Haghighat Setareh, Siadat Seyed Davar, Akhavan Sepahi Abbas, Mahdavi Mehdi
Department of Microbiology, Faculty of Advanced Science and Technology, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran.
Department of Mycobacteriology & Pulmonary Research, Microbiology Research Center, Pasteur Institute of Iran, Tehran, Iran.
Iran J Basic Med Sci. 2022 Apr;25(4):442-450. doi: 10.22038/IJBMS.2022.59992.13303.
Methicillin-resistant (MRSA) reasons extreme infections, can resist various conventional antimicrobial agents, and cause morbidity and mortality worldwide. Vaccination seems to help modulate MRSA infections. Nanovaccine is considered a novel strategy in vaccine technology. The primary purpose of the present study was to develop a conjugate vaccine based on recombinant PBP2a and MRSA autolysin formulated in PLGA as a nanoparticle capable of enhancing protective responses against MRSA in the murine model.
Recombinant PBP2a and autolysin have been expressed and purified by nickel-nitrilotriacetic acid (Ni-NTA) affinity column and characterized by SDS-PAGE and western blot. PLGA was bound to recombinant proteins by using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDAC) and adipic acid dihydrazide (ADH) as a linker and spacer, respectively. Conjugation of recombinant proteins to PLGA was confirmed by the AFM assay, zeta potential, and size distribution, and its efficacy was evaluated in mice. Total IgG, IgG1, IgG2a, IgG2b, and IgM titers were analyzed to assess immune responses. Lastly, the bioactivity of antibodies was tested by using the opsonophagocytosis assay.
Mice immunized with the r-PBP2a-r-autolysin-PLGA nanovaccine led to increased levels of opsonic antibodies and IgG1, IgG2a, IgG2b, and IgM when compared with other experimental groups. Our results confirmed that vaccination with nanovaccine could reduce the mortality rate against the sub-lethal dose of MRSA challenge. Furthermore, the nanovaccine could eliminate MRSA from the kidney of infected mice.
This study may provide valuable insights into the protective power of the r-PBP2a-r-autolysin-PLGA conjugate vaccine against MRSA infection.
耐甲氧西林金黄色葡萄球菌（MRSA）引发严重感染，能抵抗多种传统抗菌药物，在全球范围内导致发病和死亡。疫苗接种似乎有助于调节MRSA感染。纳米疫苗被认为是疫苗技术中的一种新策略。本研究的主要目的是开发一种基于重组PBP2a和MRSA自溶素的结合疫苗，该疫苗以聚乳酸-羟基乙酸共聚物（PLGA）为纳米颗粒制剂，能够增强小鼠模型中针对MRSA的保护性反应。
重组PBP2a和自溶素已通过镍-亚氨基三乙酸（Ni-NTA）亲和柱进行表达和纯化，并通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）和蛋白质免疫印迹法进行表征。分别使用1-乙基-3-（3-二甲基氨基丙基）碳二亚胺盐酸盐（EDAC）和己二酸二酰肼（ADH）作为连接剂和间隔物，将PLGA与重组蛋白结合。通过原子力显微镜（AFM）检测、zeta电位和尺寸分布确认重组蛋白与PLGA的结合，并在小鼠中评估其效果。分析总免疫球蛋白G（IgG）、免疫球蛋白G1（IgG1）、免疫球蛋白G2a（IgG2a）、免疫球蛋白G2b（IgG2b）和免疫球蛋白M（IgM）滴度以评估免疫反应。最后，通过调理吞噬作用试验检测抗体的生物活性。
与其他实验组相比，用重组PBP2a-重组自溶素-PLGA纳米疫苗免疫的小鼠导致调理抗体以及IgG1、IgG2a、IgG2b和IgM水平升高。我们的结果证实，用纳米疫苗接种可降低亚致死剂量MRSA攻击后的死亡率。此外，纳米疫苗可从感染小鼠的肾脏中清除MRSA。
本研究可能为重组PBP2a-重组自溶素-PLGA结合疫苗对MRSA感染的保护作用提供有价值的见解。
