Cercarial Antigen Preparation Research Articles

The Efficiency of Schistosoma mansoni Crude Antigens in Inhibition of Heat Shock Protein, Apoptosis, and Lysosomal Activity: An Immunohistochemical Study.

The adverse impact of schistosomiasis on tissues is considered in generating a schistosomal vaccine. The purpose of this study was to evaluate the effectiveness of Schistosoma mansoni crude antigens as a therapeutic and prophylactic formulation in the inhibition of heat shock protein, apoptosis, and CD3/CD20 expression in a liver and spleen mouse models using the immunohistochemistry method. A total of 65 mice were divided into five groups: (i) infected untreated group (G1), (ii) therapeutic treated group (G2) with egg soluble egg antigen (SEA), and soluble worm antigen preparation (SWAP), (iii) prophylactically treated group (G3) with cercarial antigen preparation (CAP), (iv) combined treated group with three antigens (G4), and (v) control group (G5). The results we obtained showed that CAP, SEA, and SWAP antigens mitigated the deterioration and inflammation induced by infection. Apoptosis and sinusoidal injuries were significantly reduced when treated with CAP antigen before infection. After infection, using SEA and SWAP antigens may help lighten the liver's load. A high degree of activation in T and B cells in the liver and spleen is linked to this. Our findings shed light on the immunological mechanisms that contribute to the recovery from therapy and vaccination against schistosome damage.

Elevation of C-reactive protein, P-selectin and Resistin as potential inflammatory biomarkers of urogenital Schistosomiasis exposure in preschool children

BackgroundSchistosomiasis is known to induce inflammatory immune responses. C-reactive protein (CRP), resistin and P-selectin are serological inflammatory markers that rise during the acute stages of infection. Here, we propose such inflammatory biomarkers have a potential for use in urogenital schistosomiasis diagnostic screening for exposure and infection in preschool-aged children.MethodsAs part of a larger study on urogenital schistosomiasis, 299 preschool children aged 1–5 years were included in this cross-sectional study. Parasitological diagnosis was conducted using urine filtration for Schistosoma haemtobium infection, and Kato Katz for S. mansoni infection. Serum levels of P-selectin, resistin, CRP, and antibodies against S. haematobium cercarial antigen preparation (CAP) and soluble worm antigen preparation (SWAP) were measured by ELISA.ResultsOf the 299 participants, 14% were egg positive for S. haematobium. Serology showed 46 and 9% of the participants to have been exposed to S. haematobium cercarial antigens and adult worm antigens, respectively. Levels of P-selectin were significantly higher in participants infected with S. haematobium (egg-positive) than in uninfected participants (p = 0.001). Levels of P-selectin were also higher in those exposed to cercarial antigen than in unexposed participants (p = 0.019). There was a positive correlation between P-selectin and infection intensity (r = 0.172; p = 0.002), as well as with IgM responses to CAP and SWAP (r = 0.183; p = 0.001); (r = 0.333; p < 0.0001) respectively. CRP significantly correlated with IgM responses to CAP (r = 0.133; p = 0.029) while resistin correlated with IgM responses to CAP and SWAP (r = 0.127; p = 0.016); (r = 0.197; p = 0.0004). CRP levels were higher in those exposed to cercarial and adult worm antigens than unexposed participants (p = 0.035); (p = 0.002) respectively, while resistin was higher in participants exposed to cercarial antigen than unexposed participants (p = 0.024).ConclusionIn this preschool population, P-selectin is significantly associated with urogenital schistosome infection and intensity; hence a potential biomarker for infection diagnosis and disease monitoring. The inflammatory biomarkers (P-selectin, Resistin and CRP) were significantly higher in participants exposed to cercarial antigens than unexposed individuals indicating an underlying inflammatory environment.

Studies on the role of tumor necrosis factor- alpha (TNF-α) in hepatocytes induced apoptosis in vaccinated, Schistosoma mansoni-challenged mice.

Tumour Necrosis Factor-alpha (TNF-α) plays a complex role in pathophysiological changes caused by schistosomiasis in the liver cells as induced apoptosis. So, The highlighted experimentally the role of TNF-α in hepatocytes apoptosis, using that as an assessment of the efficacy of antischistosomal vaccination by mixed crude antigens preparations [Cercarial antigen preparation (CAP) + soluble worm antigen preparation (SWAP) + soluble egg antigen(SEA)] by parasitological, histo-pathological and histochemical studies using Feulgen stain of hepatoytes DNA, a serological study also of serum TNF-α level by ELISA. Fifty two laboratory bred Albino male mice, were used in this study. They were classified into four groups (13 mice in each group), G1: normal control, G2 as infected control while G3 supported by Freund's Adjuvant (F. Adj) then infected and G4 vaccinated with combined antigens (CAP, SWAP and SEA) + F. Adj, then infected. Mice were sacrificed by cervical dislocation 9 weeks post infection, parasitological (Kato-Katz thick smear for egg count), histopathologial {haematoxylin and eosin (H&E) staining of hepatic sections}, histochemical (feulgen staining of hepatocytes DNA) and ELISA to estimate serum TNF-α level were performed. The data showed that vaccination with combined antigens showed protective effect on vaccinated then Schistosoma challenged mice, hepatocytes induced apoptosis was directly proportional with the TNF-α serum level, and the protection degree of potential combined vaccine was inversely proportional with serum TNF-α level and induced apoptosis.

Effect of Ultra Violet Attenuated Cercariae on Antibody Response in Rabbits

Ultraviolet (UV) light is an electromagnetic radiation below the visible wavelengths. Their use in cercariae attenuation needs to be examined. In the present work, Schistosoma mansoni cercariae was exposed to UV radiation. Few studies showed previously an inflicted damage seen on the adult schistosome worms developed from irradiated cercariae. The aim of the study was to find out whether this damage was attributed to direct effect of UV-irradiation on cercariae or due to the host's immunogenicity induced by UV- irradiated cercariae. Single as well as multiple exposures of cercariae to UV Light were examined. They were exposed to UV light, for 1, 2 and 3 hrs and then, were subjected to different treatments: the first one was for the assessment of cercarial viability after one hour of the 3 different treatments. The second one was for rabbit infection and the third one was for cercarial antigen preparation. The cercarial antigens (Ag) were recognized by protective antibodies (IgG1 fractions) which then, were separated and purified from vaccinated rabbit's serum. This cercarial Ag was identified as UVISmC1 gene encoding a protein showing 100% identity at the amino acid level with previously identified S. mansoni clones; theses clones are encoding 51.7 kDa antigens elicited as a result of direct effect of UV radiation on cercariae as well as host's immunogenicity induced by UV irradiated cercariae. This was verified by the recognition of this Ag prepared from E coli clones isolated from cDNA expression library. In conclusion, the data showed a remarkable potency of the UV-radiation-attenuated cercaria in eliciting differential high effectiveness in Ab response under laboratory conditions at 1, 2 and 3 hrs of UV light output at 254 nm. An understanding of the protective immune response elicited by RA cercaria may help in designing a candidate vaccine which is still needed.

In vitroandin vivoeffects of hesperidin treatment on adult worms ofSchistosoma mansoni

Hesperidin has been reported to exert a wide range of pharmacological effects, including antifungal, antiviral, antioxidant, anti-inflammatory and anticarcinogenic activities. Herein, the schistosomicidal activity of this compound was evaluated in vitro and in vivo. Using an in vitro assay, a concentration of 200 μg/ml of hesperidin resulted in the mortality of 100% adult worms of Schistosoma (S.) mansoni within 72 h and a partial tegumental alteration in 10% of worms. However, after 144 h incubation, 50 and 100 μg/ml concentrations showed 0% and 10% mortality in adult worms, respectively, without any changes to the tegument. Sublethal doses did not influence egg output nor the development of eggs deposited by pairs of adult worms. In an in vivo study, mice infected with S. mansoni and treated with 600 mg hesperidin/kg body weight showed a respective reduction of 50, 45.2, 50 and 47.5% of males, females, worm pairs and total worm burden. In addition, a respective reduction, based on the number of eggs/g tissue, of 41.5, 63.7 and 58.6% was observed in the liver, intestine and liver/intestinal tissue combined. Furthermore, S. mansoni-specific IgG level significantly increased with hesperidin treatment, whereas IgA and IgE levels were not significantly changed. IgM levels decreased in response to cercarial antigen preparation but were not altered in response to soluble worm or soluble egg antigen. As in hesperidin-treated mice, praziquantel-treated mice showed a similar pattern of specific antibody response to S. mansoni antigens. The present study represents the first report on the effects of the schistosomicidal activity of hesperidin.

Efficacy of L-carnosine against Schistosoma mansoni antigens in liver, heart, kidney and brain of rabbits

Article history: The present investigation was an attempt to study the enhancememt effect of L-carnosine (beta alanyl-l-histidine) on the influence of vaccination on healthy (non-infected) rabbits treated with Schistosoma mansoni egg antigens, cercariae antigens, and worms antigens as protective agents against infection. This study involved individual injection of three Schistosoma mansoni antigens: soluble egg antigen (SEA), cercarial antigen preparation (CAP) and soluble worm antigen preparation (SWAP), in three rabbit groups, respectively. Three other groups each received the same specific antigen in conjunction with the administration of L-carnosine, biochemical parameters including DNA, RNA, DNA/RNA ratio concentrations in addition ATPase, and acetyl cholinesterase activities were measured in liver, heart, kidney and brain in all groups. Elevation in most parameters was observed in the immunized groups. Carnosine treatment of rabbit groups immunized with SEA, CAP and SWAP in comparison to the non-carnosine-treated immunized groups resulted in amelioration the changes of DNA, RNA, ATPase and acetyl cholinesterase activities in most groups. L-carnosine has a beneficial effect in the amelioration of the changes in biochemical parameters as a result of S. mansoni antigen immunization.

Comparison of Schistosoma mansoni Soluble Cercarial Antigens and Soluble Egg Antigens for Serodiagnosing Schistosome Infections

A Schistosoma mansoni cercarial antigen preparation (cercarial transformation fluid – SmCTF) was evaluated for detection of anti-schistosome antibodies in human sera in 4 collaborating laboratories. The performance of SmCTF was compared with that of S. mansoni egg antigens (SmSEA) in an indirect enzyme-immunoassay (ELISA) antigen assay, the latter being used routinely in 3 of the 4 participating laboratories to diagnose S. mansoni and S. haematobium infections. In the fourth laboratory the performance of SmCTF was compared with that of S. japonicum egg antigens (SjSEA) in ELISA for detection of anti-S. japonicum antibodies. In all 4 laboratories the results given by SmCTF in ELISA were very similar to those given by the antigen preparation routinely used in the respective laboratory to detect anti-schistosome antibodies in human infection sera. In so far as the ELISA results from SmCTF are thus so little different from those given by schistosome egg antigens and also cheaper to produce, the former is a potentially useful new diagnostic aid for schistosomiasis.

A HIGH POTENT ANTIBODY RESPONSE IN RABBITS INFECTED WITH ULTRA VIOLET ATTENUATED CERCARIAE

Ultraviolet (UV) light is an electromagnetic radiation below the visible wavelengths. Their use in cercariae attenuation needs to be examined. In the present work, Schistosoma mansoni cercariae was exposed to UV radiation. Few studies showed previously an inflicted damage seen on the adult schistosome worms developed from irradiated cercariae. The aim of the study was to find out whether this damage was attributed to direct effect of UV-irradiation on cercariae or due to the host's immunogenicity induced by UV- irradiated cercariae. Single as well as multiple exposures of cercariae to UV Light were examined. They were exposed to UV light, for 1, 2 and 3 hrs and then, were subjected to different treatments: the first one was for the assessment of cercarial viability after one hour of the 3 different treatments. The second one was for rabbit infection and the third one was for cercarial antigen preparation. The cercarial antigens (Ag) were recognized by protective antibodies (IgG1 fractions) which then, were separated and purified from vaccinated rabbit's serum. This cercarial Ag was identified as UVISmC1 gene encoding a protein showing 100% identity at the amino acid level with previously identified S. mansoni clones; theses clones are encoding 51.7 kDa antigens elicited as a result of direct effect of UV radiation on cercariae as well as host's immunogenicity induced by UV irradiated cercariae. This was verified by the recognition of this Ag prepared from E coli clones isolated from cDNA expression library. In conclusion, the data showed a remarkable potency of the UV-radiation-attenuated cercaria in eliciting differential high effectiveness in Ab response under laboratory conditions at 1, 2 and 3 hrs of UV light output at 254 nm. An understanding of the protective immune response elicited by RA cercaria may help in designing a candidate vaccine which is still needed.

Evaluation of the immunological effect of beta alanyl-l-histidine against Schistosoma mansoni antigens in rabbits

The influence of vaccination on healthy (non-infected) rabbits treated with Schistosoma mansoni egg antigens, cercariae, and worms as prophylactic agents against infection, and the benefit of beta alanyl-l-histidine treatment against Schistosoma mansoni infection were investigated. This study involved individual injection of three Schistosoma mansoni antigens: soluble egg antigen (SEA), cercarial antigen preparation (CAP) and soluble worm antigen preparation (SWAP), in three rabbit groups, respectively. Three other groups each received the same specific antigen in conjunction with the administration of (beta alanyl-l-histidine) L-carnosine. Hepatic total protein, glycogen and glycogen phosphorylase, total serum protein and one diminution electrophoresis of protein fractions (180 KDa; 116 KDa; 97, 4 KDa, serum albumin 66 KDa; 48. 5 KDa; 29 KDa; 18.400 KDa; 14.200 KDa; 6.5 KDa) were measured in all the rabbit groups. Elevation in most parameters was observed in the immunized groups. Carnosine treatment of rabbit groups immunized with SEA, CAP and SWAP in comparison to the non-carnosine-treated immunized groups resulted in amelioration of serum protein fractions in all SEA and SWAP-immunized animals, and reduction in glycogen phosphorylase b in SWAP animals alone. In addition, changes in glycogen content were observed in the CAP-immunized group. L-carnosine has a beneficial effect in the amelioration of most biochemical parameters as a result of S. mansoni antigen immunization.

Comparative sensitivities of various tests for diagnosing early Schistosoma mansoni infection in mice

Objective: We compared the diagnostic values of cercarial antigen preparation, cercarial secretions, soluble worm antigen preparation and worm vomit prepared from the parasite Schistosoma mansoni. Methods: Enzyme linked immunosorbant assay was used to detect IgG in plasma from Schistosoma mansoni infected mice. In parallel, specific primers for the parasite genome was used to detect S. mansoni DNA in plasma and urine from infected mice and hemolymph and tissues of infected Biomphalaria alexandrina snails by Polymerase chain reaction. Results: The results showed that all the above diagnostic approaches enabled infection to be diagnosed as early as three days post mice exposure to parasite cercariae. Conclusion: Cercarial secretions and worm vomit represent new useful economic crude antigens for preliminary detection of parasite active transmission or response to therapy in an endemic setting. Also, it was found that the detection of Schistosoma mansoni DNA in urine from infected mice was the most sensitive and specific (although expensive) method for infection diagnosis than all the others.

Immune Responses in Mice after Immunization with Antigens from Different Stages of the Parasite Schistosoma mansoni

Mice responses to immunization with Schistosoma mansoni antigens were investigated. Priming with cercarial antigen preparation (CAP) induced significant (P < 0.05) IgM, IgG, IgG2a, IgG2b, and IgA increases, while booster caused a significant IgG1 increase. A soluble worm antigen preparation (SWAP) caused significant IgG elevation. Priming with soluble egg antigen (SEA) caused significant IgM and IgG2a increases, while booster induced significant IgM, IgG and IgA increases. CAP-immunized mice sera (IMS) recognized CAP peptides ranging from 23-78 kDa. SWAP-IMS recognized SWAP peptides ranging from 40-75 kDa. SEA-IMS recognized SEA peptides ranging from 33-101 kDa. The cross-reactive peptides among the 3 antigens were identified. CAP caused significant increases in mesenteric lymph nodes (MLNs) CD(4,8)+, B lymphocytes, CD8+ thymocytes, CD4+ T and B splenocytes. SWAP priming caused significant increases in MLNs CD(4,8)+ thymocytes and B splenocytes. SWAP booster caused significant increases in MLNs CD8+ T and B lymphocytes, CD(4,8)+ thymocytes and CD4+ T and B splenocytes. SEA caused significant increase in CD4+ T cells.

Development of a new assay for the diagnosis of schistosomiasis, using cercarial antigens

We have developed a new ELISA for detection of anti-schistosome antibodies using an extract from Schistosoma mansoni cercariae. We evaluated the new assay on serum samples sent to the Hospital for Tropical Diseases, Department of Clinical Parasitology, London, UK, by comparing it with our routinely used S. mansoni soluble egg antigen (SEA) assay. We also evaluated the new assay for cross-reactivity with a number of helminth and other infections. We demonstrate that the cercarial antigen assay is equivalent to the SEA assay for serodiagnosis of schistosomiasis in a non-endemic setting. The cercarial antigen preparation is more easily produced than SEA, and for this reason this assay may be preferred for routine clinical use and may be amenable to scaling up. Further assessment is needed before it can be recommended for use in an endemic area, as chronic disease and co-infection with other helminths are likely to be under-represented in our sample set.

Escherichia coli shares T- and B-lymphocyte epitopes with Schistosoma mansoni

In this study, we tested the cross-reaction between crude Escherichia coli antigen (ECA) and 3 crude Schistosoma mansoni antigens. The schistosomal antigens used were cercarial antigen preparation (CAP), soluble worm antigen preparation (SWAP), and soluble egg antigen (SEA). Four groups each of 3 mice received 2 intraperotineal immunizations with the above-mentioned antigens at a two-week interval. The dose of the ECA was 20 microg/100 microl PBS/mouse and that of any of the used schistosomal antigens was 50 microg/100 microl PBS/mouse. IgM and IgG reactivities and cross-reactivities were tested in individual immunized mice sera (IMS) against the above-mentioned antigens by ELISA and Western blotting. The changes in the B, CD4+ and CD8+ -T cells' counts post immunization were recorded. Priming with ECA caused significant increases in IgM (P<0.05) against CAP and SWAP, while both priming and boosting with ECA caused a significant elevation in the IgG only against SWAP. Priming and boosting with ECA or schistosomal antigens caused significant increases in IgM against ECA. Priming with ECA or SWAP caused significant elevation in IgG against ECA. In Western blotting, ECA-IMS recognized 16, 33, 38 and 94 kDa ECA peptides that cross-reacted with CAP-IMS. ECA peptides at 30 and 38 kDa cross-reacted among ECA, SWAP and SEA-IMS. CAP peptides at 40, 71, 85 and 97 kDa cross-reacted with ECA-IMS. A 59 kDa SWAP peptide cross-reacted with ECA. SEA peptides at approximately 55, 96 and 101 kDa cross-reacted with ECA-IMS. Immunization with ECA, CAP, SWAP or SEA caused significant increases in mesentric lymph nodes (MLN)-CD4+, CD8+ -T cells and MLN-B cells. For thymocytes, CD4+ -T cells significantly increased upon immunization with ECA and SWAP while CD8+-T cells significantly increased upon immunization with SWAP. It is necessary to include E. coli antigens as controls while establishing schistosomal antigens-based diagnostic tests to ensure the specificity of the detected immune responses. Characterization of the cross-reactive ECA antigens with protective potential against S. mansoni infection remains a future research objective.

The blocking of human antibody-dependent, eosinophil-mediated killing of Schistosoma mansoni schistosomula by monoclonal antibodies which cross-react with a polysaccharide-containing egg antigen.

Three IgM monoclonal antibodies, M22G11P, M7B7 and M22P3G, which reacted with the surface of Schistosoma mansoni schistosomula in an indirect fluorescent antibody assay, were found to recognize a polysaccharide-containing egg antigen previously designated K3. The monoclonal antibodies and a monospecific rabbit anti-K3 serum also recognized a cross-reacting antigen in a crude cercarial antigen preparation. In an eosinophil-mediated schistosomulum killing assay, all three monoclonal antibodies significantly inhibited the level of killing produced by human infection serum. An IgG3 monoclonal antibody, M22C1C, which also recognized the egg antigen K3, did not inhibit eosinophil-mediated killing. However, when lower concentrations of human serum were used in the assay, this monoclonal antibody significantly enhanced the level of killing, despite having no capacity to induce eosinophil-mediated damage in the absence of human infection serum. On the basis of these and other results we suggest the possibility that antibodies to S. mansoni egg antigens which cross-react with the surface of the early post-penetration schistosomulum may influence the effective expression of antibody-dependent, eosinophil-mediated effector mechanisms in human infections.

Early Diagnosis of Schistosoma mansoni in Mice Using Assays Directed against Cercarial Antigens Isolated by Hydrophobic Chromatography

Two diagnostic assays are described for the early diagnosis of acute schistosomiasis, using a defined cercarial antigen preparation obtained by hydrophobic chromatography. Circulating IgM antibodies against this antigen fraction could be detected by ELISA as early as 1 wk after exposure in experimentally infected mice; IgM levels against other antigens and IgG levels against all the preparations examined were not significantly elevated until approximately 4-5 wk postinfection. Circulating antigen was detected as early as 3 days after exposure by a competitive inhibition ELISA using rabbit serum prepared against the cercarial antigen; antigen levels in the serum of mice with a 100-worm infection were found to exceed 100 ng/ml. Studies using sera from infected humans indicate that the assay can also recognize chronic S. mansoni, S. haematobium or S. japonicum infections. In a very limited field study, the specificity of the circulating antigen assay with regard to other helminthic infections was found to be 85%; sensitivity 100%. Preliminary characterization of the relevant antigen indicates that it is a relatively hydrophobic polypeptide with a molecular weight of approximately 41,000 daltons. The implications of these findings with regard to the treatment of travelers or the conduct of seroepidemiological studies in endemic areas are discussed.

Immunocytochemical Studies on Schistosoma mansoni. II. Soluble Cercarial Antigens in Cercariae and Schistosomules

The localization of soluble, cercarial antigen preparation (CAP) in cercariae and schistosomules of Schistosoma mansoni was performed at the light and electron microscope levels using the unlabeled antibody method. Reaction produce was observed associated with the contents of the pre- and postacetabular glands and with the filamentous coat of mature cercariae. No reaction product was observed associated with the glycoacalyx of schistosomules. However, several schistosomules did retain remnants of their filamentous coats and reaction product was observed associated with those remains. CAP components were also observed in the area surrounding the intrasporocyst cercariae.

Induction of cellular and humoral immunological responsiveness to a soluble cercarial antigen preparation from Schistosoma mansoni.

The development of immunological responsiveness to a soluble cercarial antigenic preparation (CAP) from Schistosoma mansoni was analyzed in inbred CBA/J mice infected with cercariae, one or multiple times, or sensitized using CAP. Repeated exposure to 75 cercariae at three weekly intervals (3X-75) or subcutaneous administration of 20 mug of CAP (CAP/complete Freund adjuvant [CFA]) stimulated the development of specific anti-CAP lymph node cell blastogenesis. The degree of responsiveness was dependent upon the concentration of CAP in the culture system and was optimal in the dose range of 20 to 30 mug of protein of CAP per culture. Animals exposed once to 75 or 225 cercariae or to two sequential weekly infections with 75 cercariae exhibited a minimal response to CAP in comparison to the responsiveness of 3X-75 or CAP/CFA lymph node cells. Assessment of anti-CAP agglutinating antibody by application of a microtiter passive hemagglutination technique revealed that both 3X-75 and CAP/CFA animals possessed low titers of activity. In addition, both 3X-75 and CAP/CFA sera contained reagin-like antibodies to CAP as detected by the heterologous (rat), 72-h latent period, passive cutaneous anaphylaxis technique.