Pushpakumara Pradeep D, Jeewandara Chandima, Bary Farha, Madushanka Deshan, Perera Lahiru, Aberathna Inoka Sepali, Nimasha Thashmi, Jayamali Jeewantha, Ranasinghe Thushali, Kuruppu Heshan, Danasekara Saubhagya, Wijewickrama Ananda, Ogg Graham S, Malavige Gathsaurie Neelika
Allergy Immunology and Cell Biology Unit, Department of Immunology and Molecular Medicine, University of Sri Jayewardenepura, Nugegoda, Sri Lanka.
National Institute of Infectious Diseases, Angoda, Sri Lanka.
medRxiv. 2023 Jan 7:2023.01.05.23284247. doi: 10.1101/2023.01.05.23284247.
As there are limited data on B cell epitopes for the nucleocapsid protein in SARS-CoV-2, we sought to identify the immunodominant regions within the N protein, recognized by patients with varying severity of natural infection with the Wuhan strain (WT), delta, omicron and in those who received the Sinopharm vaccines, which is an inactivated, whole virus vaccine.
Using overlapping peptides representing the N protein, with an in-house ELISA, we mapped the immunodominant regions within the N protein, in seronegative (n=30), WT infected (n=30), delta infected (n=30), omicron infected+vaccinated (n=20) and Sinopharm (BBIBP-CorV) vaccinees (n=30). We then investigated the sensitivity and specificity of these immunodominant regions and analysed their conservation with other SARS-CoV-2 variants of concern, seasonal human coronaviruses and bat Sarbecoviruses. We then investigated the kinetics of responses to these regions in those with varying severity of acute COVID-19.
We identified four immunodominant regions aa 29-52, aa 155-178, aa 274 to 297 and aa 365 to 388, were highly conserved within SARS-CoV-2 and the bat coronaviruses. The magnitude of responses to these regions varied based on the infecting SARS-CoV-2 variants, with WT infected individuals predominantly recognizing aa155 to 178 regions, delta infected individuals and vaccinated+omicron infected individuals predominantly recognizing regions aa 29 to 52 and aa 274 to 294 regions. Sinopharm vaccinees recognized all four regions, with the magnitude of responses significantly lower than other groups. >80% of individuals gave responses above the positive cut-off threshold to many of the four regions, with some differences with individuals who were infected with different VoCs. These regions were found to be 100% specific, as none of the seronegative individuals gave any responses.
N-protein specific responses appear to be detectable in over 90% of those who were naturally infected or vaccinated with a whole virus inactivated vaccine, with responses mainly directed against four regions of the protein, which were highly conserved. As these regions were highly specific with high sensitivity, they have a potential to be used to develop diagnostic assays and to be used in development of vaccines.
由于关于严重急性呼吸综合征冠状病毒2（SARS-CoV-2）核衣壳蛋白B细胞表位的数据有限，我们试图确定N蛋白内的免疫显性区域，这些区域在感染武汉毒株（WT）、德尔塔、奥密克戎的不同严重程度的自然感染患者以及接种国药疫苗（一种灭活全病毒疫苗）的患者中被识别。
使用代表N蛋白的重叠肽，通过内部酶联免疫吸附测定（ELISA），我们绘制了N蛋白内的免疫显性区域，这些区域来自血清阴性者（n = 30）、感染WT者（n = 30）、感染德尔塔者（n = 30）、感染奥密克戎并接种疫苗者（n = 20）以及接种国药（BBIBP-CorV）疫苗者（n = 30）。然后，我们研究了这些免疫显性区域的敏感性和特异性，并分析了它们与其他值得关注的SARS-CoV-2变异株、季节性人类冠状病毒和蝙蝠Sarbecoviruses的保守性。接着，我们研究了急性新冠肺炎严重程度不同的患者对这些区域的反应动力学。
我们确定了四个免疫显性区域，即氨基酸29 - 52、氨基酸155 - 178、氨基酸274至297以及氨基酸365至388，它们在SARS-CoV-2和蝙蝠冠状病毒中高度保守。对这些区域的反应强度因感染的SARS-CoV-2变异株而异，感染WT的个体主要识别氨基酸155至178区域，感染德尔塔的个体以及接种疫苗并感染奥密克戎的个体主要识别氨基酸29至52区域和氨基酸274至294区域。接种国药疫苗者识别所有四个区域，但其反应强度明显低于其他组。超过80%的个体对这四个区域中的许多区域的反应高于阳性临界阈值，不同变异株感染个体之间存在一些差异。这些区域被发现具有100%的特异性，因为血清阴性个体均无任何反应。
在超过90%的自然感染或接种全病毒灭活疫苗的个体中似乎可检测到N蛋白特异性反应，反应主要针对该蛋白的四个高度保守区域。由于这些区域具有高特异性和高敏感性，它们有潜力用于开发诊断检测方法和疫苗。
