Huang Bin, Kwang Jimmy, Loh Hilda, Frey Joachim, Tan Hai-Meng, Chua Kim-Lee
Department of Biochemistry, Faculty of Medicine, Institute of Molecular Agrobiology, National University of Singapore, 117604, Singapore, Singapore.
Vet Microbiol. 2002 Sep 24;88(4):339-49. doi: 10.1016/s0378-1135(02)00123-2.
Riemerella anatipestifer, a gram-negative bacillus, is the causative agent of duck septicemia, a disease which could incur much economic loss in the duck industry. An indirect enzyme-linked immunosorbent assay (ELISA) has been developed to facilitate early detection of R. anatipestifer infection in ducks. The antigen used was a recombinant 41 kDa N-terminal fragment (rP45N') of a newly characterized R. anatipestifer potential surface protein, P45, which was expressed in Escherichia coli as an N-terminal GST fusion protein. The rP45N'-based ELISA successfully detected P45 antibodies in the sera of 20 ducks immunized with bacterin preparations of R. anatipestifer serotypes 1, 10 15, 19 and the ATCC11845 strain. Antibodies to P45 were also detected in the sera of 25% (75/296) of White Pekin ducks which were imported into Singapore from three different farms. Successful discrimination was obtained between sera from infected ducks and that of specific-pathogen free ducks (p<0.01). The rP45N'-GST antigen did not cross-react with antibodies in sera from guinea pigs which were infected with other gram-negative and gram-positive bacterial pathogens, including Aeromonas hydrophila, Citrobacter freundii, E. coli, Klebsiella pneumoniae, Pastuerella multocida, Proteus mirabilis, Salmonella spp., Serratia maccescens, Shigella sonnei and Yersinia enterocolitica. In addition, the DNA sequence encoding P45 was detected in R. anatipestifer serotypes 1, 2, 3, 4, 5, 6, 7, 8, 9, 11, 14, 15, 16, 17, 18, 19 and the ATCC11845 strain, suggesting that P45 is probably also universally expressed in these R. anatipestifer serotypes. Thus, the ELISA described is applicable to the detection of R. anatipestifer infection in ducks.
鸭疫里默氏杆菌是一种革兰氏阴性杆菌，是鸭败血症的病原体，这种疾病会给养鸭业造成巨大经济损失。已开发出一种间接酶联免疫吸附测定（ELISA）方法，以促进鸭疫里默氏杆菌感染的早期检测。所用抗原是一种新鉴定的鸭疫里默氏杆菌潜在表面蛋白P45的重组41 kDa N端片段（rP45N'），它在大肠杆菌中作为N端GST融合蛋白表达。基于rP45N'的ELISA成功检测到用鸭疫里默氏杆菌血清型1、10、15、19及ATCC11845菌株的菌体制剂免疫的20只鸭血清中的P45抗体。在从三个不同农场进口到新加坡的25%（75/296）的北京鸭血清中也检测到了P45抗体。感染鸭血清与无特定病原体鸭血清之间成功实现了区分（p<0.01）。rP45N'-GST抗原与感染其他革兰氏阴性和革兰氏阳性细菌病原体（包括嗜水气单胞菌、弗氏柠檬酸杆菌、大肠杆菌、肺炎克雷伯菌、多杀巴斯德菌、奇异变形杆菌、沙门氏菌属、粘质沙雷氏菌、宋内志贺氏菌和小肠结肠炎耶尔森氏菌）的豚鼠血清中的抗体无交叉反应。此外，在鸭疫里默氏杆菌血清型1、2、3、4、5、6、7、8、9、11、14、15、16、17、18、19及ATCC11845菌株中检测到了编码P45的DNA序列，这表明P45可能在这些鸭疫里默氏杆菌血清型中也普遍表达。因此，所述ELISA适用于鸭疫里默氏杆菌感染的检测。
