Dynamics of the double burden of malnutrition in Guatemala: a secondary data analysis of the demographic and health surveys from 1998-2015.

OBJECTIVE: We estimated the prevalence and time trends of the double burden of malnutrition (DBM) in Guatemala and explored its occurrence based on socio-demographic factors. STUDY DESIGN: This was a secondary data analysis using information from four Demographic and Health Surveys covering the period 1998-2015. METHODS: The unit of analysis was the household within which information was gathered from women 18-49 years and their children, 6-59 months. The main outcome was the prevalence of any DBM in the household (co-existence of undernutrition and overnutrition in a woman, her children or both). We estimated the prevalence of any DBM by survey and analysed time trends. Stepwise logistic regression was used to explore the occurrence of DBM and socio-demographic factors. RESULTS: We analysed 39,749 households across all surveys. The prevalence of any DBM was 25.3% (95%CI: 22.1-28.7) in 1998-99, 23.8% (22.0-25.8) in 2002, 25.9% (24.3-27.5) in 2008-09 and 24.2% (22.9-25.5) in 2014-15, with no significant change over time (P = 0.782). Characteristics associated with lower odds of any DBM were rural residence, female-headed household, wealth and women's secondary education. Higher odds were seen for households with electricity, women >25y, indigenous and with >2 children. CONCLUSION: Our findings revealed that a quarter of Guatemala's households suffer from DBM, which has remained unchanged for 17 years. Interventions should prioritise urban areas, households of lower socio-economic status and those less educated. To increase awareness of policymakers of this pressing public health concern, further research on DBM could be strengthened by prospective study designs, integrating all household members and expanding the types of malnutrition.

Micronutrient status of populations and preventive nutrition interventions in South East Asia.

Objectives Since the 1990s, programs for the control of micronutrient deficiencies became a public health priority for many governments, including the countries partnering the project "Sustainable Micronutrient Interventions to Control Deficiencies and Improve Nutritional Status and General Health in Asia" (SMILING): Cambodia, Indonesia, Laos-PDR, Thailand and Vietnam. The aim of this study was to map which micronutrient deficiencies have been addressed and which interventions were in place in the SMILING countries. Methods The mapping covered the period up to 2012. Updated information from relevant surveys after 2012 is included in this paper after the completion of the SMILING project. The mapping of micronutrient status was limited to either national or at least large-scale surveys. Information on nutrition interventions obtained through a systematic mapping of national programs combined with a snowball collection from various sources. Results Among the five SMILING countries, Thailand differed historically by an early implementation of a nationwide community-based nutrition program, contributing to reductions in undernutrition and micronutrient deficiencies. For Cambodia, Indonesia, Laos PDR, and Vietnam, some national programs addressing micronutrients have been implemented following adjusted international recommendations. National surveys on micronutrient status were scattered and inconsistent across the countries in design and frequency. Conclusion for practice In conclusion, some micronutrient deficiencies were addressed in national interventions but the evidence of effects was generally lacking because of limited nationally representative data collected. Improvement of intervention programs to efficiently reduce or eliminate micronutrient deficiencies requires more systematic monitoring and evaluation of effects of interventions in order to identify best practices.

What Approaches are Most Effective at Addressing Micronutrient Deficiency in Children 0-5 Years? A Review of Systematic Reviews.

Introduction Even though micronutrient deficiency is still a major public health problem, it is still unclear which interventions are most effective in improving micronutrient status. This review therefore aims to summarize the evidence published in systematic reviews on intervention strategies that aim at improving micronutrient status in children under the age of five. Methods We searched the literature and included systematic reviews that reported on micronutrient status as a primary outcome for children of 0-5 years old, had a focus on low or middle income countries. Subsequently, papers were reviewed and selected by two authors. Results We included 4235 reviews in this systematic review. We found that (single or multiple) micronutrient deficiencies in pre-school children improved after providing (single or multiple) micronutrients. However home fortification did not always lead to significant increase in serum vitamin A, serum ferritin, hemoglobin or zinc. Commercial fortification did improve iron status. Cord clamping reduced the risk of anemia in infants up to 6 months and, in helminth endemic areas, anthelminthic treatment increased serum ferritin levels, hemoglobin and improved height for age z-scores. Anti-malaria treatment improved ferritin levels. Discussion Based on our results the clearest recommendations are: delayed cord clamping is an effective intervention for reducing anemia in early life. In helminth endemic areas iron status can be improved by anthelminthic treatment. Anti-malaria treatment can improve ferritin. In deficient populations, single iron, vitamin A and multimicronutrient supplementation can improve iron, vitamin A and multimicronutrient status respectively. While the impact of home-fortification on multimicronutrient status remains questionable, commercial iron fortification may improve iron status.

Intestinal parasites: Associations with intestinal and systemic inflammation.

The aim of the present study was to evaluate associations between intestinal parasitic infection with intestinal and systemic inflammatory markers in school-aged children with high rates of obesity. Plasma concentrations of C-Reactive Protein (CRP), leptin, TNF-α, IL-6 and IL-10 were measured as systemic inflammation markers and count of stool leukocytes as marker of intestinal inflammation in 291 children (6-10 years). Intestinal parasitic infection was measured by stool examination. Logistic regression analyses were performed to determine the odds of having high inflammatory markers for each parasite or group of parasites as compared to parasite-free children while adjusting for sex, age, mother's educational level and percentage of body fat. The prevalence of soil-transmitted helminths and intestinal protozoa infections was 12% and 36%, respectively. Parasitic infection was not associated with CRP, IL-6, IL-10 or TNF-α. Children infected with Ascaris lumbricoides (aOR: 5.91, 95% CI: 1.97-17.70) and Entamoeba coli (aOR: 8.46, 95% CI: 2.85-25.14) were more likely to have higher stool leucocytes than parasite-free children. Children with multiple infections (aOR: 10.60, 95% CI: 2.85-25.14) were more likely to have higher leptin concentrations than parasite-free children. Intestinal parasitic infection was not associated with systemic inflammation, but was associated with intestinal inflammation. Having multiple infections were associated with higher leptin concentrations.

Combined stool-based multiplex PCR and microscopy for enhanced pathogen detection in patients with persistent diarrhoea and asymptomatic controls from Côte d'Ivoire.

Infectious diarrhoea ranks among the leading causes of morbidity worldwide. Although most acute diarrhoeal episodes are self-limiting, the diagnosis and treatment of persistent diarrhoea (≥2 weeks) are cumbersome and require laboratory identification of the causative pathogen. Stool-based PCR assays have greatly improved the previously disappointing pathogen detection rates in high-income countries, but there is a paucity of quality data from tropical settings. We performed a case-control study to elucidate the spectrum of intestinal pathogens in patients with persistent diarrhoea and asymptomatic controls in southern Côte d'Ivoire. Stool samples from 68 patients and 68 controls were obtained and subjected to molecular multiplex testing with the Luminex(®) Gastrointestinal Pathogen Panel (GPP), microscopy and rapid antigen detection tests for the diagnosis of diarrhoeagenic pathogens. Overall, 20 different bacteria, parasites and viruses were detected by the suite of diagnostic methods employed. At least one pathogen was observed in 84% of the participants, and co-infections were observed in >50% of the participants. Enterotoxigenic Escherichia coli (32%), Giardia intestinalis (29%) and Shigella species (20%) were the predominant pathogens, and Strongyloides stercoralis (10%) was the most prevalent helminth. Pathogen frequencies and numbers of co-infections were similar in patients and controls. Although the Luminex(®) GPP detects a broad range of pathogens, microscopy for helminths and intestinal protozoa remains necessary to cover the full aetiological spectrum in tropical settings. We conclude that highly sensitive multiplex PCR assays constitute a useful screening tool, but that positive results might need to be confirmed by independent methods to discriminate active infection from asymptomatic faecal shedding of nucleic acids.

[Prevalence of geohelminth infections in children living in Kinshasa]. / Prévalence des géohelminthiases chez les enfants à Kinshasa.

BACKGROUND: Data on geohelminth infections in children in the Democratic Republic of Congo are sparse. The objective of this study was to document and compare the profile and prevalence in children living in Kinshasa. METHODS: A prospective cohort study was conducted from May to October 2009 in children in the Biyela health area in Kinshasa, Democratic Republic of Congo. Stool samples were collected from representative members of these two populations and analyzed for geohelminths (GH) using the Kato-Katz technique. RESULTS: In this series, 438 school-age children were included. There were 235 children recruited in schools and 203 in households (77.8%). Overall prevalence of geohelminths was 66.9%. The specific prevalence was 69.4% in children recruited in schools and 64.0% in children recruited in households. The frequency of Ascaris lumbricoides, Trichuris trichiura, and Ancylostoma species were, respectively, 56.2%, 38.7%, and 1.7% in schools and 39.9%, 51.7%, and 1.0% in households. A. lumbricoides was significantly more prevalent in schools (56.2% vs 39.9%; OR=2.0; 95%CI: 1.3-3.0), T. trichiura was significantly less prevalent in schools (38.7% vs 51.7%; OR=0.6; 95% CI: 0.4-0.9). There were no significant differences in the prevalence of Ancylostoma between schools and households. CONCLUSION: GH is a health problem among Biyela children. Preventive measures and education of the population need to be emphasized in attempts to reduce the prevalence of geohelminths in these children.

Inbreeding within human Schistosoma mansoni: do host-specific factors shape the genetic composition of parasite populations?

The size, structure and distribution of host populations are key determinants of the genetic composition of parasite populations. Despite the evolutionary and epidemiological merits, there has been little consideration of how host heterogeneities affect the evolutionary trajectories of parasite populations. We assessed the genetic composition of natural populations of the parasite Schistosoma mansoni in northern Senegal. A total of 1346 parasites were collected from 14 snail and 57 human hosts within three villages and individually genotyped using nine microsatellite markers. Human host demographic parameters (age, gender and village of residence) and co-infection with Schistosoma haematobium were documented, and S. mansoni infection intensities were quantified. F-statistics and clustering analyses revealed a random distribution (panmixia) of parasite genetic variation among villages and hosts, confirming the concept of human hosts as 'genetic mixing bowls' for schistosomes. Host gender and village of residence did not show any association with parasite genetics. Host age, however, was significantly correlated with parasite inbreeding and heterozygosity, with children being more infected by related parasites than adults. The patterns may be explained by (1) genotype-dependent 'concomitant immunity' that leads to selective recruitment of genetically unrelated worms with host age, and/or (2) the 'genetic mixing bowl' hypothesis, where older hosts have been exposed to a wider variety of parasite strains than children. The present study suggests that host-specific factors may shape the genetic composition of schistosome populations, revealing important insights into host-parasite interactions within a natural system.

Prediction of Asthma by Common Risk Factors: A Follow-up Study in Cuban Schoolchildren

Objective: To determine which common risk factors, including environmental factors, are predictors for the development of asthma in Cuban schoolchildren. Methods: A longitudinal study was conducted in 1042 schoolchildren without asthma at baseline in 2 Cuban municipalities. Asthma status in 2007, diagnosed using the International Study of Asthma and Allergies in Childhood questionnaire, was related to a set of common risk factors assessed in 2003/2004 in a multivariable logistic regression model. Multiple imputation was used for missing values. The final prediction model was obtained by backward selection (P<.15). The model’s prognostic accuracy (R2) and discriminative ability (area under the receiver operating characteristic curve [AUC]) were assessed and internal validation by bootstrapping was performed. Results: A family history of atopic diseases (odds ratio [OR], 2.19; 95% CI, 1.19-4.04), allergic sensitization (OR, 1.83; 95% CI, 0.94-3.55), municipality (OR, 0.34; 95% CI, 0.15-0.74), and use of antibiotics in the child’s first year of life (OR, 1.66; 95% CI, 0.89-3.11) were predictors for asthma development. The model had an R2 of 8.0% and a moderate discriminative ability (AUC, 0.69; 95% CI, 0.60-0.78). Internal validation hardly influenced the model’s performance. Conclusions: Antibiotics use, genetic predisposition, and allergic sensitization were predictors of asthma in Cuban schoolchildren. Although known as common risk factors they could only partly predict asthma development. Poverty-related factors, such as low income and education, and parasitic infections, did not have an effect. Other or additional environmental predictors need to be identified, as these are potential targets for prevention and control of childhood asthma in affluent as well as nonaffluent countries (AU)

Objetivo: Determinar cuáles de los factores de riesgo habituales, incluyendo factores ambientales, tienen capacidad predictiva para el desarrollo de asma en población escolar cubana. Métodos: Estudio longitudinal realizado en una muestra de 1042 escolares cubanos, sin asma al comienzo del estudio, residentes en dos municipios. La presencia de asma en el año 2007, diagnosticada mediante el cuestionario ISAAC, se relacionó con la presencia de factores de riesgo habituales, presentes al inicio del estudio en los años 2003-2004 mediante un modelo de regresión logística multivariable. Se utilizó el criterio de imputación múltiple para los valores no registrados. El modelo final predictivo se obtuvo mediante selección retrógrada. Se evaluaron la capacidad predictiva del modelo (R cuadrada) y capacidad discriminante (Área bajo la curva ROC) y se realizó una validación interna mediante la técnica de muestreo “bootstrapping”. Resultados: Una historia familiar de enfermedades alérgicas (OR 2.19, 95%CI 1.19-4.04), la presencia de sensibilización frente a alérgenos comunes (OR 1.83, 95%CI 0.94-3.55), el municipio de residencia (OR 0.34, 95%CI 0.15-0.74), y el uso de antibióticos en el primer año del vida del niño (OR 1.66, 95%CI 0.89-3.11) fueron todos ellos factores predictivos del desarrollo de asma. La R cuadrada del modelo fue el 8,0% y poseía una moderada capacidad discriminativa (AUC 0.69, 95%CI 0.60-0.78). La validez interna influencia notablemente el funcionamiento del modelo. Conclusiones: Los antecedentes familiares de enfermedad alérgica, la sensibilización frente a alérgenos comunes y el uso de antibióticos en el primer año de vida fueron factores predictivos de la aparición de asma en los escolares cubanos, si bien solo fueron capaces de explicar en parte esta predisposición. Otros factores relacionados con un nivel socioeconómico bajo, como un nivel de ingresos o de educación bajos o las infecciones por parásitos, no tuvieron ningún efecto. Se hace por tanto necesario, identificar otros o adicionales factores predictivos ambientales, que sean también dianas de programas de prevención y control de la enfermedad asmática tanto en países de niveles económicos altos o bajos (AU)

Hybridisation between the two major African schistosome species of humans.

It is generally accepted that Schistosoma mansoni and Schistosoma haematobium, causing intestinal and urinary schistosomiasis, respectively, are not able to hybridise, due to the high phylogenetic distance between them. Cloning of nuclear internal transcribed spacer rDNA and partial mitochondrial cytochrome c oxidase 1 fragments revealed two internal transcribed spacer rDNA genotypes within single eggs and miracidia, one identical to S. mansoni and the other identical to S. haematobium, suggesting hybrid ancestry. The cytochrome c oxidase 1 clones always belonged to only one of the parental species. This demonstrates that offspring of heterologous pairing between these two species is not (always) parthenogenetic.

Deworming is not a risk factor for the development of atopic diseases: a longitudinal study in Cuban school children.

BACKGROUND: Soil-transmitted helminth (STH) infections have been suggested to protect from allergic sensitization and atopic diseases. Consequently, anthelminthic treatment would increase the prevalence of atopic disease in STH endemic populations. OBJECTIVE: To investigate the effect of deworming on allergic sensitization and atopic diseases in Cuban schoolchildren. METHODS: We followed up 108 STH positive schoolchildren aged 5-13 in six-monthly intervals for 24 months. Four consecutive groups of, respectively, 104, 56, 68, and 53 STH positive children were used as 'untreated' reference groups to assess general time trends. STH infections were diagnosed by stool examination. Asthma, allergic rhinoconjunctivitis, and atopic dermatitis were diagnosed by International Study of Asthma and Allergies in Childhood (ISAAC) questionnaire and allergic sensitization by skin prick testing (SPT). At each time point, STH positive children were treated with one single dose of 500 mg mebendazole. RESULTS: After deworming, the frequency of asthma significantly decreased (P < 0.001) while the frequency of allergic rhinoconjunctivitis and atopic dermatitis was not affected (P = 0.129 and P = 0.751, respectively). The percentage of SPT positives temporarily increased (P < 0.001) and subsequently returned to nearly baseline values (P = 0.093). In the references groups, no change over time was observed in the proportion of children with allergic sensitization and atopic diseases (P > 0.05). CONCLUSION & CLINICAL RELEVANCE: Our results indicate that atopic diseases do not increase after anthelminthic treatment. Allergic sensitization on the other hand increases after deworming. As this increase appears only temporarily, deworming of schoolchildren does not seem to be a risk factor for the development of allergic sensitization, nor for atopic diseases.

Regular treatments of praziquantel do not impact on the genetic make-up of Schistosoma mansoni in Northern Senegal.

The Senegal River Basin (SRB) experienced a major epidemic of intestinal schistosomiasis in the early nineties, after the construction of a dam for irrigation purposes. Exceptionally low cure rates following praziquantel (PZQ) treatment at the onset of the epidemic raised concerns about PZQ resistant strains of Schistosoma mansoni, although they could also be attributed to the intense transmission at that time. A field study in the same region more than 15 years later found cure rates for S. mansoni still to be low, whereas Schistosomahaematobium responded well to treatment. We collected S. mansoni miracidia from children at base-line prior to treatment, six months after two PZQ treatments and two years after the start of the study when they had received a total of five PZQ treatments. In total, 434 miracidia from 12 children were successfully genotyped with at least six out of nine DNA microsatellite loci. We found no significant differences in the genetic diversity of, and genetic differentiation between parasite populations before and after repeated treatment, suggesting that PZQ treatment does not have an impact on the neutral evolution of the parasite. This is in stark contrast with a similar study in Tanzania where a significant decrease in genetic diversity was observed in S. mansoni miracidia after a single round of PZQ treatment. We argue that PZQ resistance might play a role in our study area, although rapid re-infection cannot be excluded. It is important to monitor this situation carefully and conduct larger field studies with short-term follow-up after treatment. Since PZQ is the only general schistosomicide available, the possibility of PZQ resistance is of great concern both for disease control and for curative use in clinical practice.

An agent-based model of exposure to human toxocariasis: a multi-country validation.

Seroprevalence data illustrate that human exposure to Toxocara is frequent. Environmental contamination with Toxocara spp. eggs is assumed to be the best indicator of human exposure, but increased risk of exposure has also been associated with many other factors. Reported associations are inconsistent, however, and there is still ambiguity regarding the factors driving the onset of Toxocara antibody positivity. The objective of this work was to assess the validity of our current conceptual understanding of the key processes driving human exposure to Toxocara. We constructed an agent-based model predicting Toxocara antibody positivity (as a measure of exposure) in children. Exposure was assumed to depend on the joint probability of 3 parameters: (1) environmental contamination with Toxocara spp. eggs, (2) larvation of these eggs and (3) the age-related contact with these eggs. This joint probability was linked to processes of acquired humoral immunity, influencing the rate of antibody seroreversion. The results of the simulation were validated against published data from 5 different geographical settings. Using simple rules and a stochastic approach with parameter estimates derived from the respective contexts, plausible serological patterns emerged from the model in nearly all settings. Our approach leads to novel insights in the transmission dynamics of Toxocara.

Toxocara seropositivity, atopy and asthma: a study in Cuban schoolchildren.

INTRODUCTION: Evidence suggests that human toxocariasis (HT) could stimulate the onset of allergic diseases such as asthma. More specifically, in subjects having a hypothetical 'atopic genotype', HT could boost preexistent allergy symptoms. We tested the latter hypothesis in Cuba, a country where both asthma and HT are prevalent. MATERIAL AND METHODS: In a group of Cuban school-aged children (n = 958), we investigated the association of Toxocara seropositivity and atopic status with asthma. Toxocara seropositivity was diagnosed with ELISA and atopy by allergen skin prick test. Both physician-diagnosed asthma and current wheeze, as determined by International Study of Asthma and Allergies in Childhood (ISAAC) questionnaire, were considered. Associations were assessed using multivariable logistic regression analyses, with either 'physician-diagnosed asthma' or 'current wheeze' as outcome variable. RESULTS: 40.1% of the children were Toxocara seropositive. Prevalences were 21.7% for current wheeze and 32.7% for physician-diagnosed asthma. The odds of having asthma were almost two times higher in atopic children, but only reached borderline significance (OR=1.90, CI 95%: 0.95-3.80 for physician-diagnosed asthma and OR=1.94, CI 95%: 0.98-3.85 for current wheeze). Toxocara seropositivity and physician-diagnosed asthma were associated (OR=1.51, CI 95%: 1.01-2.26). Moreover, in children without antibodies to Toxocara, being atopic was significantly associated with having physician-diagnosed asthma (OR=2.53, CI 95%: 1.63-3.90), while this association was not present in Toxocara positives (OR=1.38, CI 95%: 0.82-2.37). CONCLUSION: Our data confirm previous observations of higher Toxocara seropositivity rates in asthmatic children. Toxocara seropositivity appeared to abrogate the apparent association between atopy and asthma in Cuban children. Although this observation was limited to physician-diagnosed asthma, it challenges the hypothesis that HT stimulates the onset of allergic diseases such as asthma in atopic individuals.

Schistosome infection is associated with enhanced whole-blood IL-10 secretion in response to cercarial excretory/secretory products.

Infection of the human host by schistosome parasites follows exposure of skin to free-swimming cercariae and is aided by the release of excretory/secretory (E/S) material, which is rich in proteases and glycoconjugates. This material provides the initial stimulus to cells of the innate immune system. The study presented here is the first to examine human innate/early immune responsiveness to cercarial E/S in subjects from an area co-endemic for Schistosoma mansoni and S. haematobium. We report that in infected participants, stimulation of whole-blood cultures with cercarial E/S material (termed 0-3 hRP) caused the early (within 24 h) release of greater quantities of regulatory IL-10, compared with uninfected controls. Elevated levels of IL-10 but not pro-inflammatory TNFα or IL-8 were most evident in participants co-infected with S. mansoni and S. haematobium and were accompanied by a higher 0-3 h RP-specific IL-10: TNFα ratio. We also report that glycosylated components within 0-3 h RP appear to be important factors in the stimulation of IL-8, TNFα and IL-10 production by whole-blood cells.

Prediction of asthma by common risk factors: a follow-up study in Cuban schoolchildren.

OBJECTIVE: To determine which common risk factors, including environmental factors, are predictors for the development of asthma in Cuban schoolchildren. METHODS: A longitudinal study was conducted in 1042 schoolchildren without asthma at baseline in 2 Cuban municipalities. Asthma status in 2007, diagnosed using the International Study of Asthma and Allergies in Childhood questionnaire, was related to a set of common risk factors assessed in 2003/2004 in a multivariable logistic regression model. Multiple imputation was used for missing values. The final prediction model was obtained by backward selection (P<.15). The model's prognostic accuracy (R2) and discriminative ability (area under the receiver operating characteristic curve [AUC]) were assessed and internal validation by bootstrapping was performed. RESULTS: A family history of atopic diseases (odds ratio [OR], 2.19; 95% CI, 1.19-4.04), allergic sensitization (OR, 1.83; 95% CI, 0.94-3.55), municipality (OR, 0.34; 95% CI, 0.15-0.74), and use of antibiotics in the child's first year of life (OR, 1.66; 95% CI, 0.89-3.11) were predictors for asthma development. The model had an R2 of 8.0% and a moderate discriminative ability (AUC, 0.69; 95% CI, 0.60-0.78). Internal validation hardly influenced the model's performance. CONCLUSIONS: Antibiotics use, genetic predisposition, and allergic sensitization were predictors of asthma in Cuban schoolchildren. Although known as common risk factors they could only partly predict asthma development. Poverty-related factors, such as low income and education, and parasitic infections, did not have an effect. Other or additional environmental predictors need to be identified, as these are potential targets for prevention and control of childhood asthma in affluent as well as nonaffluent countries.

Frequency of antibodies to Toxocara in Cuban schoolchildren.

OBJECTIVE: The aim of the study was to determine the frequency of antibodies to Toxocara in Cuban schoolchildren. METHODS: The frequency of antibodies to Toxocara canis was assessed with a commercial enzyme-linked immunosorbent assays kit in school-aged children from two municipalities of Cuba. Univariate analysis and a multivariable logistic regression analysis adjusted for age, sex, municipality and co-infection with helminth and/or protozoa were conducted. RESULTS: The percentage of children with antibodies to Toxocara was 38.8% (392/1011; 95% CI = 36.8-42.8). Antibody positivity was significantly associated with gender and co-infections with intestinal parasites, but not with age or municipality. CONCLUSION: Cuban children are highly exposed to the Toxocara parasite, corresponding well with reported environmental contamination with Toxocara parasite eggs and T. canis prevalences in dogs in Cuba. Relevant policy makers and the Cuban population need to be better informed about this preventable infection.

Optimal sample storage and extraction procotols for reliable multilocus genotyping of the human parasite Schistosoma mansoni.

Genotyping individual larval stages and eggs of natural parasite populations is complicated by the difficulty of obtaining reliable genotypes from low quantity DNA template. A suitable storage and extraction protocol, together with a thorough quantification of genotyping errors are therefore crucial for molecular epidemiological studies. Here we test the robustness, handling time, ease of use, cost effectiveness and success rate of various fixation (Whatman FTA(®) Classic and Elute Cards, 70% EtOH and RNAlater(®)) and subsequent DNA extraction methods (commercial kits and proteinase K protocol). None of these methods require a cooling chain and are therefore suitable for field collection. Based on a multiplex microsatellite PCR with nine loci the success and reliability of each technique is evaluated by the proportion of samples with at least eight scored loci and the proportion of genotyping errors. If only the former is taken into account, FTA(®) Elute is recommended (83% success; 44% genotyping error; 0.2 €/sample; 1h 20 m handling time). However, when also considering the genotyping errors, handling time and ease of use, we opt for 70% EtOH with the 96-well plate technology followed by a simple proteinase K extraction (73% success; 0% genotyping error; 0.2 €/sample; 15m handling time). For eggs we suggest (1) to pool all eggs per person in 1.5 ml tubes filled with 70% EtOH for transport and (2) to identify each egg to species level prior to genotyping. To this end we extended the Rapid diagnostic PCR developed by Webster et al. (2010) with a S. mansoni-specific primer to discriminate between S. mansoni, S. haematobium and S. bovis in a single PCR reaction. The success rate of genotyping eggs was 75% (0% genotyping error). This is the first study to incorporate genotyping errors through re-amplification for the evaluation of schistosome sampling protocols and the identification of error-prone loci.

Helminths and malignancy.

It has been estimated that chronic infections with viruses, bacteria and parasites contribute to 17.8% of the global burden of cancer, although only a relatively small proportion of the infection-related cancers can be attributed to helminth infections. These are important because of the high number of people who are exposed or infected worldwide. Carcinogenesis associated with helminth infections is a complex process, which may involve several different mechanisms, but chronic inflammation is a key feature. Host immune responses and immunopathological processes mediate inflammatory responses and any failure in the control of the immunological components involved in this response can lead to chronic inflammation. This may generate a microenvironment that might be conducive to the initiation and development of cancer. Inflammatory cells generate free radicals and nitrogen species, which can oxidize and damage DNA and lead to genetic instabilities and malignant transformation. Physical damage caused by the parasites, their eggs or secreted products leads to restorative hyperplasia of the damaged tissue. This may promote the propagation of cells, in which genotoxic damage and pre-malignant change has taken place. Three helminth infections have been classified as definitely carcinogenic to humans (group 1 carcinogens), namely Schistosoma haematobium, which is associated with cancer of the urinary bladder and the food-borne liver flukes Clonorchis sinensis and Opisthorchis viverrini associated with cholangiocarcinoma of the liver. Reducing the level of infection and the risk of getting (re)infected will reduce the risk of cancer development later in life. Helminth infections are thus a preventable cause of cancer, emphasizing the need for sustainable helminth control in endemic areas coupled with health education, especially in relation to food-borne liver fluke infections.