Roberto Sena Rocha

Analysis of anti-keyhole limpet haemocyanin antibody in Brazilians supports its use for the diagnosis of acute schistosomiasis mansoni

Antibody (immunoglobulin (Ig) G) to the haemocyanin of the keyhole limpet (KLH) (Megathura crenulata), which shares a well defined carbohydrate epitope with the surface of schistosomula of Schistosoma mansoni, was determined by enzyme-linked immunosorbent assay (ELISA) in the sera of Brazilians with acute schistosomiasis. Of 53 such individuals tested, 51 had a level of KLH reactivity in excess of the mean +2 standard deviations of that exhibited by chronically infected individuals. This difference in reactivity allowed the acute cases to be readily identified by visual inspection of ELISA plates. The levels of IgG in patients with hepatointestinal and hepatosplenic schistosomiasis, as well as in non-infected, seropositive residents of endemic areas and infected children from endemic areas, were not statistically different from those of intestinal patients. Significant levels of anti-KLH IgG were not detected in patients with leishmaniasis, Chagas disease, ancylostomiasis or ascariasis. The results support the use of KLH as a means of rapidly and easily identifying individuals with acute schistosomiasis.

The control of the schistosome-transmitting snail Biomphalaria glabrata by the plant Molluscicide Euphorbia splendens var. hislopii (syn milli Des. Moul): a longitudinal field study in an endemic area in Brazil

Under laboratory conditions, latex from Euphorbia splendens has shown promise as a plant molluscicide for control of Biomphalaria species, intermediate hosts for Schistosoma mansoni. The purpose of this study was to evaluate its efficiency under field conditions. Application of filtered latex at 12 ppm to one stream in an endemic rural area in Minas Gerais state, Brazil, in September 1995, did result in a reduction in snail density as compared to an untreated stream but the snail population recovered quickly. However, two applications with a two-week interval of unfiltered E. splendens latex at 5 ppm in November 1996 in the same stream resulted in complete disappearance of B. glabrata and snails did not reappear until the 14th month after the applications. In the control stream, without treatment, the snails were found during all months. Laboratory studies confirmed that unfiltered latex is a more potent molluscicide than filtered latex. Considering the advantages of the latex such as its low toxicity to other aquatic animals and its photobiodegradability, as well as the simple method of application, this natural product is promising as an effective molluscicide.

A multivariate analysis of socio-demographic factors, water contact patterns and Schistosoma mansoni infection in an endemic area in Brazil

Associations between socio-demographic factors, water contact patterns and Schistosoma mansoni infection were investigated in 506 individuals (87% of inhabitants over 1 year of age) in an endemic area in Brazil (Divino), aiming at determining priorities for public health measures to prevent the infection. Those who eliminated S. mansoni eggs (n = 198) were compared to those without eggs in the stools (n = 308). The following explanatory variables were considered: age, sex, color, previous treatment with schistosomicide, place of birth, quality of the houses, water supply for the household, distance from houses to stream, and frequency and reasons for water contact. Factors found to be independently associated with the infection were age (10-19 and > or = 20 yrs old), and water contact for agricultural activities, fishing, and swimming or bathing (Adjusted relative odds = 5.0, 2.4, 3.2, 2.1 and 2.0, respectively). This suggests the need for public health measures to prevent the infection, emphasizing water contact for leisure and agricultural activities in this endemic area.

Evaluation of the molluscicidal properties of Euphorbia splendens var. hislopii (N.E.B.) latex: experimental test in an endemic area in the State of Minas Gerais, Brazil.

Following the positive results obtained regarding the molluscicidal properties of the latex of Euphorbia splendens that were corroborated in laboratory and field tests under restricted conditions, a field study was conducted in experimental streams located in an endemic area. After recording the average annual fluctuations of vectors in three streams, a solution of E. splendens latex at 12 ppm was applied in stream A, a solution of niclosamide at 3 ppm that was applied in stream B and a third stream (C) remained untreated for negative control. Applications of E. splendens and niclosamide resulted in a mortality of 100% among the snails collected in the streams A and B. No dead snails were found in the negative control stream. A monthly follow-up survey conducted during three consecutive months confirmed the return of vectors to both experimental streams treated with latex and niclosamide. This fact has called for a need to repeat application in order to reach the snails that remained buried in the mud substrate or escaped to the water edge, as well as, newly hatched snails that did not respond to the concentration of these molluscicides. Adults snails collected a month following treatment led us to believe that they had migrate from untreated areas of the streams to those previously treated.

Production and characterization of human monoclonal antibodies against Schistosoma mansoni

Summary We have produced a panel of human monoclonal antibodies (MoAb) from patients infected with Schistosoma mansoni in order to analyse more carefully the human immune response to this helminth infection. This study describes the production, characterization and analysis of these MoAbs. Briefly, peripheral blood mononuclear cells from chronically infected patients were (1) isolated and stimulated with parasite antigens in vitro, (2) positively selected for B–cells on anti–Ig columns, and (3) then transformed with Epstein–Barr virus (EBV). Once EBV cell lines were established, they were selected for anti–5. mansoni antibodies using an ELISA, cloned, retested and then fused with the mouse–human heteromyeloma SHM–D33. In this study, we describe five MoAbs which have different antigenic specificities for life–cycle stages based on ELISA to soluble crude antigen preparations, membrane immunofluorescence on whole intact organisms, and immunofluorescent staining of cryostat frozen sections. The importance of these reagents with regard to the human immune response to S. mansoni is currently being evaluated.

Dot-dye-immunoassay and dot-ELISA for the serological differentiation of acute and chronic schistosomiasis mansoni using keyhole limpet haemocyanin as antigen

Two immunoassays, dot enzyme-linked immunosorbent assay (dot-ELISA) and dot-dye immunoassay (dot-DIA), using soluble egg antigen and keyhole limpet haemocyanin as antigens, were evaluated for the serological differentiation of 25 acute and 37 chronic patients infected with Schistosoma mansoni and 20 non-infected individuals, in comparison with ELISA. Efficiency was 92.7%, 90.0% for ELISA, dot-ELISA and dot-DIA, respectively. Dipstick dot-ELISA and dot-DIA are described and shown to be reliable cheap and simple methods for the serological differentiation of acute and chronic schistosomiasis.

Granulomatous hypersensitivity to Schistosoma mansoni egg antigens in human Schistosomiasis: I. Granuloma formation and modulation around polyacrylamide antigen-conjugated beads

We have developed an in vitro model of granuloma formation for the purpose of studying the immunological components of delayed type hypersensitivity granuloma formation in patients infected with Schistosoma mansoni. Our data show that 1) granulomatous hypersensitivity can be studied by examining the cellular reactivity manifested as multiple cell layers surrounding the antigen conjugated beads; 2) this reactivity is a CD4 cell dependent, macrophage dependent, B cell independent response and 3) the in vitro granuloma response is antigenically specific for parasite egg antigens. Studies designed to investigate the immune regulation of granulomatous hypersensitivity using purified populations of either CD4 or CD8 T cells have demonstrated the complexity of cellular interactions in the suppression of granulomatous hypersensitivity. The anti-S. mansoni egg immune responses of individual patients with chronic intestinal schistosomiasis can be classified either as soluble egg antigen (SEA) hypersensitive with maximal granulomatous hypersensitivity or SEA suppressive with activation of the T cell suppressor pathway with effective SEA granuloma modulation. Our data suggest that T cell network interactions are active in the generation of effective granuloma modulation in chronic intestinal schistosomiasis patients.

QUESTIONNAIRES IN THE SCREENING FOR Schistosoma mansoni INFECTION: A STUDY OF SOCIO DEMOGRAPHIC AND WATER CONTACT VARIABLES IN FOUR COMMUNITIES IN BRAZIL

The use of questionnaires has been recommended for identifying, at a lower cost, individuals at risk for schistosomiasis. In this study, validity of information obtained by questionnaire in the screening for Schistosoma mansoni infection was assessed in four communities in the State of Minas Gerais, Brazil. Explanatory variables were water contact activities, sociodemographic characteristics and previous treatment for schistosomiasis. From 677, 1474, 766 and 3290 individuals eligible for stool examination in the communities, 89 to 97% participated in the study. The estimated probability of individuals to be infected, if they have all characteristics identified as independently associated with S.mansoni infection, varied from 15% in Canabrava, to 42% in Belo Horizonte, 48% in Comercinho and 80% in São José do Acácio. Our results do not support the hypothesis that a same questionnaire on risk factors could be used in screening for S.mansoni infection in different communities.

Plasmodium vivax Duffy binding protein: baseline antibody responses and parasite polymorphisms in a well-consolidated settlement of the Amazon Region

Objective To investigate risk factors associated with the acquisition of antibodies against Plasmodium vivax Duffy binding protein (PvDBP) – a leading malaria vaccine candidate – in a well‐consolidated agricultural settlement of the Brazilian Amazon Region and to determine the sequence diversity of the PvDBP ligand domain (DBPII) within the local malaria parasite population.

Stool examination and rectal biopsy in the diagnosis and evaluation of therapy of schistosomiasis mansoni

Em um grupo de 217 adultos do sexo masculino selecionados atraves de ELISA ou intradermoreacao, realizaram-se seis exames de fezes pelos metodos de Lutz/Hoffman, Pons e Janer (Lutz/HPJ) e Kato/Katz e um oograma da mucosa retal, observando-se positividade em 44,7%, 47,5% e 40,1%, respectivamente. A analise comparativa dos resultados mostrou: 1) aumento da sensibilidade de ambos os metodos de exames de fezes ate a terceira amostra examinada; 2) maior sensibilidade da biopsia retal quando comparada ao exame de uma unica amostra fecal pelos metodos de Lutz/HPJ ou Kato/Katz; 3) igual sensibilidade pelo exame de duas a cinco amostras fecais pelo metodo de Kato/Katz ou duas a seis amostras pelo metodo de Lutz/HPJ e a biopsia retal; 4) sensibilidade superior de seis amostras fecais pelo metodo de Kato/Katz, quando comparado a biopsia retal; 5) igual sensibilidade entre o mesmo numero de amostras examinadas pelos metodos de Lutz/HPJ e Kato/Katz. Para avaliar a cura parasitologica, os ultimos 40 pacientes do grupo A, tratados com oxamniquine oral, dose unica, 15 mg/kg foram acompanhados durante seis meses (Grupo B). O controle de cura incluiu tres exames de fezes mensais, pelos metodos de Lutz/HPJ e Kato/Katz no 1o, 3o e 6o meses e uma biopsia retal, entre o 4o e 6o meses apos o tratamento. Os exames foram negativos em 87,5%, 90,0% e 95,0% dos pacientes tratados e avaliados pelos metodos de Kato/Katz, Luiz/HPJ e biopsia retal, respectivamente. O percentual de cura com a oxamniquine foi de 82,5%, quando considerados os tres metodos de controle parasitologico associados e, nao houve diferenca estatistica entre o numero de casos considerados curados por um deles.From each of a group of 217 adult males selected through enzyme-immunoassay or skin-test (Group A), six stool samples were examined by both the Lutz/Hoffman, Pons & Janer (Lutz/HPJ) and Kato/Katz methods. In addition, one oogram of the rectal mucosa was performed. By these methods, schistosomiasis was detected in 44.7%, 47.5% and 40.1% of the individuals respectively. To evaluate the methods in the assessment of cure, the last 40 patients from group A, treated with a single oral dose of oxamniquine at 15 mg/kg were followed up for six months (Group B). The criteria for parasitological cure included three stool examinations by Kato/Katz and Lutz/HPJ methods, one, three and six months post-treatment and a rectal biopsy between the fourth and sixth months post-treatment. The examinations were negative in 87.5%, 90% and 95% of the patients, respectively. The efficacy of oxamniquine was 82.5% when the three methods were considered together and there was no statistically significant difference between the sensitivity of the individual methods.