Sączyńska Violetta, Romanik-Chruścielewska Agnieszka, Florys-Jankowska Katarzyna, Cecuda-Adamczewska Violetta, Kęsik-Brodacka Małgorzata
ŁUKASIEWICZ Research Network - Industrial Chemistry Institute, Rydygiera 8 Street, 01-793 Warsaw, Poland.
ŁUKASIEWICZ Research Network - Industrial Chemistry Institute, Rydygiera 8 Street, 01-793 Warsaw, Poland.
Vet Immunol Immunopathol. 2022 May;247:110406. doi: 10.1016/j.vetimm.2022.110406. Epub 2022 Mar 8.
Avian influenza viruses (AIVs) and especially highly pathogenic (HP) AIVs of the H5 and H7 subtypes are of both veterinary and public health concern worldwide. In response to the demand for effective vaccines against H5N1 HPAIVs, we produced recombinant protein based on hemagglutinin (HA), a protective viral antigen. A fragment of the HA ectodomain, with a multibasic cleavage site deletion, was expressed in Escherichia coli, refolded, and chromatographically purified from inclusion bodies. Finally, the protein was formulated in Tris-HCl buffer of pH 8.0 or PBS of pH 7.4 to obtain antigens denoted rH5-1 and rH5-2, respectively. The systemic prime and boost immunizations proved that rH5-1 adsorbed to aluminum hydroxide induces anti-H5 HA neutralizing antibodies and protective immune responses against H5N1 HPAIVs in chickens. The present studies were aimed at stimulating immune responses via the mucosal routes using the systemic prime-mucosal boost strategy. Efficacy trials were performed in commercial layer chickens. For systemic and mucosal immunizations, H5 HA antigens were adjuvanted with aluminum hydroxide and chitosan glutamate, respectively. The first dose of rH5-2 was administered subcutaneously, while its second dose was administered subcutaneously, intraocularly, oculo-nasally, or intranasally. rH5-1 was delivered to the subcutaneously primed chickens by the intranasal route. Post-vaccination sera were analyzed for anti-H5 HA antibodies, using homologous ELISA and heterologous FluAC H5 and hemagglutination inhibition tests. Intraocularly and oculo-nasally delivered rH5-2 mixed with chitosan glutamate was capable of stimulating anti-H5 HA IgY antibody responses in the subcutaneously primed chickens; however, it was ineffective when administered by the intranasal route. Efficient intranasal boosting was achieved using rH5-1. The enhanced production of antigen-specific antibodies was reflected in the development of H5-subtype specific and hemagglutination inhibiting antibodies. Conclusively, the subcutaneous prime and oculo-nasal boost vaccination is proposed as the target strategy for future optimization.
禽流感病毒（AIVs），尤其是H5和H7亚型的高致病性（HP）AIVs，在全球范围内都受到兽医和公共卫生领域的关注。为了满足对抗H5N1高致病性禽流感病毒有效疫苗的需求，我们基于血凝素（HA）这种保护性病毒抗原生产了重组蛋白。HA胞外域的一个片段，带有多碱性切割位点缺失，在大肠杆菌中表达，复性，并从包涵体中通过色谱法纯化。最后，将该蛋白分别用pH 8.0的Tris-HCl缓冲液或pH 7.4的磷酸盐缓冲盐水（PBS）配制，以获得分别称为rH5-1和rH5-2的抗原。全身初次免疫和加强免疫证明，吸附到氢氧化铝上的rH5-1能诱导鸡产生抗H5 HA中和抗体以及针对H5N1高致病性禽流感病毒的保护性免疫反应。本研究旨在通过全身初次免疫-黏膜加强策略，经由黏膜途径刺激免疫反应。在商品蛋鸡中进行了效力试验。对于全身免疫和黏膜免疫，H5 HA抗原分别用氢氧化铝和谷氨酸壳聚糖佐剂。rH5-2的第一剂通过皮下注射给药，而其第二剂通过皮下注射给药、眼内给药、眼鼻给药或鼻内给药。rH5-1通过鼻内途径递送至皮下初次免疫的鸡。使用同源酶联免疫吸附测定（ELISA）以及异源流感病毒血凝抑制试验（FluAC H5），对疫苗接种后的血清进行抗H5 HA抗体分析。与谷氨酸壳聚糖混合的眼内和眼鼻给药的rH5-2能够刺激皮下初次免疫的鸡产生抗H5 HA IgY抗体反应；然而，通过鼻内途径给药时无效。使用rH5-1实现了有效的鼻内加强免疫。抗原特异性抗体产量的增加反映在H5亚型特异性抗体和血凝抑制抗体的产生上。总之，皮下初次免疫和眼鼻加强免疫接种被提议作为未来优化的目标策略。
