Yuanxi Shen1, Rongyi Ji1, Rui Chai1, Nana Yuan1, Jiyue Zhang1, Yi Jing1, Man Yang1, Lanqi Zhang1,2, Yang Hong1, Jiaojiao Lin1,3, Chuangang Zhu4. 1. Key Laboratory of Animal Parasitology, Ministry of Agriculture of China, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai, China. 2. University of Reading, Whiteknights, Reading, Berkshire, RG26UA, England. 3. Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, China. 4. Key Laboratory of Animal Parasitology, Ministry of Agriculture of China, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai, China. zcg@shvri.ac.cn.
Abstract
BACKGROUND: Schistosomiasis japonica is a severe zoonosis. Domestic animals are the primary source of infection and play an important role in disease transmission. Surveillance and diagnosis play key roles in schistosomiasis control; however, current techniques for the surveillance and diagnosis of the disease have limitations. In this study, we developed a novel fluorescence immunochromatographic assay (FICA) strip to detect anti-Schistosoma japonicum antibodies in host serum. METHODS: A FICA strip was developed for the diagnosis of Schistosoma japonicum in domestic animals. Streptococcus protein G (SPG) and soluble egg antigen (SEA) were transferred onto a nitrocellulose (NC) membrane to form the control line (C) and the test line (T), respectively. With fluorescence activity as well as binding activity to multispecies IgG, the recombinant protein rSPG-RFP was expressed and employed as an antibody indicator in the FICA strips. RESULTS: The dual gene fusion plasmid was verified by PCR and restriction enzyme digestion. The expressed recombinant protein was 39.72 kDa in size, which was consistent with the predicted molecular weight. The western blot results showed binding activity between rSPG-RFP and IgGs from different hosts. Fluorescence microscopy also showed the fluorescence activity of the protein present. The affinity constant (Ka) values of rSPG-RFP with rabbit, donkey, mouse and goat IgG were 1.9 × 105, 4.1 × 105, 1.7 × 105 and 5.4 × 105, respectively. Moreover, based on the recombinant protein, the test strip for detecting S. japonicum in buffaloes could distinguish positive from negative serum. The lower limit of detection of the FICA strip was 1:10,000. Compared with ELISA, the FICA strips exhibited similar results in the diagnosis of infection in clinical bovine serum samples, with a kappa value of 0.9660 and P < 0.01. The cross-reactivities of the FICA strips with Haemonchus contortus and Schistosoma turkestanicum (30.15% and 91.66%, respectively) were higher than those of ELISA (26.98% and 87.5%, respectively). CONCLUSIONS: Based on the rSPG-RFP protein that we developed, strip detection can be completed within 15 min. Heightened sensitivity allows the strip to accurately identify schistosome antibodies in serum. In conclusion, this method is convenient, feasible, rapid and effective for detecting S. japonicum.
BACKGROUND:Schistosomiasis japonica is a severe zoonosis. Domestic animals are the primary source of infection and play an important role in disease transmission. Surveillance and diagnosis play key roles in schistosomiasis control; however, current techniques for the surveillance and diagnosis of the disease have limitations. In this study, we developed a novel fluorescence immunochromatographic assay (FICA) strip to detect anti-Schistosoma japonicum antibodies in host serum. METHODS: A FICA strip was developed for the diagnosis of Schistosoma japonicum in domestic animals. Streptococcus protein G (SPG) and soluble egg antigen (SEA) were transferred onto a nitrocellulose (NC) membrane to form the control line (C) and the test line (T), respectively. With fluorescence activity as well as binding activity to multispecies IgG, the recombinant protein rSPG-RFP was expressed and employed as an antibody indicator in the FICA strips. RESULTS: The dual gene fusion plasmid was verified by PCR and restriction enzyme digestion. The expressed recombinant protein was 39.72 kDa in size, which was consistent with the predicted molecular weight. The western blot results showed binding activity between rSPG-RFP and IgGs from different hosts. Fluorescence microscopy also showed the fluorescence activity of the protein present. The affinity constant (Ka) values of rSPG-RFP with rabbit, donkey, mouse and goat IgG were 1.9 × 105, 4.1 × 105, 1.7 × 105 and 5.4 × 105, respectively. Moreover, based on the recombinant protein, the test strip for detecting S. japonicum in buffaloes could distinguish positive from negative serum. The lower limit of detection of the FICA strip was 1:10,000. Compared with ELISA, the FICA strips exhibited similar results in the diagnosis of infection in clinical bovine serum samples, with a kappa value of 0.9660 and P < 0.01. The cross-reactivities of the FICA strips with Haemonchus contortus and Schistosoma turkestanicum (30.15% and 91.66%, respectively) were higher than those of ELISA (26.98% and 87.5%, respectively). CONCLUSIONS: Based on the rSPG-RFP protein that we developed, strip detection can be completed within 15 min. Heightened sensitivity allows the strip to accurately identify schistosome antibodies in serum. In conclusion, this method is convenient, feasible, rapid and effective for detecting S. japonicum.