诺如病毒GⅡ.4 IgG抗体的荧光素酶免疫吸附方法的建立及在动物血清中的应用

Detection of GⅡ.4 Norovirus Specific IgG in Animal Serum Using A Newly Developed Luciferase Immunosorbent Assay

诺如病毒(Norovirus,NoV)是引起全球范围内人类急性胃肠炎(Acutegastroenteritis,AGE)的主要病原体,对全球健康和经济造成了重大威胁。已有研究在不同种类的动物粪便中检出人类诺如病毒(Human norovirus,HuNoV)的核酸或在家畜血清中检测出可识别HuNoV病毒样颗粒(Virus like particles,VLP)的抗体,这引起了公众对NoV是否为潜在人兽共患病的关注与讨论。本研究基于HuNoV主要流行株GⅡ.4建立了一种检测GⅡ.4IgG的荧光素酶免疫吸附试验(Luciferase immunosorbent assay,LISA)。LISA法具有较高的灵敏度(94.87%)和特异度(98.18%);最低检测限是ELISA法的2~16倍;GⅡ.4-LISA检测人血清的批间和批内变异系数值均小于10%,重复性良好。GⅡ.4 IgG在狗、猪、蝙蝠、大鼠和鼩鼱等不同的家畜和野生动物中均有检出,阳性率在3.37%~39.53%之间,其中蝙蝠的检出率最高(39.53%),狗次之(9.72%)。综上所述,本研究建立的GⅡ.4-LISA方法适用于人和不同种属动物中GⅡ.4特异性IgG检测。针对HuNoV的IgG抗体广泛分布,提示NoV可能是潜在的人畜共患病。

Norovirus(NoV)is the leading cause of acute gastroenteritis.It poses a major threat to the health and economy of societies worldwide.The nucleic acids of human norovirus(HuNoV)have been detected in the feces of different species of animals.Antibodies to HuNoV virus-like particles have been detected in the serum of domestic animals.These phenomena have attracted public attention and discussion on whether NoV is potential zoonotic pathogen.We developed a new luciferase immunosorbent assay(LISA)to detect GⅡ.4immunoglobulin(Ig)G based on the main epidemic strain of HuNoV GⅡ.4.Our LISA had high sensitivity(94.87%)and specificity(98.18%).The limit of detection was 2–16-times higher than that of an enzyme-linked immunosorbent assay.The inter-and intra-assay coefficients of variation of GⅡ.4-LISA were<10%,indicating good repeatability.GⅡ.4 IgG was detected in different domestic and wild animals(dogs,pigs,bats,rats and shrews)with positive rates ranging from 3.37%to 39.53%.The positive rate for the bat was the highest(39.53%),followed by the dog(9.72%).In conclusion,the GⅡ.4-LISA established in this study was suitable for the detection of GⅡ.4-specific IgG in humans and different species of animals.The wide distribution of IgG antibodies against HuNoV suggests that NoV may be a zoonotic pathogen.