氏考克斯体的摄取和随后的复制。当使用 30 个基因组当量的感染复数时，在 72 小时 PI 时，30%-40%的暴露细胞被观察到有小的、圆形的空泡，而在 7 天和 11 天 PI 时，60%-70%的细胞含有含有大量细菌的扩大空泡。总基因组 DNA 的定量 PCR 分析证实，从 24 小时到 11 天 PI，伯氏考克斯体基因组数量显著增加。在 7 天 PI 时通过逆转录 PCR 证明了伯氏考克斯体四种分泌系统同源物的表达。最后，间接免疫荧光法证明在 IDE8 细胞内增殖的伯氏考克斯体对哺乳动物细胞具有感染性。这些研究表明，培养的蜱细胞系可作为一种模型，用于研究伯氏考克斯体与节肢动物载体的分子相互作用。
