Rajeev Sreekumari, Nair Rajeev V, Kania Stephen A, Bemis David A
Animal Disease Diagnostic Laboratory, Ohio Department of Agriculture, 8995 East Main Street, Reynoldsburg, OH 43068, USA.
Vet Microbiol. 2003 Jul 30;94(4):313-23. doi: 10.1016/s0378-1135(03)00137-8.
Mild or subclinical respiratory infections caused by Bordetella bronchiseptica are widespread in pigs despite multiple control efforts. Infection with virulent B. bronchiseptica strains is a common risk factor in the establishment of toxin-producing strains of Pasteurella multocida in the nasal cavity of pigs leading to the disease, atrophic rhinitis (AR). This study was designed to explore the possibility of expressing a protective epitope of P. multocida toxin (PMT) in B. bronchiseptica to create single-component mucosal vaccine to control atrophic rhinitis in pigs. To achieve this, a P. multocida toxin fragment (PMTCE), that was non-toxic and protective against lethal challenge in mice, was cloned into a broad-host-range plasmid, PBBR1MCS2, and introduced into B. bronchiseptica by electroporation. The Pasteurella gene construct was placed under the regulatory control of a promoter region that was separately isolated from B. bronchiseptica and appears to be part of the heat shock protein gene family. B. bronchiseptica harboring the plasmid under antibiotic selection expressed the 80kDa PMTCE as determined by PAGE and Western blot with a PMT-specific monoclonal antibody. When introduced into the respiratory tracts of mice, B. bronchiseptica harboring the plasmid construct was reisolated in declining numbers for 72h post-inoculation. Antibody responses (IgM, IgA and IgG) to B. bronchiseptica were detected in serum and respiratory lavage, but PMTCE-specific antibodies were not detected. While further refinements of PMT expression in B. bronchiseptica are necessary, this study provides a basis for the development of a single-component, live-attenuated vaccine against atrophic rhinitis.
尽管采取了多种防控措施，但支气管败血波氏杆菌引起的轻度或亚临床呼吸道感染在猪群中仍广泛存在。感染强毒支气管败血波氏杆菌菌株是猪鼻腔中多杀性巴氏杆菌产毒株定植并导致萎缩性鼻炎（AR）疾病的常见危险因素。本研究旨在探索在支气管败血波氏杆菌中表达多杀性巴氏杆菌毒素（PMT）保护性表位以制备单组分黏膜疫苗来防控猪萎缩性鼻炎的可能性。为此，将无毒且能保护小鼠抵御致死性攻击的多杀性巴氏杆菌毒素片段（PMTCE）克隆到广宿主范围质粒PBBR1MCS2中，并通过电穿孔法导入支气管败血波氏杆菌。多杀性巴氏杆菌基因构建体置于从支气管败血波氏杆菌中单独分离出的启动子区域的调控之下，该启动子区域似乎是热休克蛋白基因家族的一部分。在抗生素选择压力下，携带该质粒的支气管败血波氏杆菌通过PAGE和用PMT特异性单克隆抗体进行的Western印迹法测定表达了80kDa的PMTCE。将携带质粒构建体的支气管败血波氏杆菌引入小鼠呼吸道后，接种后72小时内其重新分离的数量逐渐减少。在血清和呼吸道灌洗液中检测到了针对支气管败血波氏杆菌的抗体反应（IgM、IgA和IgG），但未检测到PMTCE特异性抗体。虽然有必要进一步优化支气管败血波氏杆菌中PMT的表达，但本研究为开发针对萎缩性鼻炎的单组分减毒活疫苗提供了基础。
