College of Life Sciences, Capital Normal University, Beijing 100048, China.
Vet Microbiol. 2010 Aug 26;144(3-4):377-83. doi: 10.1016/j.vetmic.2010.01.023. Epub 2010 Feb 1.
Haemophilus parasuis is the causative agent of Glässer's disease. Up to now 15 serovars of H. parasuis have been identified, with significant differences existing in virulence between serovars. In this study, suppression subtractive hybridization (SSH) was used to identify the genetic difference between Nagasaki (H. parasuis serovar 5 reference strain, highly virulent) and SW114 (H. parasuis serovar 3 reference strain, non-virulent). A total of 191 clones were obtained from the SSH library. Using dot hybridization and PCR, 15 clones were identified containing fragments that were present in the Nagasaki genome while absent in the SW114 genome. Among these 15 fragments, three fragments (ssh1, ssh13, ssh15) encode cell surface-associated components; three fragments (ssh2, ssh5, ssh9) are associated with metabolism and stress response; one fragment (ssh8) is involved in assembly of fimbria and one fragment (ssh6) is a phage phi-105 ORF25-like protein. The remaining seven fragments are hypothetical proteins or unknown. Based on PCR analysis of the 15 serovar reference strains, eight fragments (ssh1, ssh2, ssh3, ssh6, ssh8, ssh10, ssh11 and ssh12) were found in three to five of most virulent serovars (1, 5, 10, 12, 13 and 14), zero to two in three moderately virulent serovars (2, 4 and 15), but absent in the low virulent serovar (8) and non-virulent serovars (3, 6, 7, 9 and 11). In vivo transcription fragments ssh1, ssh2, ssh8 and ssh12 were identified in total RNA samples extracted from experimental infected pig lung by RT-PCR. This study has provided some evidence of genetic differences between H. parasuis strains of different virulence.
副猪嗜血杆菌是格拉泽氏病的病原体。迄今为止，已经鉴定出 15 种副猪嗜血杆菌血清型，血清型之间的毒力存在显著差异。在本研究中，采用抑制性消减杂交（SSH）技术来鉴定长崎（副猪嗜血杆菌血清 5 型参考株，高致病性）和 SW114（副猪嗜血杆菌血清 3 型参考株，非致病性）之间的遗传差异。从 SSH 文库中获得了总共 191 个克隆。通过点杂交和 PCR，鉴定出 15 个克隆含有在长崎基因组中存在而在 SW114 基因组中不存在的片段。在这 15 个片段中，有 3 个片段（ssh1、ssh13、ssh15）编码细胞表面相关成分；3 个片段（ssh2、ssh5、ssh9）与代谢和应激反应有关；1 个片段（ssh8）参与菌毛的组装，1 个片段（ssh6）是噬菌体 phi-105 ORF25 样蛋白。其余 7 个片段是假定蛋白或未知蛋白。基于对 15 个血清型参考株的 PCR 分析，在三个到五个最毒力血清型（1、5、10、12、13 和 14）中发现了 8 个片段（ssh1、ssh2、ssh3、ssh6、ssh8、ssh10、ssh11 和 ssh12），在三个中度毒力血清型（2、4 和 15）中发现了零到两个片段，而在低毒力血清型（8）和非毒力血清型（3、6、7、9 和 11）中不存在。通过 RT-PCR 从实验感染猪肺的总 RNA 样品中鉴定出体内转录片段 ssh1、ssh2、ssh8 和 ssh12。本研究为不同毒力副猪嗜血杆菌菌株之间的遗传差异提供了一些证据。
