Role of Bordetella pertussis virulence factors in pathogenesis of whooping cough.

1.皮肤坏死毒素（dermonecrotic toxin，DNT）是引起百日咳的一个原因。为了建立一种高灵敏度的检测DNT的方法，我们尝试制备抗DNT作用产生的多胺Rho GTP酶的抗体，并从多胺Rho免疫的家兔中获得血清。然而，血清中，表现出更高的抗体滴度对完整的Rho比多胺之一。然后，我们尝试用兔抗DNT抗体直接ELISA法检测皮肤坏死毒素。该方法可以检测低至0.2 ng/ml的低浓度DNT。使用这种方法，我们确定了10个不同的菌株百日咳博德特氏菌和支气管败血症博德特氏菌的培养上清液中的DNT的浓度。发现培养物上清液中DNT的量随着生物体的增殖而增加，并且对于B.pertussis和B.bronchiseptica分别达到平均5.4 ng/ml和2.0 ng/ml。这些结果表明，DNT存在 ...更多信息 在细菌外环境中以完全有效的浓度存在，并可能影响被博德特氏菌属感染的组织。另外，在上呼吸道组织中寻找靶细胞的过程中，我们发现猪气管平滑肌细胞对DNT敏感。综上所述，我们推测有效剂量的DNT从感染气管中的生物体中释放出来，通过上皮到达平滑肌细胞，并使它们容易收缩，2.博德特氏菌感染体外模型的病理生理学分析我们检测了博德特氏菌感染引起的来自气管上皮的各种细胞系的变化，发现大鼠肺泡上皮L2细胞对实验性感染B.pertussis而非B.bronchiseptica表现出特征性的形态学变化，这意味着L2细胞可用作B.pertussis感染的体外细胞模型。少

1.The function of dermonecrotic toxin(DNT) as a cause of whooping coughTo establish a highly sensitive method detecting DNT, we attempted to generate antibody against polyaminated Rho GTPases, which are yielded by DNT action, and obtained serum from rabbits immunized with polyaminated Rho. The serum, however, showed higher antibody titer against intact Rho than polyaminated one. Then, we attempted to directly detect dermonecrotic toxin by ELISA method, using rabbit anti-DNT antibody. This method made it possible to detect DNT at a low concentration as little as 0.2 ng/ml. Using this method, we determined the concentrations of DNT in culture supernatants of 10 different strains of Bordetella pertussis and B.bronchiseptica. The amounts of DNT in the culture supernatants were found to increase in accordance with proliferation of the organisms and to reach to an average of 5.4 ng/ml and 2.0 ng/ml for B.pertussis and B.bronchiseptica, respectively. These results indicate that DNT is present … More at fully effective concentrations in the extrabacterial milieu and likely affect the tissues which are infected with Bordetella spp. In addition, in the course of searching for target cells in the upper respiratory tissues, we found that tracheal smooth muscle cells from pig are sensitive to DNT. Taken together, we speculate that an effective dose of DNT is liberated from the organisms in the infected trachea, reach to smooth muscle cells through the epithelium, and make them easily constrictive, leading to the paroxysmal cough in whooping cough.2.Pathophysiologic analysis of Bordetella infections in in vitro models.We examined alterations caused by Bordetella infection in various lines of cells derived from the tracheal epithelium and found that rat alveolar epithelial L2 cells showed a characteristic morphological change in response to experimental infection with B.pertussis but not with B.bronchiseptica, implying that L2 cells could be useful as an in vitro cellular model for B.pertussis infection. Less