David Diemert

Soil-transmitted helminth infections: ascariasis, trichuriasis, and hookworm

The three main soil-transmitted helminth infections, ascariasis, trichuriasis, and hookworm, are common clinical disorders in man. The gastrointestinal tract of a child living in poverty in a less developed country is likely to be parasitised with at least one, and in many cases all three soil-transmitted helminths, with resultant impairments in physical, intellectual, and cognitive development. The benzimidazole anthelmintics, mebendazole and albendazole, are commonly used to remove these infections. The use of these drugs is not limited to treatment of symptomatic soil-transmitted helminth infections, but also for large-scale prevention of morbidity in children living in endemic areas. As a result of data showing improvements in child health and education after deworming, and the burden of disease attributed to soil-transmitted helminths, the worldwide community is awakening to the importance of these infections. Concerns about the sustainability of periodic deworming with benzimidazole anthelmintics and the emergence of resistance have prompted efforts to develop and test new control tools.

Phase 1 Clinical Trial of Apical Membrane Antigen 1: an Asexual Blood-Stage Vaccine for Plasmodium falciparum Malaria

ABSTRACT Apical membrane antigen 1 (AMA1), a polymorphic merozoite surface protein, is a leading blood-stage malaria vaccine candidate. A phase 1 trial was conducted with 30 malaria-naïve volunteers to assess the safety and immunogenicity of the AMA1-C1 malaria vaccine. AMA1-C1 contains an equal mixture of recombinant proteins based on sequences from the FVO and 3D7 clones of Plasmodium falciparum. The proteins were expressed in Pichia pastoris and adsorbed on Alhydrogel. Ten volunteers in each of three dose groups (5 μg, 20 μg, and 80 μg) were vaccinated in an open-label study at 0, 28, and 180 days. The vaccine was well tolerated, with pain at the injection site being the most commonly observed reaction. Anti-AMA1 immunoglobulin G (IgG) was detected by enzyme-linked immunosorbent assay (ELISA) in 15/28 (54%) volunteers after the second immunization and in 23/25 (92%) after the third immunization, with equal reactivity to both AMA1-FVO and AMA1-3D7 vaccine components. A significant dose-response relationship between antigen dose and antibody response by ELISA was observed, and the antibodies were predominantly of the IgG1 isotype. Confocal microscopic evaluation of sera from vaccinated volunteers demonstrated reactivity with P. falciparum schizonts in a pattern similar to native parasite AMA1. Antigen-specific in vitro inhibition of both FVO and 3D7 parasites was achieved with IgG purified from sera of vaccinees, demonstrating biological activity of the antibodies. To our knowledge, this is the first AMA1 vaccine candidate to elicit functional immune responses in malaria-naïve humans, and our results support the further development of this vaccine.

Developing vaccines to combat hookworm infection and intestinal schistosomiasis

Hookworm infection and schistosomiasis rank among the most important health problems in developing countries. Both cause anaemia and malnutrition, and schistosomiasis also results in substantial intestinal, liver and genitourinary pathology. In sub-Saharan Africa and Brazil, co-infections with the hookworm, Necator americanus, and the intestinal schistosome, Schistosoma mansoni, are common. The development of vaccines for these infections could substantially reduce the global disability associated with these helminthiases. New genomic, proteomic, immunological and X-ray crystallographic data have led to the discovery of several promising candidate vaccine antigens. Here, we describe recent progress in this field and the rationale for vaccine development.

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.

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.

Generalized urticaria induced by the Na-ASP-2 hookworm vaccine: Implications for the development of vaccines against helminths

BACKGROUND Necator americanus Ancylostoma-secreted protein 2 (Na-ASP-2) is secreted by infective hookworm larvae on entry into human hosts. Vaccination of laboratory animals with recombinant Na-ASP-2 provides significant protection against challenge infections. In endemic areas antibodies to Na-ASP-2 are associated with reduced risk of heavy N americanus infections. OBJECTIVE To assess the safety and immunogenicity of recombinant Na-ASP-2 adjuvanted with Alhydrogel in healthy Brazilian adults previously infected with N americanus. METHODS Participants were randomized to receive Na-ASP-2 or hepatitis B vaccine. Major IgG and IgE epitopes of the Na-ASP-2 molecule were mapped by using sera from these same subjects. Seroepidemiologic studies in adults and children residing in hookworm-endemic areas were conducted to assess the prevalence of IgE responses to Na-ASP-2. RESULTS Vaccination with a single dose of Na-ASP-2 resulted in generalized urticarial reactions in several volunteers. These reactions were associated with pre-existing Na-ASP-2-specific IgE likely induced by previous hookworm infection. Surveys revealed that a significant proportion of the population in hookworm-endemic areas had increased levels of IgE to Na-ASP-2. Epitope mapping demonstrated sites on the Na-ASP-2 molecule that are uniquely or jointly recognized by IgG and IgE antibodies. CONCLUSION Infection with N americanus induces increased levels of total and specific IgE to Na-ASP-2 that result in generalized urticaria on vaccination with recombinant Na-ASP-2. These data advance knowledge of vaccine development for helminths given their propensity to induce strong T(H)2 responses. Study data highlight the important differences between the immune responses to natural helminth infection and to vaccination with a recombinant helminth antigen.

Randomized, placebo-controlled, double-blind trial of the Na-ASP-2 hookworm vaccine in unexposed adults.

Necator americanus Ancylostoma Secreted Protein-2 (Na-ASP-2) is a leading larval-stage hookworm vaccine candidate. Recombinant Na-ASP-2 was expressed in Pichia pastoris and formulated with Alhydrogel. In a phase 1 trial, 36 healthy adults without history of hookworm infection were enrolled into 1 of 3 dose cohorts (n=12 per cohort) and randomized to receive intramuscular injections of either Na-ASP-2 or saline placebo. Nine participants in the first, second and third cohorts were assigned to receive 10, 50 and 100 microg of Na-ASP-2, respectively, on study days 0, 56 and 112, while 3 participants in each cohort received placebo. The most frequent adverse events were mild-to-moderate injection site reactions; in 8 participants these were delayed and occurred up to 10 days after immunization. No serious adverse events occurred. Anti-Na-ASP-2 IgG endpoint titers as determined by ELISA increased from baseline in all vaccine groups and peaked 14 days after the third injection, with geometric mean titers of 1:7066, 1:7611 and 1:11,593 for the 10, 50 and 100 microg doses, respectively, compared to <1:100 for saline controls (p<0.001). Antibody titers remained significantly elevated in all vaccine groups until the end of the study, approximately 8 months after the third vaccination. In vitro stimulation of PBMCs collected from participants with Na-ASP-2 resulted in robust proliferative responses in those who received vaccine, which increased with successive immunizations and remained high in the 50 and 100 microg dose groups through the end of the study. This first trial of a human hookworm vaccine demonstrates that the Na-ASP-2 vaccine is well-tolerated and induces a prolonged immune response in adults not exposed to hookworm, justifying further testing of this vaccine in an endemic area.

Prevention and Self-Treatment of Traveler's Diarrhea

SUMMARY Of the millions who travel from the industrialized world to developing countries every year, between 20% and 50% will develop at least one episode of diarrhea, making it the most common medical ailment afflicting travelers. Although usually a mild illness, travelers diarrhea can result in significant morbidity and hardship overseas. Precautions can be taken to minimize the risk of developing travelers diarrhea, either through avoidance of potentially contaminated food or drink or through various prophylactic measures, including both nonpharmacological and antimicrobial strategies. If diarrhea does develop despite the precautions taken, effective treatment—usually a combination of an antibiotic and an antimotility agent—can be brought by the traveler and initiated as soon as symptoms develop. In the future, vaccines—several of which are in the advanced stages of clinical testing—may be added to the list of prophylactic measures.

Population structure of the genes encoding the polymorphic Plasmodium falciparum apical membrane antigen 1: Implications for vaccine design

Immunization with the highly polymorphic Plasmodium falciparum apical membrane antigen 1 (PfAMA1) induces protection in animals but primarily against parasites that express the same or similar alleles. One strategy to overcome the obstacle of polymorphism is to combine PfAMA1 proteins representing major haplotypes into one vaccine. To determine the minimum number of haplotypes that would confer broad protection, we sequenced the coding region of PfAMA1 from 97 clones from around the world and 61 isolates from Mali, identifying 150 haplotypes for domains 1 to 3 that included previous sequences. A clustering algorithm grouped the 150 haplotypes into six populations that were independent of geographic location. Each of the six populations contained haplotypes predominantly of that population (predominant haplotypes) and haplotypes that were a mixture of haplotypes represented in other populations (admixed haplotypes). To determine the biological relevance of the populations identified through the clustering algorithm, antibodies induced against one predominant haplotype of population 1 (3D7) and one admixed haplotype of population 5 (FVO) were tested for their ability to block parasite invasion of erythrocytes. Parasites expressing PfAMA1s belonging to population 1 were efficiently inhibited by 3D7-specific antibodies, whereas parasites expressing PfAMA1s belonging to other populations were not. For FVO-specific antibodies, we observed growth inhibition against itself as well as isolates belonging to populations 3 and 6. Our data suggests that the inclusion of PfAMA1 sequences from each of the six populations may result in a vaccine that induces protective immunity against a broad range of malaria parasites.

The Human Hookworm Vaccine.

Highlights ► Human hookworm infection is a leading cause of iron deficiency anemia. ► An estimated 700 million people in developing countries are affected. ► The Sabin Vaccine Institute PDP is developing the vaccine in collaboration with FIOCRUZ. ► The vaccine comprises two recombinant protein antigens on alum and a TLR4 agonist. ► The partnerships plan is that the vaccine will be licensed by 2020.