Middle East Respiratory Syndrome Coronavirus (MERS-CoV) spike protein as a DNA candidate vaccine

A. Ghamry, E. Abushady, M. Shehata, Mahmoud Shahata, M. Ali
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Abstract

Background Middle East respiratory syndrome coronavirus (MERS-CoV) has become a global public health threat as it is capable of causing life-threatening disease with lower respiratory tract involvement, with a case fatality rate of ∼37.5%. So, ongoing efforts to develop MERS-CoV vaccines are mandatory, and their immunity profiles against different antigens and correlation with protection should be assessed. Aim The present study aimed to assess the neutralizing capacity of MERS-CoV spike (S) structural protein as a DNA-based candidate vaccine in mice models. Materials and methods The spike structural protein gene of MERS-CoV was amplified and cloned using pcDNA3.1 (negative) mammalian expression vector and competent Escherichia coli, for immunization of BALB/c mice as a DNA candidate vaccine, followed by a booster dose after 2 weeks. Sera of mice were collected within 8 weeks after prime vaccination for evaluation of the neutralizing capacity of DNA vaccine using plaque reduction neutralization test (PRNT) assay compared with the neutralizing capacity of inactivated whole virus vaccine, and also a group of mice was injected with empty vector in phosphate-buffered saline (PBS); PBS-pcDNA3.1 (negative) was used as a negative control. Results PRNT50 showed complete neutralization in mice vaccinated with inactivated MERS-CoV vaccine (PRNT50 titer, ∼1 : 160) 6 and 8 weeks of first immunization (P<0.01). The negative control group of mice injected with PBS-pcDNA3.1 (negative) did not show any neutralizing antibodies against MERS-CoV at 2, 4, 6, and 8 weeks after prime vaccination. The mice vaccinated with S gene-based DNA vaccine (pcDNA3.1-S) showed a significant increase of neutralizing antibodies against MERS-CoV strain NC163/2014 at week 8 after prime vaccination with PRNT50 titer, ∼1 : 80. Conclusion These results reported that the spike gene-expressed protein is a major immunogenic protein in MERS-CoV, so it would be recommended in future vaccine development.
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中东呼吸综合征冠状病毒(MERS-CoV)刺突蛋白作为DNA候选疫苗
中东呼吸综合征冠状病毒(MERS-CoV)已成为全球公共卫生威胁,因为它能够引起危及生命的疾病,并累及下呼吸道,病死率约为37.5%。因此,开发MERS-CoV疫苗的持续努力是强制性的,并应评估其对不同抗原的免疫特性及其与保护的相关性。目的研究MERS-CoV刺突(S)结构蛋白作为一种基于dna的候选疫苗在小鼠模型中的中和能力。材料与方法利用pcDNA3.1(阴性)哺乳动物表达载体和大肠杆菌扩增和克隆MERS-CoV刺突结构蛋白基因,作为DNA候选疫苗免疫BALB/c小鼠,2周后接种加强剂。在预接种后8周内收集小鼠血清,采用斑块减少中和试验(PRNT)法比较DNA疫苗与灭活全病毒疫苗的中和能力,并将空载体注射于磷酸盐缓冲盐水(PBS)中;PBS-pcDNA3.1(阴性)作为阴性对照。结果接种MERS-CoV灭活疫苗(PRNT50滴度,~ 1:16 0)后第6周和第8周小鼠PRNT50完全中和(P<0.01)。注射PBS-pcDNA3.1(阴性)的阴性对照组小鼠在预接种后2、4、6和8周均未显示出任何针对MERS-CoV的中和抗体。接种S基因DNA疫苗(pcDNA3.1-S)的小鼠在PRNT50滴度为1:80的预接种后第8周,对MERS-CoV株NC163/2014的中和抗体显著增加。结论刺突基因表达蛋白是MERS-CoV的主要免疫原性蛋白,可作为今后疫苗开发的参考蛋白。
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