Fit for genomic and proteomic purposes: Sampling the fitness of nucleic acid and protein derivatives from formalin fixed paraffin embedded tissue.

The demand for nucleic acid and protein derivatives from formalin-fixed paraffin-embedded (FFPE) tissue has greatly increased due to advances in extraction and purification methods, making these derivatives available for numerous genomic and proteomic platforms. Previously, DNA, RNA, microRNA (miRNA), or protein derived from FFPE tissue blocks were considered "unfit" for such platforms, as the process of tissue immobilization by FFPE resulted in cross-linked, fragmented, and chemically modified macromolecules. We conducted a systematic examination of nucleic acids and proteins co-extracted from 118 FFPE blocks sampled from the AIDS and Cancer Specimen Resource (ACSR) at The George Washington University after stratification by storage duration and the three most common tumor tissue types at the ACSR (adenocarcinoma, squamous cell carcinoma, and papillary carcinoma). DNA, RNA, miRNA, and protein could be co-extracted from 98% of the FFPE blocks sampled, with DNA and miRNA "fit" for diverse genomic purposes including sequencing. While RNA was the most labile of the FFPE derivatives, especially when assessed by RNA integrity number (RIN), it was still "fit" for genomic methods that use smaller sequence lengths, e.g., quantitative PCR. While more than half of the protein derivatives were fit for proteomic purposes, our analyses indicated a significant interaction effect on the absorbance values for proteins derived from FFPE, implying that storage duration may affect protein derivatives differently by tumor tissue type. The mean absorbance value for proteins derived from more recently stored FFPE was greater than protein derived from older FFPE, with the exception of adenocarcinoma tissue. Finally, the fitness of one type of derivative was weakly associated with the fitness of derivatives co-extracted from the same FFPE block. The current study used several novel quality assurance approaches and metrics to show that archival FFPE tissue blocks are a valuable resource for contemporary genomic and proteomic platforms.

Impact of gender on the decision to participate in a clinical trial: a cross-sectional study.

BACKGROUND: In order for Informed Consent to be ethical and valid each clinical trial participant must be able to make a voluntary decision to participate, free from pressure or coercion. Nonetheless, many factors may influence the decision reached, and such influences may be different for male and female volunteers. Being aware of these differences may help researches develop better processes for obtaining consent that safeguard the right of autonomy for all participants. The goal of this study was to evaluate potential gender-based differences in the factors influencing clinical trial participation. METHODS: This cross-sectional study was conducted in the Northeast region of Minas Gerais, Brazil, in October 2011. A structured questionnaire was administered to 143 volunteers (48 male, 95 female) screened for participation in a clinical study of an investigational functional food with potential anthelminthic properties. Answers regarding their decision to participate in the study were compared, by gender, using chi-square and Mann Whitney tests. Odds ratios (OR) was used to measure association. RESULTS: A majority of subjects (58% of males, 59% of females) listed the desire to collaborate with the development of a product against parasitic worms as their main reason for participation. Females were significantly more likely to report a decision influenced by friends, family, or researchers (OR 3.14, 3.45, and 3.46 respectively, p < 0.005). Females were also significantly more likely to report a decision influenced by general altruistic considerations (OR 8.45, p < 0.005). There was no difference, by gender, in the report of decisions influenced by informational meetings, understanding of the disease, or the availability of medical treatments or exams. There was also no difference in knowledge of the rights of research participants. CONCLUSION: Study results indicate that there is a strong difference between male and female participants regarding social influences on the decision to participate in clinical research. Further research into the impact this may have on autonomy is warranted.

Development of cognitive abilities of children infected with helminths through health education.

INTRODUCTION: The aim of this study was to evaluate the effect of health education in learning and cognitive development of children infected, previously treated in an endemic area for helminthiasis. METHODS: It is a longitudinal, experimental, with random allocation of participants. The study included 87 children of both sexes enrolled in the school hall of Maranhão, State of Minas Gerais, Brazil, and divided into two groups: intervention and control. Initially the children were submitted to the parasitological fecal examination for infection diagnosis and, when positive, they were treated. For the data collection, a structured questionnaire and the psychological tests Raven, Wisc-III and DAP III were applied, before and after the educational intervention. For the group comparison, the Mann Whitney test was used, and established significance level of 5%. RESULTS: It was found that previously infected children who received the educational intervention, children showed higher performance than the control group in strutured questionnaire (p<0.05). CONCLUSIONS: It is acceptable to suppose the positive influence and the importance in the use of educational interventions in the cognitive recovery and learning of children previously treated with anthelmintics.

Development of cognitive abilities of children infected with helminths through health education / Desenvolvimento de habilidades cognitivas de crianças infectadas por helmintos através da Educação em Saúde

INTRODUCTION: The aim of this study was to evaluate the effect of health education in learning and cognitive development of children infected, previously treated in an endemic area for helminthiasis. METHODS: It is a longitudinal, experimental, with random allocation of participants. The study included 87 children of both sexes enrolled in the school hall of Maranhão, State of Minas Gerais, Brazil, and divided into two groups: intervention and control. Initially the children were submitted to the parasitological fecal examination for infection diagnosis and, when positive, they were treated. For the data collection, a structured questionnaire and the psychological tests Raven, Wisc-III and DAP III were applied, before and after the educational intervention. For the group comparison, the Mann Whitney test was used, and established significance level of 5%. RESULTS: It was found that previously infected children who received the educational intervention, children showed higher performance than the control group in strutured questionnaire (p<0.05). CONCLUSIONS: It is acceptable to suppose the positive influence and the importance in the use of educational interventions in the cognitive recovery and learning of children previously treated with anthelmintics.

INTRODUÇÃO: O objetivo do estudo foi avaliar o efeito da educação em saúde na aprendizagem e desenvolvimento cognitivo de crianças infectadas, previamente tratadas, em área endêmica para helmintoses. MÉTODOS: Trata-se de um estudo longitudinal, experimental e com distribuição aleatória dos participantes. Participaram do estudo 87 crianças, de ambos os sexos, matriculadas na escola municipal de Maranhão, Minas Gerais, Brasil, e distribuídas em dois grupos: intervenção e controle. Inicialmente, as crianças foram submetidas ao exame parasitológico de fezes para diagnóstico de infecção e, em caso positivo, foram tratadas. Para a coleta de dados, foi aplicado um questionário estruturado e os testes psicológicos Raven, Wisc-III e DFH III, antes e após intervenção educativa. Para a comparação entre os grupos foi utilizado o teste Mann Whitney e estabelecido nível de significância de 5%. RESULTADOS: Verificou-se que as crianças previamente infectadas, que receberam a intervenção educativa, apresentaram desempenho superior as crianças do grupo controle no questionário estruturado (p<0,05). CONCLUSÕES: É aceitável supor a influencia positiva e a importância da utilização de intervenções educativas na recuperação cognitiva e aprendizagem das crianças previamente tratadas com antihelmínticos.

Necator americanus and helminth co-infections: further down-modulation of hookworm-specific type 1 immune responses.

BACKGROUND: Helminth co-infection in humans is common in tropical regions of the world where transmission of soil-transmitted helminths such as Ascaris lumbricoides, Trichuris trichiura, and the hookworms Necator americanus and Ancylostoma duodenale as well as other helminths such as Schistosoma mansoni often occur simultaneously. METHODOLOGY: We investigated whether co-infection with another helminth(s) altered the human immune response to crude antigen extracts from either different stages of N. americanus infection (infective third stage or adult) or different crude antigen extract preparations (adult somatic and adult excretory/secretory). Using these antigens, we compared the cellular and humoral immune responses of individuals mono-infected with hookworm (N. americanus) and individuals co-infected with hookworm and other helminth infections, namely co-infection with either A. lumbricoides, Schistosoma mansoni, or both. Immunological variables were compared between hookworm infection group (mono- versus co-infected) by bootstrap, and principal component analysis (PCA) was used as a data reduction method. CONCLUSIONS: Contrary to several animal studies of helminth co-infection, we found that co-infected individuals had a further downmodulated Th1 cytokine response (e.g., reduced INF-γ), accompanied by a significant increase in the hookworm-specific humoral immune response (e.g. higher levels of IgE or IgG4 to crude antigen extracts) compared with mono- infected individuals. Neither of these changes was associated with a reduction of hookworm infection intensity in helminth co-infected individuals. From the standpoint of hookworm vaccine development, these results are relevant; i.e., the specific immune response to hookworm vaccine antigens might be altered by infection with another helminth.

Excretory-secretory products from hookworm l(3) and adult worms suppress proinflammatory cytokines in infected individuals.

We compared the effects of larval and adult worm excretory-secretory (ES) products from hookworm on the proliferative responses and cytokine secretion in peripheral blood mononuclear cells (PBMCs) from hookwormpatients and egg-negative, nonendemic controls. When compared with negative controls, mitogen-stimulated PBMC from hookworm-infected individuals showed a significantly reduced proliferative response when adult worm ES antigen was added to the cultures. Furthermore, in hookworm-infected individuals a significant downmodulation of inflammatory interleukin (IL)-6 and tumor necrosis factor (TNF)-α secretion resulted when PBMCs were stimulated with mitogen in combination with larval or adult worm ES. Both, interferon (IFN)-γ and IL-10 secretion were significantly lower in stimulated PBMC from infected individuals; however the IFN-γ/IL-10 ratio was much lower in hookworm-infected patients. Comparable effects, although at lower concentrations, were achieved when PBMCs from both groups were incubated with living hookworm third-stage larvae. We suggest that hookworm ES products downmodulate proliferative responses and inflammation during the chronic phase of the disease and facilitate early larval survival or adult worm persistence in the gut.

Hookworm, Ascaris lumbricoides infection and polyparasitism associated with poor cognitive performance in Brazilian schoolchildren.

OBJECTIVE: To investigate the relationship between hookworm and Ascaris lumbricoides infection and performance on three subsets of the Wechsler Intelligence Scale for Children - third edition (WISC-III) (Digit Span, Arithmetic and Coding) and Raven Colored Progressive Matrices. METHODS: Cross-sectional study of 210 children between the ages of 6 and 11 years in Americaninhas, Minas Gerais, Brazil. Separate proportional odds models were used to measure the association between the intensity of helminth infections and poor performance on each of the four cognitive tests. RESULTS: After adjusting for sex, age, socioeconomic status and other helminth infections, moderate-to-high-intensity hookworm infection was associated with poor performance on the WISC-III Coding subtest [OR = 3.20; 95% confidence interval (CI) = 1.43-7.17], low intensity of hookworm infection was associated with poor performance on the WISC-III Coding subtest [odds ratio (OR) = 3.71; 95% CI = 1.80-7.66] and moderate-to-high-intensity A. lumbricoides infection was associated with poor performance on the Raven test (OR = 2.03; 95% CI = 1.04-3.99), all in comparison with uninfected children. Children co-infected with A. lumbricoides infection and hookworm infection had greater odds of poor performance on some WISC-III subtests than children with only A. lumbricoides infection. CONCLUSIONS: These findings suggest that hookworm infection may be associated with poorer concentration and information processing skills, as measured on the WISC-III Coding subtest, and that A. lumbricoides infection may be associated with poorer general intelligence, as measured through the Raven Colored Progressive Matrices. This study also presents evidence that polyparasitized children experience worse cognitive outcomes than children with only one helminth infection.

Age patterns in undernutrition and helminth infection in a rural area of Brazil: associations with ascariasis and hookworm.

OBJECTIVE: To investigate the nutritional status of individuals from a rural area of Brazil, and associations with helminth infections in an age-stratified sample. METHOD: A total of 1113 individuals aged from 6 months to 83 years from the rural community of Americaninhas in Minas Gerais were investigated. Assessments comprised anthropometric measurements of weight, height and body composition, examining faecal samples for helminth eggs, and peripheral blood assays for albumin, haemoglobin and ferritin concentrations. RESULTS: Ten percent of the participants were underweight, 12.8% were overweight and 28.3% of the children and adolescents were stunted. 11.6% had low lean body mass and 28.8% had low fat body mass. Hypoalbuminaemia was seen in 5.5%, anaemia in 12.5% and iron deficiency in 13.1%, although the prevalence of these two indices increased with age. Multivariate analysis showed that, after controlling for age, sex and socio-economic status, stunting was significantly associated with Ascaris lumbricoides infection among children and adolescents, whereas low body mass was significantly associated with hookworm infection among adults and the elderly. CONCLUSIONS: Helminth infections are associated with undernutrition in endemic populations, with important differences between the effects of hookworm and A. lumbricoides on age-related nutritional status.

Immunogenicity in dogs of three recombinant antigens (TSA, LeIF and LmSTI1) potential vaccine candidates for canine visceral leishmaniasis.

Control of canine visceral leishmaniasis (VL) remains a difficult and serious problem mostly because there is no reliable and effective vaccine available to prevent this disease. A mixture of three recombinant leishmanial antigens (TSA, LeIF and LmSTI1) encoded by three genes highly conserved in the Leishmania genus have been shown to induce excellent protection against infection in both murine and simian models of cutaneous leishmaniasis. A human clinical trial with these antigens is currently underway. Because of the high degree of conservation, these antigens might be useful vaccine candidates for VL as well. In the present study, using the dog model of the visceral disease, we evaluated the immunogenicity of these three antigens formulated with two different adjuvants, MPL-SE and AdjuPrime. The results were compared with a whole parasite vaccine formulated with BCG as the adjuvant. In order to investigate if sensitization with the recombinant antigens would result in recognition of the corresponding native parasite antigens upon infection, the animals were exposed for four weeks after the termination of the immunization protocol with the recombinant antigens to a low number of L. chagasi promastigotes, an etiological agent of VL. Immune response was evaluated by quantitative ELISA in the animal sera before and after exposure to the viable parasites. Both antigen specific IgG1 and IgG2 antibody levels were measured. Immunization of dogs with the recombinant antigens formulated in either MPL-SE or AdjuPrime resulted in high antibody levels particularly to LmSTI1. In addition, this immunization although to low levels, resulted in the development of antibody response to the whole parasite lysate. Importantly, experimental exposure with low numbers of culture forms of L. chagasi promastigotes caused a clear boost in the immune response to both the recombinant antigens and the corresponding native molecules. The boost response was predominantly of the IgG2 isotype in animals primed with the recombinant antigens plus MPL-SE. In contrast, animals primed with the recombinant antigens formulated in AdjuPrime as well as animals vaccinated with crude antigen preparation responded with mixed IgG1/IgG2 isotypes. These results point to the possible use of this antigen cocktail formulated with the adjuvant MPL-SE in efficacy field trials against canine VL.