Study On Diagnosis Of Chronic Schistosomiasis Japonicum With The Recombinant SJ26GST-SJ32 Fusion Protein

ObjectiveTo study the value of rSj26GST-Sj32 fusion protein for diagnosis of chronic schistosomiasis japonicum, and to investigate amplification of Sj26GST, Sj32 and Sj14-3-3 coding gene by PCR or RT-PCR for gene diagnosis of the patients with chronic schistosomiasis japonicum.MethodsThe recombinant plasmid pET32α-Sj26GST-Sj32 was transformed into E.coli BL21(DE3) and was induced with IPTG, then the expression product were purified by Ni-NTA kits and were identified by SDS-PAGE. To detect the IgG and IgG4 in the sera of patients with chronic schistosomiasis japonicum with rSj26GST-Sj32 fusion protein and SjAWA by ELISA, Dot-ELISA and Dipstick. The sera of patients with clonorchiasis sinensis, paragonimiasis westermani, alveolar echinococcosis, cystic echinococcosis, hepatitis B, pulmonary tuberculosis and health donors were taken as controls.The encoding gene of Sj26GST, Sj32 and Sj14-3-3 were amplified by PCR or RT-PCR and identified by 1.2% agarose gel electrophoresis. The sera of patients with paragonimiasis westermani, clonorchiasis sinensis and the health donors were taken as controls.ResultsThe recombinant Sj26GST-Sj32 fusion protein was successfully obtained.The sensitivity and specificity of rSj26GST-Sj32-IgG-ELISA were 95.00% and 97.67% respectively, 92.50% and 97.67% in SjAWA groups. There were cross reactions with two patients with alveolar echinococcosis by SjAWA, but no cross reaction with rSj26GST-Sj32.The sensitivity and specificity of rSj26GST-Sj32-IgG4-ELISA were 95.00% and 100% respectively, but 97.50% and 93.02% by SjAWA. There were cross reactions with the sera of patients with alveolar echinococcosis, cystic echinococcosis and hepatitis B with SjAWA, but no cross reaction with rSj26GST-Sj32.The sensitivity and specificity of rSj26GST-Sj32-HRP-IgG-Dot-ELISA were 92.50% and 95.35%, they were 95.00% and 93.02% in SjAWA groups. There were cross reactions with the sera of patients with clonorchiasis sinensis, paragonimiasis westermani and alveolar echinococcosis with the two antigens, but no cross reaction with the sera of patients with cystic echinococcosis, hepatitis B and pulmonary tuberculosis. The sensitivity and specificity of rSj26GST-Sj32-Immunogold-IgG-Dot-ELISA were 95.00% and 97.67% respectively, and 97.50% and 95.35% in SjAWA groups. Cross reactions were emerged in clonorchiasis sinensis and paragonimiasis westermani with SjAWA, but not with rSj26GST-Sj32.The sensitivity and specificity of rSj26GST-Sj32-Immunogold-IgG-Dipstick were 92.50% and 97.67% respectively, and 97.50% and 95.35% in SjAWA groups. There were cross reactions with sera of patients with clonorchiasis sinensis, paragonimiasis westermani and alveolar echinococcosis with SjAWA, but there was no cross reaction with sera of patients with clonorchiasis sinensis, paragonimiasis westermani, alveolar echinococcosis, cystic echinococcosis, hepatitis B and pulmonary tuberculosis with rSj26GST-Sj32.A about 400 bp fragment of Sj14-3-3 coding gene was obtained by PCR or RT-PCR, but Sj26GST (676 bp) and Sj32 (1270 bp) coding gene were not obtained. Both control groups were negative.Conclusions1. The rSj26GST-Sj32 fusion protein may be used for the immunodiagnosis of chronic schistosomiasis japonicum.2. Amplification Sj14-3-3 coding gene by PCR or RT-PCR may be used for the gene diagnosis of the chronic schistosomiasis japonicum.