Serological proteomic screening and evaluation of a recombinant egg antigen for the diagnosis of low-intensity infections in endemic area in Brazil Antigen target in schistosomiasis diagnosis

Abstract

52Background: Despite decades of employment of control programs, schistosomiasis 53 remains a global public health problem. To further reduce prevalence and intensity of 54 infection, or to achieve the goal of elimination in low-endemic areas, there need to be 55 better diagnostic tools to detect low intensity infections in low-endemic areas as Brazil. 56The rationale for development of new diagnostic tools is because in low-endemic 57 settings, the standard Kato-Katz diagnostic method loses its sensitivity and misses low 58 intensity infections. To develop new diagnostic tools, we employed a proteomics 59 approach search for biomarkers associated with schistosome-specific immune responses 60 to develop sensitive and specific new methods for immunodiagnosis. 61 62 Methods and findings: Immunoproteomic analysis was performed on an egg extract of 63 Schistosoma mansoni using pooled sera from infected or non-infected individuals from 64 a low-endemic area of Brazil. Cross reactivity with other helminth parasites was 65 determined using pooled sera from individuals infected with different parasitic 66helminths. Using this approach we identified 23 spots recognized by schistosome acute 67 and chronic sera samples. To identify immunoreactive spots that are likely glycan 68 epitopes, we compared immunoreactivity of spots treated by sodium metaperiodate 69 oxidation of egg extract. This treatment yielded 12/23 spots maintaining 70 immunoreactivity, suggesting they are protein epitopes. From these 12 spots, 11 spots 71 cross-reacted with sera from infection with other helminths and 10 spots cross-reacted 72 with the negative control group. Spot number 5 was exclusively immunoreactive with 73 sera from schistosome-infected groups in native and deglycosylated conditions and 74 corresponds to a Major Egg Antigen. We expressed the Major egg antigen as a 75 recombinant protein and showed by western blot a similar recognition pattern of that of 4 76 the native protein. IgG-ELISA gave a sensitivity of 87.10% and specificity of 89.09% 77 represented by area under ROC curve of 0.95. IgG-ELISA performed better than the 78 conventional K-K (2 slides) identifying 56/64 cases harboring 1-10 egg per gram of 79 feces that were undiagnosed by K-K parasitological technique.80 81 Conclusions: The serological proteome approach was able to identify a new diagnostic 82 candidate. The recombinant egg antigen provided good performance in IgG-ELISA to 83 detect individuals with extreme low-intensity infections (1 egg per gram of feces). 84 Therefore, the IgG-ELISA using this newly identified recombinant major egg antigen 85 can be a useful tool to be combined with other techniques in low-endemic areas to 86 determine the true prevalence of schistosome infection that is underestimated by Kato-87 Katz method. Further, to overcome the complexity of ELISA in the field, a second-88 generation of antibody-based rapid diagnostic tests (RDT) can be developed. 89 90 Recombinant Protein, Endemic Areas. 92 93 94 95 96 97 98 99 5 100 Author Summary 101 102 ...