ABSTRACT

Bordetella (B.) bronchiseptica and Streptococcus (S.) suis are major pathogens in pigs, which are frequently isolated from co-infections in the respiratory tract and contribute to the porcine respiratory disease complex (PRDC). Despite the high impact of co-infections on respiratory diseases of swine (and other hosts), very little is known about pathogen-pathogen-host interactions and the mechanisms of pathogenesis. In the present study, we established a porcine precision-cut lung slice (PCLS) model to analyze the effects of B. bronchiseptica infection on adherence, colonization, and cytotoxic effects of S. suis. We hypothesized that induction of ciliostasis by a clinical isolate of B. bronchiseptica may promote subsequent infection with a virulent S. suis serotype 2 strain. To investigate this theory, we monitored the ciliary activity by light microscopy, measured the release of lactate dehydrogenase, and calculated the number of PCLS-associated bacteria. To study the role of the pore-forming toxin suilysin (SLY) in S. suis-induced cytotoxicity, we included a SLY-negative isogenic mutant and the complemented mutant strain. Furthermore, we analyzed infected PCLS by histopathology, immunofluorescence microscopy, and field emission scanning electron microscopy. Our results showed that pre-infection with B. bronchiseptica promoted adherence, colonization, and, as a consequence of the increased colonization, the cytotoxic effects of S. suis, probably by reduction of the ciliary activity. Moreover, cytotoxicity induced by S. suis is strictly dependent on the presence of SLY. Though the underlying molecular mechanisms remain to be fully clarified, our results clearly support the hypothesis that B. bronchiseptica paves the way for S. suis infection.

摘要

Bordetella (B.) bronchiseptica和Streptococcus (S.) suis是猪的主要病原体，它们经常从呼吸道合并感染中分离出来，并导致猪呼吸道疾病复合体 (PRDC)。尽管共感染对猪（和其他宿主）呼吸道疾病的影响很大，但对病原体-病原体-宿主相互作用和发病机制知之甚少。在本研究中，我们建立了猪精确切割肺切片 (PCLS) 模型来分析支气管败血杆菌感染对猪链球菌的粘附、定植和细胞毒作用的影响。我们假设支气管败血杆菌的临床分离株诱导纤毛停滞可能会促进随后的猪链球菌血清型 2 毒株感染。为了研究这一理论，我们通过光学显微镜监测纤毛活性，测量乳酸脱氢酶的释放，并计算 PCLS 相关细菌的数量。为了研究成孔毒素水溶素 (SLY) 在猪链球菌诱导的细胞毒性中的作用，我们包括了 SLY 阴性等基因突变体和互补突变体菌株。此外，我们通过组织病理学、免疫荧光显微镜和场发射扫描电子显微镜分析了受感染的 PCLS。我们的结果表明，支气管败血杆菌的预感染促进了粘附、定植，并且由于定植增加，猪链球菌的细胞毒作用，可能是由于睫状活动的减少。此外，猪链球菌诱导的细胞毒性严格依赖于 SLY 的存在。尽管潜在的分子机制仍有待完全阐明，但我们的结果清楚地支持了支气管败血杆菌为猪链球菌感染铺平道路的假设。