The NIH-NIAID Filariasis Research Reagent Resource Center

Filarial worms cause a variety of tropical diseases in humans; however, they are difficult to study because they have complex life cycles that require arthropod intermediate hosts and mammalian definitive hosts. Research efforts in industrialized countries are further complicated by the fact that some filarial nematodes that cause disease in humans are restricted in host specificity to humans alone. This potentially makes the commitment to research difficult, expensive, and restrictive. Over 40 years ago, the United States National Institutes of Health–National Institute of Allergy and Infectious Diseases (NIH-NIAID) established a resource from which investigators could obtain various filarial parasite species and life cycle stages without having to expend the effort and funds necessary to maintain the entire life cycles in their own laboratories. This centralized resource (The Filariasis Research Reagent Resource Center, or FR3) translated into cost savings to both NIH-NIAID and to principal investigators by freeing up personnel costs on grants and allowing investigators to divert more funds to targeted research goals. Many investigators, especially those new to the field of tropical medicine, are unaware of the scope of materials and support provided by the FR3. This review is intended to provide a short history of the contract, brief descriptions of the fiilarial species and molecular resources provided, and an estimate of the impact the resource has had on the research community, and describes some new additions and potential benefits the resource center might have for the ever-changing research interests of investigators

Web-Based Virtual Microscopy for Parasitology: A Novel Tool for Education and Quality Assurance

Here, we describe a novel tool to observe parasites by virtual microscopy on the Internet. Microscopy-based identification of parasites is the basis for both diagnostics and epidemiological assessment of parasite burden globally. Yet, quality assessment of diagnostic parasitology laboratories is difficult, as delivering identical educational specimens has been impossible. In this study, a series of parasite specimens on ordinary glass slides were digitized using a recently developed microscope scanner technique. Up to 50,000 images captured at high magnification are digitally stitched together to form a representation of the entire glass slide. These “virtual slides” digitized at a thousand-fold magnification can hold more than 60 gigabytes of data. Handling such large amounts of data was made possible because of efficient compression techniques and a viewing system adopted from the geospatial imaging industry. Viewing the samples on the Internet very much resembles, for example, the use of Google Maps, and puts only modest requirements on the viewer's computer. In addition, we captured image stacks at different focal planes, and developed a web-based viewing system for three-dimensional navigation in the specimens. This novel technique is especially valuable for detailed visualization of large objects such as helminth eggs in stool specimens

The secreted triose phosphate isomerase of Brugia malayi is required to sustain microfilaria production in vivo

Human lymphatic filariasis is a major tropical disease transmitted through mosquito vectors which take up microfilarial larvae from the blood of infected subjects. Microfilariae are produced by long-lived adult parasites, which also release a suite of excretory-secretory products that have recently been subject to in-depth proteomic analysis. Surprisingly, the most abundant secreted protein of adult Brugia malayi is triose phosphate isomerase (TPI), a glycolytic enzyme usually associated with the cytosol. We now show that while TPI is a prominent target of the antibody response to infection, there is little antibody-mediated inhibition of catalytic activity by polyclonal sera. We generated a panel of twenty-three anti-TPI monoclonal antibodies and found only two were able to block TPI enzymatic activity. Immunisation of jirds with B. malayi TPI, or mice with the homologous protein from the rodent filaria Litomosoides sigmodontis, failed to induce neutralising antibodies or protective immunity. In contrast, passive transfer of neutralising monoclonal antibody to mice prior to implantation with adult B. malayi resulted in 60–70% reductions in microfilarial levels in vivo and both oocyte and microfilarial production by individual adult females. The loss of fecundity was accompanied by reduced IFNγ expression by CD4+ T cells and a higher proportion of macrophages at the site of infection. Thus, enzymatically active TPI plays an important role in the transmission cycle of B. malayi filarial parasites and is identified as a potential target for immunological and pharmacological intervention against filarial infections

Long-Term Weekly Iron-Folic Acid and De-Worming Is Associated with Stabilised Haemoglobin and Increasing Iron Stores in Non-Pregnant Women in Vietnam

BACKGROUND: The prevalence of anaemia and iron deficiency in women remains high worldwide. WHO recommends weekly iron-folic acid supplementation where anaemia rates in non-pregnant women of reproductive age are higher than 20%. In 2006, a demonstration project consisting of weekly iron-folic acid supplementation and regular de-worming was set up in two districts in a northern province in Vietnam where anaemia and hookworm rates were 38% and 76% respectively. In 2008 the project was expanded to all districts in the province, targeting some 250,000 women. The objectives of this study were to: 1) examine changes in haemoglobin, iron stores and soil transmitted helminth infection prevalence over three years and 2) assess women's access to and compliance with the intervention. METHODS AND FINDINGS: The study was a semi-cross-sectional, semi-longitudinal panel design with a baseline survey, three impact surveys at three-, twelve- and thirty months after commencement of the intervention, and three compliance surveys after ten weeks, eighteen and thirty six months. RESULTS: After thirty months, mean haemoglobin stabilised at 130.3 g/L, an increase of 8.2 g/L from baseline, and mean serum ferritin rose from 23.9 µg/L to 52 µg/L. Hookworm prevalence fell from 76% to 22% over the same period. After thirty six months, 81% of the target population were receiving supplements and 87% were taking 75% or more of the supplements they received. CONCLUSIONS: Weekly iron-folic acid supplementation and regular de-worming was effective in significantly and sustainably reducing the prevalence of anaemia and soil transmitted helminth infections and high compliance rates were maintained over three years

A community based field research project investigating anaemia amongst young children living in rural Karnataka, India: a cross sectional study

<p>Abstract</p> <p>Background</p> <p>Anaemia is an important problem amongst young children living in rural India. However, there has not previously been a detailed study of the biological aetiology of this anaemia, exploring the relative contributions of iron, vitamin B12, folate and Vitamin A deficiency, inflammation, genetic haemoglobinopathy, hookworm and malaria. Nor have studies related these aetiologic biological factors to household food security, standard of living and child feeding practices. Barriers to conducting such work have included perceived reluctance of village communities to permit their children to undergo venipuncture, and logistical issues. We have successfully completed a community based, cross sectional field study exploring in detail the causes of anaemia amongst young children in a rural setting.</p> <p>Methods and design</p> <p>A cross sectional, community based study. We engaged in extensive community consultation and tailored our study design to the outcomes of these discussions. We utilised local women as field workers, harnessing the capacity of local health workers to assist with the study. We adopted a programmatic approach with a census rather than random sampling strategy in the village, incorporating appropriate case management for children identified to have anaemia. We developed a questionnaire based on existing standard measurement tools for standard of living, food security and nutrition. Specimen processing was conducted at the Primary Health Centre laboratory prior to transport to an urban research laboratory.</p> <p>Discussion</p> <p>Adopting this study design, we have recruited 415 of 470 potentially eligible children who were living in the selected villages. We achieved support from the community and cooperation of local health workers. Our results will improve the understanding into anaemia amongst young children in rural India. However, many further studies are required to understand the health problems of the population of rural India, and our study design and technique provide a useful demonstration of a successful strategy.</p

Brugia malayi microfilariae adhere to human vascular endothelial cells in a C3-dependent manner

Brugia malayi causes the human tropical disease, lymphatic filariasis. Microfilariae (Mf) of this nematode live in the bloodstream and are ingested by a feeding mosquito vector. Interestingly, in a remarkable co-evolutionary adaptation, Mf appearance in the peripheral blood follows a circadian periodicity and reaches a peak when the mosquito is most likely to feed. For the remaining hours, the majority of Mf sequester in the lung capillaries. This circadian phenomenon has been widely reported and is likely to maximise parasite fitness and optimise transmission potential. However, the mechanism of Mf sequestration in the lungs remains largely unresolved. In this study, we demonstrate that B. malayi Mf can, directly adhere to vascular endothelial cells under static conditions and under flow conditions, they can bind at high (but not low) flow rates. High flow rates are more likely to be experienced diurnally. Furthermore, a non-periodic nematode adheres less efficiently to endothelial cells. Strikingly C3, the central component of complement, plays a crucial role in the adherence interaction. These novel results show that microfilariae have the ability to bind to endothelial cells, which may explain their sequestration in the lungs, and this binding is increased in the presence of inflammatory mediators

A 6 year Geohelminth infection profile of children at high altitude in Western Nepal

<p>Abstract</p> <p>Background</p> <p>Geohelminth infections are a major problem of children from the developing countries. Children with these infections suffer from developmental impairments and other serious illnesses. This study aimed to measure the prevalence of geohelminth infection, infection intensity as well as the change in the intensity in children from Western Nepal over years.</p> <p>Methods</p> <p>This 6-year hospital based prospective study at the Manipal Teaching Hospital, Pokhara included children (< 15 years) visiting the hospital from Kaski and 7 surrounding districts. Samples were also collected from children in the community from different medical camps. Three stool samples from every child were processed using direct and concentration methods. The Kato-Katz technique was used for measuring the intensity of infection.</p> <p>Results</p> <p>The overall prevalence in hospital - attending children was 9.2% with 7.6% in preschool (0 – 5 y) and 11.0% in school-age (6 – 15 y) children, and in community 17.7% with 14.8% in pre-school and 20.5% in school-age children. <it>Ascaris lumbricoides</it>, <it>Trichuris trichiura</it>, <it>Ancylostoma deodenale </it>and <it>Strongyloides stercoralis </it>were the common geohelminths with a gradual decrease in worm load over the years. School-age children were found to be significantly more prone to geohelminth infection as compared to preschool children, but no statistical difference was detected by gender, district as well as season.</p> <p>Conclusion</p> <p>This heavy infection of geohelminths in children should be corrected by appropriate medication and maintaining strict personal hygiene. Health education, clean water, good sewage management and a congenial environment should be ensured to minimise infection.</p

A free weekly iron-folic acid supplementation and regular deworming program is associated with improved hemoglobin and iron status indicators in Vietnamese women

Background Anemia due to iron deficiency is recognized as one of the major nutritional deficiencies in women and children in developing countries. Daily iron supplementation for pregnant women is recommended in many countries although there are few reports of these programs working efficiently or effectively. Weekly iron-folic acid supplementation (WIFS) and regular deworming treatment is recommended for non-pregnant women living in areas with high rates of anemia. Following a baseline survey to assess the prevalence of anemia, iron deficiency and soil transmitted helminth infections, we implemented a program to make WIFS and regular deworming treatment freely and universally available for all women of reproductive age in two districts of a province in northern Vietnam over a 12 month period. The impact of the program at the population level was assessed in terms of: i) change in mean hemoglobin and iron status indicators, and ii) change in the prevalence of anemia, iron deficiency and hookworm infections. Method Distribution of WIFS and deworming were integrated with routine health services and made available to 52,000 women. Demographic data and blood and stool samples were collected in baseline, and three and 12-month post-implementation surveys using a population-based, stratified multi-stage cluster sampling design. Results The mean Hb increased by 9.6 g/L (95% CI, 5.7, 13.5, p < 0.001) during the study period. Anemia (Hb<120 g/L) was present in 131/349 (37.5%, 95% CI 31.3, 44.8) subjects at baseline, and in 70/363 (19.3%, 95% CI 14.0, 24.6) after twelve months. Iron deficiency reduced from 75/329 (22.8%, 95% CI 16.9, 28.6) to 33/353 (9.3%, 95% CI 5.7, 13.0) by the 12-mnth survey, and hookworm infection from 279/366 (76.2%,, 95% CI 68.6, 83.8) to 66/287 (23.0%, 95% CI 17.5, 28.5) over the same period. Conclusion A free, universal WIFS program with regular deworming was associated with reduced prevalence and severity of anemia, iron deficiency and h

Endosymbiont DNA in Endobacteria-Free Filarial Nematodes Indicates Ancient Horizontal Genetic Transfer

Background: Wolbachia are among the most abundant symbiotic microbes on earth; they are present in about 66% of all insect species, some spiders, mites and crustaceans, and most filarial nematode species. Infected filarial nematodes, including many pathogens of medical and veterinary importance, depend on Wolbachia for proper development and survival. The mechanisms behind this interdependence are not understood. Interestingly, a minority of filarial species examined to date are naturally Wolbachia-free. Methodology/PrincipalFindings:We used 454 pyrosequencing to survey the genomes of two distantly related Wolbachia- free filarial species, Acanthocheilonema viteae and Onchocerca flexuosa. This screen identified 49 Wolbachia-like DNA sequences in A. viteae and 114 in O. flexuosa. qRT-PCR reactions detected expression of 30 Wolbachia-like sequences in A. viteae and 56 in O. flexuosa. Approximately half of these appear to be transcribed from pseudogenes. In situ hybridization showed that two of these pseudogene transcripts were specifically expressed in developing embryos and testes of both species. Conclusions/Significance: These results strongly suggest that the last common ancestor of extant filarial nematodes was infected with Wolbachia and that this former endosymbiont contributed to their genome evolution. Horizontally transferred Wolbachia DNA may explain the ability of some filarial species to live and reproduce without the endosymbiont while other species cannot