Immunoepidemiological studies from endemic areas have revealed age-dependent resistance correlation with increased level of IgE and decreased level of IgG4 antibodies in responses to schistosomes’ soluble worm antigen. However, there have been limited studies on analyses of major antigens that provoke IgE and IgG4 immune response during chronic stage of schistosomiasis. In this study, for the first time, immunoproteomics approach has been applied to identify S. japonicum worm antigens in liquid fractions that are recognized by IgE and IgG4 antibody using plasma from chronically infected population. ProteomeLabPF 2D fractionated 1-D and 2-D fractions of SWA antigens were screened using pooled high IgE/IgG4 reactive plasma samples by dot-blot technique. In 1-D fractions, IgE isotype was detected by fewer antigenic fractions (43.2%). The most recognized isotype was IgG3 (79.5%) followed by IgG1 (75.0%) and IgG4 (61.4%). Liquid chromatography MS/MS protein sequencing of reactive 2-D fractions revealed 18 proteins that were identified, characterized and gene ontology categories determined. 2-D fractions containing proteins such as zinc finger, RanBP2-type, domain-containing protein were strongly recognized by IgE and moderately by IgG4 whereas fractions containing proteins such as ubiquitin-conjugating enzyme and cytosolic II 5'-nucleotidase strongly recognizing by IgG subclasses (IgG1, IgG3 and IgG4) but not IgE. By this study, a simple and reproducible proteomic method has been established to identify major immunoreactive S. japonicum antigens. It is anticipated that this will stimulate further research on the immunogenicity and protective potential of proteins identified as well as discovery of novel compounds that have therapeutic importance.

Mosquito eggs laid within two hours are necessary for transgenic (injection) studies, because mosquito eggs become hard after that period. Thus, in order to have eggs available within this two-hour window, it is important to understand the ovipositional behavior of Anopheles gambiae s.s.. In the present study, the ovipositional behavior of An. gambiae s.s. (Kisumu) was investigated in several different conditions: age of mosquitoes, time post blood meal to access oviposition substrate, and light conditions. Two groups of mosquitoes, 3–5 days old and 9–11 days old were blood-fed. For those mosquito groups, an oviposition dish was set either at 48 hours or 72 hours after the blood meal either in a light condition or in an artificial dark condition. The number of laid eggs was compared among the different conditions. The 3–5 day-old mosquitoes apparently produced a higher number of eggs than 9–11 day-old mosquitoes, while there was no significant difference between the two groups. The number of laid eggs per one surviving blood-fed mosquito in the dark condition was significantly higher than that in the light condition (p = 0.03). Providing an oviposition dish at 72 hours after blood meal resulted in a significantly higher number of laid eggs per one surviving blood-fed mosquito than at 48 hours after blood meal (p = 0.03). In conclusion, the optimal condition to have readily available egg supply for transgenic analysis was as follows: 3–5 day-old mosquitoes with an oviposition dish placed at 72 hours after the blood meal in a dark environment.

Mosquito eggs laid within two hours are necessary for transgenic (injection) studies, because mosquito eggs become hard after that period. Thus, in order to have eggs available within this two-hour window, it is important for transgenic studies to understand the ovipositional behavior of Anopheles gambiae s.s.. In the present study, the ovipositional behavior of An. gambiae s.s. (Kisumu) was investigated in several different conditions: age of mosquitoes, time post blood meal to access oviposition substrate, and light conditions. Two groups of mosquitoes, 3–5 day old and 9–11 day old mosquitoes were blood-fed. For those mosquito groups, an oviposition dish was set either at 48 hours or 72 hours after the blood meal either in a light condition or in an artificial dark condition. The number of laid eggs was compared among different conditions. The 3–5 day old mosquitoes apparently produced a higher number of eggs than 9–11 day old ones, while there was no significant difference between the 2 groups. The number of laid eggs per one surviving blood-fed mosquito in the dark condition was significantly higher than that in the light condition (p = 0.03). Providing an oviposition dish at 72 hours after blood meal resulted in a significantly higher number of laid eggs per one surviving blood-fed mosquito compared to providing an oviposition dish at 48 hours after blood meal (p = 0.03). In conclusion, the optimal condition to have readily available egg supply in the present study for transgenic analysis was as follows: 3–5 days old mosquitoes with an oviposition dish placed at 72 hours after the blood meal in a dark environment.

Immunobiological roles of schistosome eggs during murine experimental infection were investigated with special reference to the induction of basophilic leukocytes. After single- or bisexual infection with Schistosoma mansoni in BALB⁄c mice, splenomegaly and liver granulomas were observed only in bisexual infection in parallel with deposition of mature parasite eggs. Comparison of the kinetics of basophil response revealed a marked increase in number in the bone marrow of mice with bisexual infection at the 7^th week post infection as opposed to a marginal increase in single- sex infections. In the spleen, bimodal response was observed in the basophil responses; a small but repeatable peak at the 4^th week after infection, increasing again at the 8^th week, which corresponded to the initiation and maturation of parasite eggs in the affected organs of infected mice. The same time course was observed for IL-4 production by the splenocytes from mice of bisexual infection. To obtain more concrete evidence of the role of eggs in the induction of basophils, we tested using the intravenous egg injection model. Injection of eggs induced basophilia, and it was accompanied by the up-regulation of IL-4 production in splenocytes from the 8^th day. Basophils induced in this model showed a high level of IL-4 production confirmed by flow cytometry, while faint levels of IL-4 production were observed for CD4⁺ T cells at this time point. In addition, we demonstrate that egg deposition is the trigger of basophil induction and activation in the murine experimental model of S. mansoni infection, which might play an essential role in the initiation of Th1⁄2 conversion during the course of S. mansoni infection in vivo.