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Asian schistosomiasis caused by Schistosoma japonicum is a serious zoonotic disease endemic in China, the Philippines and parts of Indonesia. Mass drug administration in endemic areas resulted to decline in disease severity and intensity. The low intensity of infection limits the use of current parasitological methods for schistosomiasis diagnosis. Detection of parasite circulating antigens might provide more informative result as it may indicate the true status of infection. In this study, S. japonicum thioredoxin peroxidase-1 (SjTPx-1) a 22 kDa secreted antioxidant enzyme expressed throughout the life stages of the parasite was evaluated for its potential use as a biomarker for schistosomiasis japonica infection. Rabbit polyclonal antibody and mouse monoclonal antibodies (mAbs) were raised against the recombinant SjTPx-1 (rSjTPx-1). The antibodies produced against the recombinant antigen was confirmed to detect the native SjTPx-1 in crude adult worm lysate. Likewise, the specific binding of mAbs to parasite TPx-1 and not to mammalian peroxiredoxin-1 orthologues was also confirmed. The double antibody sandwich ELISA developed in this study was able to detect at least 1 ng/ml of rSjTPx-1. In addition, this method was able to detect the antigen from all serum samples of experimentally infected rabbit and mice. The diagnostic potential of SjTPx-1 in human clinical samples was also evaluated, in which 4 out of 10 stool-confirmed serum samples had detectable levels of the antigen. The results suggest that SjTPx-1 can be a potential biomarker for Asian zoonotic schistosomiasis.

Sj7TR is a 13 kDa repetitive region of a 31 kDa protein in Schistosoma japonicum known as Sjp_0110390 that showed high sensitivity and specificity in antibody detection against schistosomiasis patients. However, the current database for S. japonicum genes characterized it only as an expressed protein. A more thorough understanding of this antigenic protein is therefore necessary to possibly give more information about the nature of this protein and its role in the parasite. In this study, immunolocalization and expression profiling were done for Sjp_0110390 on the different stages of the parasite. Immunofluorescent assay showed that Sjp_0110390 was expressed in the young stages of the parasites including the schistosomula, eggs, aquatic and intra-molluscan stages. This was supported by the reverse-transcriptase PCR which confirmed the stage-specific expression of Sjp_0110390 and Western blot test which detected the protein in the extracted eggs proteins, but not in the adults. Furthermore, it was also highly expressed in infected Oncomelania hupensis nosophora snails suggesting that Sjp_0110390 might have a role in the development of the parasite inside the intermediate host. This result also suggests that Sj7TR might be used not only for human diagnosis but to detect snail infection as well., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Background: The presence of animal reservoirs in Schistosoma japonicum infection has been a major obstacle in the control of schistosomiasis. Previous studies have proven that the inclusion of control measures on animal reservoir hosts for schistosomiasis contributed to the decrease of human cases. Animal surveillance should therefore be included to strengthen and improve the capabilities of current serological tests., Methodology/principal Findings: Thioredoxin peroxidase-1 (SjTPx-1) and four tandem repeat proteins (Sj1TR, Sj2TR, Sj4TR, Sj7TR) were initially evaluated against human sera. The previous test showed high sensitivity and specificity for antibody detection against SjTPx-1 and Sj7TR. In this study, the immunodiagnostic potential of these recombinant proteins was evaluated using enzyme-linked immunoassay on 50 water buffalo serum samples collected in Cagayan, the Philippines as compared with the soluble egg antigen (SEA). For specificity, 3 goat serum samples positive with Fasciola hepatica were used and among the antigens used, only SEA showed cross-reaction. Stool PCR targeting the S. japonicum 82 bp mitochondrial NAD 1 gene was done to confirm the true positives and served as the standard test. Twenty three samples were positive for stool PCR. SjTPx-1 and Sj1TR gave the highest sensitivity among the recombinant proteins tested for water buffalo samples with 82.61% and 78.26% respectively which were higher than that of SEA (69.57%)., Conclusions/significance: These results prove that SjTPx-1 works both for humans and water buffaloes making it a good candidate antigen for zoonotic diagnosis. Sj1TR showed good results for water buffaloes and therefore can also be used as a possible candidate for detecting animal schistosome infection.

Schistosoma mekongi causes granulomatous lesions around eggs deposited in the liver with neutrophil-rich inflammatory reactions in the early stage of the egg laying. To define the aspects of the typical pathogenesis of S. mekongi infection, we determined the difference between soluble egg antigen (SEA) from S. mekongi and S. japonicum with a focus on chemotactic factors for neutrophils or eosinophils. Mean volume and protein amount of S. mekongi eggs was 71 and 58% of those of Schistosoma japonicum eggs, respectively. Neutrophil chemotactic activity of S. mekongi SEA was about two times higher than that of S. japonicum. In contrast, eosinophil chemotactic activity of S. mekongi SEA was about half of that of S. japonicum SEA. Molecular analysis revealed that S. mekongi SEA contains higher molecular-weight components with a lower level of glycosylation, and this is likely to be related to the intense neutrophil chemotactic activity in comparison with S. japonicum SEA. The prominent chemotactic reactivity for neutrophils is likely to be involved in the typical pathogenesis of mekongi schistosomiasis.

A rapid and simplified ELISA using whole blood samples of Schistosoma japonicum-infected rabbits was compared with a conventional ELISA. This whole-blood ELISA has advantages. The volume of crude egg antigens, whole blood samples, and conjugates was only 0.05 ml. The incubation time was shortened to 5 minutes. Wells were washed three to five times with PBS-Tween after each procedure. Optical density values were measured in 10 minutes after transfer of 0.1 ml of substrate. Constant temperature was not necessary. The entire procedure took only 20-30 minutes.

An ELISA technique was developed using samples of Schistosoma japonicum-infected human whole blood based on the conventional ELISA. In this study, the following were demonstrated. 1) Whole blood samples could be used. 2) The volume of whole blood and conjugate could be reduced to 0.05 ml. 3) The incubation time was shortened to 5 minutes. 4) The optical density could be measured at 10 minutes after transferring the substrate and the volume was reduced to 0.1 ml. 5) It did not require a fixed temperature setting. 6) The operation time was as short as 20 to 30 minutes. 7) The optical density values were almost the same as the conventional ELISA and were not influenced by other common intestinal helminthic infections. 8) The observed variations from day to day including effects of sampling in stool examination were negated by the results of this ELISA technique. 9) Based on correlation with stool examination results, criteria can be formulated in which optical density values of 0.3 and above as positive, 0.1 to less than 0.3 as doubtful, and less than 0.1 as negative. Whenever an immunological field survey is necessary, before and after a selective or a mass treatment control program, this WHOLE BLOOD-ELISA, which was shown to be rapid and simple, is recommended.

Humoral immune responses of IgG, IgM, IgA, IgE and IgG subclass antibodies to Schistosoma japonicum egg antigens were determined by immunoblotting with serum samples from individuals in China with acute (n=24) or chronic (n=35) schistosomiasis. In general, IgM, IgA, and IgE in sera from acute patients exhibited strong binding to antigens but binding was much weaker in chronic cases. Reaction of IgG4 of chronic cases was stronger than that of IgG4 of acute cases. The recognition profile of each antibody isotype in sera was analyzed for 11 major antigen molecules (antigens with apparent molecular weights of 82, 76, 61, 57, 53, 46, 40, 32, 27, 10 and less than 6.5 kDa). Except for the 10 kDa molecule, they were well-recognized by IgA and IgE in sera of acute cases. In other combinations of antibody class and clinical phase, recognition patterns against these molecules differed among individuals. Notably, the 10 kDa molecule was specifically recognized by total IgG and IgG4 in sera from most of the chronic patients, but in sera from only one acute case. This result suggests that the 10 kDa molecule is one of the major target antigens of IgG4 and may be useful as a marker antigen to characterize the clinical phases of S. japonicum infection.