Ronald E. Blanton

Repeated Exposure Induces Periportal Fibrosis in Schistosoma mansoni-Infected Baboons: Role of TGF-β and IL-4

Recently, we observed that repeated Schistosoma mansoni infection and treatment boost Th2-associated cytokines and TGF-β production in baboons. Other studies have shown that some chronically infected baboons develop hepatic fibrosis. Because TGF-β, IL-2, and IL-4 have been shown to participate in development of fibrosis in murine schistosomiasis, the present study examined whether repeated exposure stimulates hepatic fibrosis in olive baboons. To test this hypothesis, animals were exposed to similar numbers of S. mansoni cercariae given once or repeatedly. After 19 wk of infection, animals were cured with praziquantel and reinfected once or multiple times. Hepatic granulomatous inflammation and fibrosis were assessed from serial liver biopsies taken at weeks 6, 9, and 16 after reinfection and egg Ag (schistosome egg Ag)-specific cytokine production by PBMC were measured simultaneously. Periportal fibroblast infiltration and extracellular matrix deposition (fibrosis), angiogenesis, and biliary duct hyperplasia developed in some animals. The presence and amount of fibrosis directly correlated with the frequency of exposure. Fibrosis was not associated with adult worm or tissue egg burden. The amount of fibrosis correlated with increased schistosome egg Ag-driven TGF-β at 6, 9, and 16 wk postinfection (rs = 0.9, 0.8, and 0.54, respectively, all p < 0.01) and IL-4 production (p = 0.02) at 16 wk postinfection and not IFN-γ, IL-2, IL-5, or IL-10. These data suggest that repeated exposure is a risk factor for periportal fibrosis by a mechanism that primes lymphocytes to produce increased levels of profibrotic molecules that include TGF-β and IL-4.

Allergies and Diabetes as Risk Factors for Dengue Hemorrhagic Fever: Results of a Case Control Study

Background The physiopathology of dengue hemorrhagic fever (DHF), a severe form of Dengue Fever, is poorly understood. We are unable to identify patients likely to progress to DHF for closer monitoring and early intervention during epidemics, so most cases are sent home. This study explored whether patients with selected co-morbidities are at higher risk of developing DHF. Methods A matched case-control study was conducted in a dengue sero-positive population in two Brazilian cities. For each case of DHF, 7 sero-positive controls were selected. Cases and controls were interviewed and information collected on demographic and socio-economic status, reported co-morbidities (diabetes, hypertension, allergy) and use of medication. Conditional logistic regression was used to calculate the strength of the association between the co-morbidities and occurrence of DHF. Results 170 cases of DHF and 1,175 controls were included. Significant associations were found between DHF and white ethnicity (OR = 4.70; 2.17–10.20), high income (OR = 6.84; 4.09–11.43), high education (OR = 4.67; 2.35–9.27), reported diabetes (OR = 2.75; 1.12–6.73) and reported allergy treated with steroids (OR = 2.94; 1.01–8.54). Black individuals who reported being treated for hypertension had 13 times higher risk of DHF then black individuals reporting no hypertension. Conclusions This is the first study to find an association between DHF and diabetes, allergy and hypertension. Given the high case fatality rate of DHF (1–5%), we believe that the evidence produced in this study, when confirmed in other studies, suggests that screening criteria might be used to identify adult patients at a greater risk of developing DHF with a recommendation that they remain under observation and monitoring in hospital.

Genetic ancestry and income are associated with dengue hemorrhagic fever in a highly admixed population

To test whether African ancestry is protective for severe dengue, we genotyped 49 hospitalized cases of dengue hemorrhagic fever (DHF) as well as 293 neighborhood cases of dengue fever and 294 asymptomatic controls in Salvador, Bahia, Brazil. Ancestry-informative markers and 282 unlinked SNPs not associated with the clinical presentation of dengue were used to estimate ancestry. After controlling for income, both self-defined Afro-Brazilian ethnicity and African ancestry were protective for DHF (P=0.02, OR=0.28 and P=0.02, OR=0.13, respectively). Income or an index of income indicators, however, was also independently associated with the diagnosis of DHF.

Characterization of a native and recombinant Schistosoma haematobium serine protease inhibitor gene product.

Immunologic screening of a Schistosoma haematobium cDNA library with species-specific human antisera identified a clone whose predicted amino acid sequence encodes a member of the serine protease inhibitor (serpin) gene family. This cDNA consists of 1397 bp with a single open reading frame that can encode a 409-amino acid protein of 46,261 Da. The native antigen is a 54-58-kDa glycoprotein and is located on the surface of adult worms. Sequence comparison with other serpins predicts the amino acid Phe at the putative reactive center of the protein. Phenylalanine is also found at the corresponding site of a vaccinia serpin that may contribute to the hemorrhagic phenotype of some strains of cowpox virus. Though the human parasites S. haematobium and Schistosoma mansoni demonstrate a close antigenic relationship, the S. haematobium serpin gene product demonstrates marked species-specific immunogenicity. By Northern blot hybridization, however, both species express a 1700-nucleotide mRNA that hybridizes with the S. haematobium serpin cDNA. The intensity of cross hybridization for the S. mansoni mRNA is 10-fold lower than that for S. haematobium. Southern blots of genomic DNA and gene titration experiments indicate that the S. haematobium gene is present in approximately 4-5 copies per haploid genome.

Schistosomal hepatic fibrosis and the interferon gamma receptor: a linkage analysis using single-nucleotide polymorphic markers

A minority of individuals infected with the parasite Schistosoma mansoni develops hepatic fibrosis. HLA studies in Egypt and a candidate gene search in a Sudanese population indicate that the hosts genetics contribute to disease susceptibility. In an Egyptian community, 32.7% of individuals 11 years and older had significant fibrosis by WHO ultrasound criteria. Linkage to 10 candidate genes was tested using 89 affected sibling pairs from 40 pedigrees in this community. The candidates included genes that initiate fibrosis, participate in collagen synthesis, or control collagen degradation. Two to four single-nucleotide polymorphisms (SNPs) were genotyped per locus, and 188 individuals were genotyped at 48 markers. Model-free modified Haseman–Elston analysis identified linkage to a SNP in the interferon gamma receptor locus (P=0.000001). There was also weak evidence for linkage to the interleukin 13-4 region and tissue growth factor beta 1.

Stage and sex specific differences in actin gene expression in Schistosoma mansoni

We have characterized actin gene expression in Schistosoma mansoni at the RNA and protein levels. Northern blot analyses showed two size classes of actin mRNA in eggs, cercariae and adult worms of both sexes, approximately 1 900 and 1 400 bases in length. A higher abundance of actin mRNA of both size classes was demonstrated in male worms than in eggs, cercariae, and females. Using a phalloidin-rhodamine conjugate, male worms were observed to contain more actin protein than females. Southern blot-hybridization indicated that the sexual differences in actin mRNA and protein levels were not related to some S. mansoni actin genes being sex linked. In addition, two other trematodes, Schistosoma japonicum and Fasciola hepatica and a cestode, Taenia pisiformis contained two classes of actin mRNA similar in size as those in S. mansoni. In contrast, a turbellarian, Dugesia tigrina contained only a single short actin message size class approximately 1 400 bases in length.

Childhood stunting in Northeast Brazil: The role of Schistosoma mansoni infection and inadequate dietary intake

Objective: To evaluate the relationship between stunting, Schistosoma mansoni infection and dietary intake in schoolchildren.Design: This is a cross-sectional study. Two stool samples were obtained from each child and examined quantitatively for the presence of S. mansoni, Ascaris lumbricoides and Trichuris trichiuria eggs. Information on dietary intake, and demographic, biologic and socioeconomic variables was elicited during the in-home survey. Logistic regression was used to evaluate the association between stunting (height for age <−2s.d.), parasitic infection and food consumption.Setting: The study was carried out in the city of Nazaré, located in the Recôncavo region of the State of Bahia, northeastern Brazil.Subjects: The sample consisted of 461 children 7–14 y old, 228 boys and 233 girls, recruited from public schools.Results: Of the children studied, 55.1% presented with S. mansoni infection and 22.1% were stunted. The median protein, lipid and carbohydrates intake were 47.8, 36.0 and 248.2 g/day, respectively. The median caloric consumption was 1527.0 kcal (6388.97 kJ/day). The analysis indicated that children heavily infected (≥400 eggs/g of stool) with S. mansoni had a 2.74-fold (95% CI: 1.32–5.67) higher risk of stunting than uninfected children, and those with inadequate intake of lipid (<36 g/day) had a 1.83-fold (95% CI: 1.05–3.20) increased risk of stunting compared to those with adequate diets.Conclusion: Heavy S. mansoni infection and inadequate dietary intake of fat in schoolchildren play a significant and independent role in the development of stunting. This meaning that nutritional interventions in this age group in S. mansoni endemic areas must include the diagnosis and treatment of the infection associated with dietary measures.Sponsorship: This study was supported by the Thrasher Foundation. Sandra Maria Conceição Pinheiro is a National Council on Technological Development Scholarship Awardee (CNPq), #302228/81-0.

Fatores de risco para anemia por deficiência de ferro em crianças e adolescentes parasitados por helmintos intestinais

OBJETIVO: Investigar os fatores de risco para anemia por deficiencia de ferro em criancas e adolescentes (7 a 17 anos) infectados por helmintos. METODOS: Foi realizado um estudo transversal com 1709 criancas e adolescentes residentes na cidade de Jequie, Estado da Bahia, Brasil, que apresentavam infeccao leve ou moderada por Schistosoma mansoni, Ascaris lumbricoides, Trichuris trichiura e ancilostomideos. Foram obtidos dados sobre niveis de hemoglobina (hemoglobinometro portatil), consumo alimentar (inquerito recordatorio de 24 horas), infeccao parasitaria (metodo Kato-Katz), condicoes ambientais e domiciliares, renda e escolaridade dos responsaveis. Os fatores de risco para anemia na populacao foram estudados com base em um modelo hierarquico de causalidade. RESULTADOS: A prevalencia de infeccao por T. trichiura, A. lumbricoides, S. mansoni e ancilostomideos foi de 74,8, 63,0, 55,5 e 15,7%, respectivamente. Constatou-se que 32,2% das criancas e adolescentes eram anemicos. Depois do ajuste para variaveis de confusao, os resultados da analise multivariada mostraram que a renda familiar per capita abaixo de um quarto do salario minimo (27 dolares), o sexo masculino, a faixa etaria de 7 a 9 anos e a ingestao inadequada de ferro biodisponivel foram significativamente associados a anemia. CONCLUSOES: As acoes para controle da anemia no grupo de maior risco, conforme identificado no presente estudo, devem visar o aumento do consumo de alimentos ricos em ferro e da biodisponibilidade do ferro ingerido, bem como a melhoria das condicoes socio-ambientais.

Lower Hookworm Incidence, Prevalence, and Intensity of Infection in Children with a Bacillus Calmette-Guerin Vaccination Scar

Bacillus Calmette-Guérin (BCG), the most common vaccine worldwide, has broad effects on the immune system. Hookworm infections are a major source of morbidity. In response to a preliminary report of BCG vaccination protection against nematodes in human immunodeficiency virus-infected adults, data from an ongoing prospective study were analyzed to determine the intensity (eggs per gram of stool), prevalence, and incidence of different helminths in children with and without a BCG vaccination scar. Adjusted prevalence and incidence ratios were estimated by using logistic regression. Children with a BCG vaccination scar were found to have statistically significantly lower hookworm prevalence (41%), incidence (37%), and mean egg counts (39%), after controlling for age, sex, and socioeconomic factors. There was no BCG association with incidence, prevalence, or intensity of infection with Schistosoma mansoni, Ascaris lumbricoides, or Trichuris trichiura. Such protection would have implications for public health and for research on mechanisms behind human immunological responses to hookworm.

Localization and pattern of expression of a female specific mRNA in Schistosoma mansoni

To understand mechanisms involved in sex-specific gene expression in Schistosoma mansoni, a cDNA (fs800) was isolated that hybridized to an 800 nucleotide mRNA present in high levels only in mature female worms. The fs800 cDNA sequence was characterized by two long open reading frames and central stretches of repeated amino acids. Fs800 did not share similarities with other known sequences in computer searches. In situ hybridization, however, revealed that the mRNA corresponding to fs800 was found only in female vitelline cells, suggesting that the product of this gene may be involved in the production or function of eggs. Fs800 is developmentally regulated as expression of this gene is dependent on the maturity of female worms. Furthermore, during in vitro culture, when female worms are known to stop egg production, expression of fs800 selectively ceased.