Ki-67 is expressed in multiplying forms of Schistosoma mansoni, but not in snail host tissues

Ki-67 is a protein expressed in the nucleus of several species during cell-division, being absent during the GO resting phase of the cellular cycle. During attempts to disclose mitosis in the so-called " amebocyte-producing organ " in Biomphalaria glabrata infected with Schistosoma mansoni, the parasite multiplying forms appeared strongly marked for Ki-67, while the snail tissues were completely negative. These data are worth registering to complement general data on Ki-67, and to help future studies on the relationship of the parasite and of its intermediate host.

Reduced protective effect of Plasmodium berghei immunization by concurrent Schistosoma mansoni infection

Studies on concomitant schistosomiasis and human and experimental malaria have shown a variation in the immunospecific response, as well as an increase in the severity of both parasitoses. In the present study, a murine co-infection model was used to determine the effects of a co-infection with Schistosoma mansoni and Plasmodium berghei on the protective immunity acquired by repeated malarial infections and subsequent curative treatment with chloroquine. Our results have demonstrated that, compared to an infection with P. berghei only, the co-infection increases the malarial parasitaemia and decreases the survival rate. Indeed, mice that were immunized by infection and treatment with drug displayed no mortality whereas co-infected mice showed a reduced protective efficacy of immunization against P. berghei (mortality > 60%). Interestingly, this high mortality rate was not associated with high levels of parasitaemia. Our findings support the idea of a suppressive effect of a Schistosoma co-infection on the anti-malarial protection by immunization. This result reveals a possible drawback of the development of anti-malarial vaccines, especially considering the wide endemic areas for both parasitoses.

Further evaluation of an updated PCR assay for the detection of Schistosoma mansoni DNA in human stool samples

A previously reported sensitive PCR assay for the detection of Schistosoma mansoni DNA was updated and evaluated. Changes in the DNA extraction method, including the use of a worldwide available commercial kit and the inclusion of additional quality control measures, increased the robustness of the test, as confirmed by the analysis of 67 faecal samples from an endemic area in Brazil. The PCR assay is at hand as a proven, reliable diagnostic test for the control of schistosomiasis in specific settings.

Inheritance of Schistosoma mansoni infection incompatibility in Biomphalaria alexandrina snails

In this study, we looked at the inheritance of susceptibility and resistance to Schistosoma mansoni infection in the first generation of crossbred Biomphalaria alexandrina snails. Our ultimate goal is to use such information to develop a biological method of controlling schistosomiasis. We infected laboratory-bred snails with S. mansoni miracidia and examined cercarial shedding to determine susceptibility and resistance. Five parental groups were used: Group I contained 30 susceptible snails, Group II contained 30 resistant snails, Group III contained 15 susceptible and 15 resistant snails, Group IV contained 27 susceptible and three resistant snails and Group V contained three susceptible and 27 resistant snails. The percentage of resistant snails in the resulting progeny varied according to the ratio of susceptible and resistant parents per group; they are 7%, 100%, 68%, 45% and 97% from Groups I, II, III, IV and V, respectively. On increasing the percentage of resistant parent snails, the percentage of resistant progeny increased, while cercarial production in their susceptible progeny decreased.

Antibody reactivity against potato apyrase, a protein that shares epitopes with Schistosoma mansoni ATP diphosphohydrolase isoforms, in acute and chronically infected mice, after chemotherapy and reinfection

Schistosoma mansoni ATP diphosphohydrolase isoforms and potato apyrase share conserved epitopes. By enzyme-linked immunosorbent assays, elevated levels of IgM, IgG2a and IgG1 antibody reactivity against potato apyrase were observed in S. mansoni-infected BALB/c mice during the acute phase of infection, while only IgM and IgG1 antibody reactivity levels maintained elevated during the chronic phase of infection. Antibody reactivity against potato apyrase was monitored over an 11-month period in chronically-infected mice treated with oxamniquine. Eleven months later, the level of seropositive IgM decreased significantly (~30%) compared to the level found in untreated, infected mice. The level of seropositive IgG1 decreased significantly four months after treatment (MAT) (61%) and remained at this level even after 11 months. The IgG2a reactivity against potato apyrase, although unchanged during chronic phase to 11 MAT, appeared elevated again in re-infected mice suggesting a response similar to that found during the acute phase. BALB/c mouse polyclonal anti-potato apyrase IgG reacted with soluble egg antigens probably due to the recognition of parasite ATP diphosphohydrolase. This study, for the first time, showed that the IgG2a antibody from S. mansoni-infected BALB mice cross-reacts with potato apyrase and the level of IgG2a in infected mice differentiates disease phases. The results also suggest that different conserved-epitopes contribute to the immune response in schistosomiasis.

Omental and pleural milky spots: different reactivity patterns in mice infected with Schistosoma mansoni reveals coelomic compartmentalisation

In vertebrate animals, pleural and peritoneal cavities are repositories of milky spots (MS), which constitute an organised coelom-associated lymphomyeloid tissue that is intensively activated by Schistosoma mansoni infection. This study compared the reactive patterns of peritoneal MS to pleural MS and concluded from histological analysis that they represent independent responsive compartments. Whole omentum, lungs and the entire mediastinum of 54 S. mansoni-infected mice were studied morphologically. The omental MS of infected animals were highly activated, modulating from myeloid-lymphocytic (60 days of infection) to lymphomyeloid (90 days of infection) and lymphocytic or lymphoplasmacytic (160 days of infection) types. The non-lymphoid component predominated in the acute phase of infection and was expressed by monocytopoietic, eosinopoietic and neutropoietic foci, with isolated megakaryocytes and small foci of late normoblasts and mast cells. Nevertheless, pleural or thoracic MS of infected mice were monotonous, consisting of small and medium lymphocytes with few mast and plasma cells and no myeloid component. Our data indicate that compartmentalisation of the MS response is dependent on the lymphatic vascularisation of each coelomic cavity, limiting the effects or consequences of any stimulating or aggressive agents, as is the case with S. mansoni infection.

Carbohydrate metabolism alterations in Biomphalaria glabrata infected with Schistosoma mansoni and exposed to Euphorbia splendens var. hislopii latex

This paper evaluates the alterations in the glycogen content of tissues (digestive gland and cephalopedal mass) and glucose in the haemolymph of Biomphalaria glabrata BH strain infected with Schistosoma mansoni BH strain and exposed to the latex of Euphorbia splendens var. hislopii. A reduction in the glycogen deposits was observed in infected snails exposed and not exposed to latex. However, the exposure to latex caused a greater depletion of the glycogen levels in both sites analysed, especially from the third week onward. The utilisation of latex as a molluscicide to control the population of infected B. glabrata selectively is proposed.

Spatial distribution of Schistosoma mansoni infection before and after chemotherapy with two praziquantel doses in a community of Pernambuco, Brazil

Praziquantel chemotherapy has been the focus of the Schistosomiasis Control Program in Brazil for the past two decades. Nevertheless, information on the impact of selective chemotherapy against Schistosoma mansoni infection under the conditions confronted by the health teams in endemic municipalities remains scarce. This paper compares the spatial pattern of infection before and after treatment with either a 40 mg/kg or 60 mg/kg dose of praziquantel by determining the intensity of spatial cluster among patients at 180 and 360 days after treatment. The spatial-temporal distribution of egg-positive patients was analysed in a Geographic Information System using the kernel smoothing technique. While all patients became egg-negative after 21 days, 17.9% and 30.9% reverted to an egg-positive condition after 180 and 360 days, respectively. Both the prevalence and intensity of infection after treatment were significantly lower in the 60 mg/kg than in the 40 mg/kg treatment group. The higher intensity of the kernel in the 40 mg/kg group compared to the 60 mg/kg group, at both 180 and 360 days, reflects the higher number of reverted cases in the lower dose group. Auxiliary, preventive measures to control transmission should be integrated with chemotherapy to achieve a more enduring impact.

Factors related to transmission of and infection with Schistosoma mansoni in a village in the South-eastern Region of Brazil

In this transversal study, factors related to infection with and transmission of Schistosoma mansoni were explored. Based on stool examinations of two Kato-Katz smears of a single sample, the prevalences of schistosomiasis and geohelminths were established. In a multivariable analysis, sets of demographic, socio-economic and water contact pattern variables were tested for strength of relation with infection. Males presented a 3.39-times higher risk for infection than females. The age groups between 10-19 years and 20-30 years showed risks of infection 7.1- and 7.5-times higher, respectively, than the control age group between 0-10 years. Individuals practicing leisure activities had a 1.96-times higher risk than those without these activities. The malacological survey identified snails of the species Biomphalaria glabrata, Biomphalaria straminea and Biomphalaria tenagophila. Two exemplars of B. glabrata (0.53%) proved positive for S. mansoni. The socio-economic improvements observed in the locality suggest a protective and preventive effect towards infection with schistosomiasis, which requires further investigation with a longitudinal and more detailed study design. Considering our findings, a proposal for an integrated control program should be based on two pillars: one horizontal, which involves social empowerment and health education, and another more vertical, which delivers treatment and infrastructure improvements.

Immunological and parasitological parameters after treatment with dexamethasone in murine Schistosoma mansoni

This work aimed to evaluate the effect of diphenyl dimethyl bicarboxylate (DDB) and dexamethasone alone and in combination with praziquantel on various parasitological, immunological and pathological parameters reflecting disease severity and morbidity in murine schistosomiasis. DDB and dexamethasone had no effect on worm burden but altered tissue egg distribution. This indicates that, under the schedule used, neither drug interfered with the development of adult worms or oviposition, but both can modulate liver pathology. Dexamethasone resulted in a greater reduction in granuloma size than did DDB. Dexamethasone-treated mice also showed lower levels of serum gamma interferon (IFN-γ), interleukin-12 (IL-12) and IL-4, together with higher IL-10 levels, than infected untreated control animals. These data suggest that dexamethasone is a convenient and promising coadjuvant agent that results in decreased morbidity in murine schistosomiasis.

Biomphalaria alexandrina snails as immunogens against Schistosoma mansoni infection in mice

Despite effective chemotherapy, schistosomiasis remains the second largest public health problem in the developing world. Currently, vaccination is the new strategy for schistosomiasis control. The presence of common antigenic fractions between Schistosoma mansoni and its intermediate host provides a source for the preparation of a proper vaccine. The objective of this paper is to evaluate the nucleoprotein extracted from either susceptible or resistant snails to protect against schistosomiasis. The vaccination schedule consisted of a subcutaneous injection of 50 µg protein of each antigen followed by another inoculation 15 days later. Analyses of marker enzymes for different cell organelles [succinate dehydrogenase, lactate dehydrogenase (LDH), glucose-6-phosphatase, acid phosphatase and 5'-nucleotidase] were carried out. Energetic parameters (ATP, ADP, AMP, phosphate potentials, inorganic phosphate, amino acids and LDH isoenzymes) were also investigated. The work was extended to record worm and ova counts, oogram determination in the liver and intestine and the histopathological pattern of the liver. The nucleoprotein of susceptible snails showed reduction in worm and ova counts by 70.96% and 51.31%, respectively, whereas the nucleoprotein of resistant snails showed reductions of 9.67% and 16.77%, respectively. In conclusion, we found that the nucleoprotein of susceptible snails was more effective in protecting against schistosomiasis.

Immunostimulatory property of a synthetic peptide belonging to the soluble ATP diphosphohydro-lase isoform (SmATPDase 2) and immunolocalisation of this protein in the Schistosoma mansoni egg

A peptide (SmB2LJ; r175-194) that belongs to a conserved domain from Schistosoma mansoni SmATPDase 2 and is shared with potato apyrase, as predicted by in silico analysis as antigenic, was synthesised and its immunostimulatory property was analysed. When inoculated in BALB/c mice, this peptide induced high levels of SmB2LJ-specific IgG1 and IgG2a subtypes, as detected by enzyme linked immunosorbent assay. In addition, dot blots were found to be positive for immune sera against potato apyrase and SmB2LJ. These results suggest that the conserved domain r175-194 from the S. mansoni SmATPDase 2 is antigenic. Western blots were performed and the anti-SmB2LJ antibody recognised in adult worm (soluble worm antigen preparation) or soluble egg antigen antigenic preparations two bands of approximately 63 and 55 kDa, molecular masses similar to those predicted for adult worm SmATPDase 2. This finding strongly suggests the expression of this same isoform in S. mansoni eggs. To assess localisation of SmATPDase 2, confocal fluorescence microscopy was performed using cryostat sections of infected mouse liver and polyclonal antiserum against SmB2LJ. Positive reactions were identified on the external surface from the miracidium in von Lichtenberg's envelope and, in the outer side of the egg-shell, showing that this soluble isoform is secreted from the S. mansoni eggs.

Pharmacodynamics of mefloquine and praziquantel combination therapy in mice harbouring juvenile and adult Schistosoma mansoni

Praziquantel (PZQ) is currently the only drug widely used for the treatment of schistosomiasis, but the antimalarial drug mefloquine (Mef) possesses interesting antischistosomal properties. Combination therapy with these two drugs has been suggested as a strategy for transmission control, as PZQ is active against adult worms and Mef is active against schistosomula. To examine the efficacy of combination therapy, Schistosoma mansoni-reinfected mice were separated into seven groups: untreated (I), treated with PZQ in doses of 200 mg/kg (II) or 1,000 mg/kg (III), treated with Mef in doses of 200 mg/kg (IV) or 400 mg/kg (V); each dose was divided equally and given on two consecutive days. Group VI was treated with doses of PZQ + Mef as in groups II and IV, respectively, while group VII was treated with PZQ + Mef as in groups III and V, respectively. PZQ + Mef at the reduced doses of 200 mg/kg each enhanced the therapeutic efficacy over the reduced PZQ dose alone as shown by a very high reduction in the total numbers of mature worms (95% vs. 49%), immature worms (96% vs. 29%) and the complete eradication of immature females, mature females and immature eggs. The reduction in worm burden was associated with the healing of hepatic granulomatous lesions and the normalisation of all liver enzymes. Therefore, the use of Mef with PZQ is more effective than PZQ alone and should be considered for clinical trials in humans as a potential treatment regimen to prevent treatment failures in areas with high rates of schistosomiasis.

Schistosoma mansoni antigens alter the cytokine response in vitro during cutaneous leishmaniasis

Schistosoma mansoni infection or associated products are able to down-modulate the type 1 CD4+ T cell inflammatory response characteristic of autoimmune diseases. In this study, we evaluated how S. mansoni antigens altered the immune response that was induced by the soluble Leishmania antigen (SLA) from cutaneous leishmaniasis (CL) patients. Cytokines were measured from the supernatants of peripheral blood mononuclear cell cultures stimulated with SLA. This was performed using the sandwich enzyme linked immunosorbent assay technique in the presence or absence of S. mansoni recombinant antigens Sm29, SmTSP-2 and PIII. The addition of S. mansoni antigens to the cultures resulted in the reduction of interferon gamma (IFN-γ) levels in 37-50% of patients. Although to a lesser extent, the antigens were also able to decrease the production of tumour necrosis factor-alpha (TNF-α). We compared patients that either had or did not have reduction in IFN-γ and TNF-α production in cultures stimulated with SLA in the presence of S. mansoni antigens. We found that there was no significant difference in the levels of interleukin (IL)-10 and IL-5 in response to S. mansoni antigens between the groups. The antigens used in this study down-modulated the in vitro proinflammatory response induced by SLA in a group of CL patients through a currently undefined mechanism.

Participation of N-acetyl-D-glucosamine carbohydrate moieties in the recognition of Schistosoma mansoni sporocysts by haemocytes of Biomphalaria tenagophila

Lectin-carbohydrate binding may be involved in the recognition of Schistosoma mansoni sporocysts by haemocytes of Biomphalaria; therefore, we tested if this interaction is associated with snail resistance against Schistosoma infection. In vitro data showed that most of the S. mansoni sporocysts cultured with haemocytes from Biomphalaria glabrata BH, a highly susceptible snail strain, had a low number of cells that adhered to their tegument and a low mortality rate. Moreover, the addition of N-acetyl-D-glucosamine (GlcNAc) did not alter this pattern of adherence and mortality. Using haemocytes and haemolymph of Biomphalaria tenagophila Cabo Frio, we observed a high percentage of sporocysts with adherent cells, but complete encapsulation was not detected. Low concentrations of GlcNAc increased haemocyte binding to the sporocysts and mortality, which returned to basal levels with high concentrations of the carbohydrate. In contrast, haemocytes plus haemolymph from B. tenagophila Taim encapsulated cellular adhesion index of level 3 and destroyed over 30% of the S. mansoni sporocysts in culture. Interestingly, the addition of GlcNAc, but not mannose, to the culture medium resulted in the significant inhibition of cellular adhesion to the parasite tegument and the reduction of parasite mortality, suggesting that GlcNAc carbohydrate moieties are important to the recognition of S. mansoni by B. tenagophila Taim.