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Diagnostic services for tuberculosis (TB) are not sufficiently accessible in low-resource settings, where most cases occur, which was aggravated by the COVID-19 pandemic. Early diagnosis of pulmonary TB can reduce transmission. Current TB-diagnostics rely on detection of

Background and purpose: Fibroblast growth factor-2 (FGF-2) and platelet-derived growth factor-A (PDGF-AA) are potent modulators of oligodendrocytes, the main responsible cells for myelin regeneration. We measured FGF-2 and PDGF-AA in the sera and cerebrospinal fluid (CSF) of patients with relapsing-remitting multiple sclerosis (RR-MS) and compared these values with control subjects. Methods: Twenty-three patients with RR-MS and 23 subjects without inflammatory and demyelinating diseases were included. Serum samples of the patients were collected in both relapse and remission phases and were analyzed with ELISA method. CSF was drawn during the relapse period. Blood and CSF were drawn from control subjects for comparison. Wilcoxon and Mann–Whitney U-test and Spearman’s rank correlation were used for analysis. P values of

A range of surface, secreted and somatic antigens from filarial parasites have been studies in order to analyse the response of human infected with these pathogens, and to develop reliable diagnostic and prophylactic agents. Diagnostic procedures, which are urgently required for targetting chemotherapy, are being developed by two techniques. Firstly, detection of host antibody is carried out using selected, specific parasite antigens in the form of recombinant peptides from a filarial DNA library. Secondly, measurement of parasite by a monoclonal antibody "antigen-capture" assay. In addition, a longer-term objective of our collaborative study is to isolate molecules which may stimulate the immune system to mount a protective immune response against filarial parasites. A major focus has been a parasite surface glycoprotein known to be closely conserved between adult worms of Brugia malayi, B. timori and Wuchereria bancrofti. This antigen has been cloned from a cDNA library, and its primary sequence established; in addition to being a constant feature of the adult surface, it is expressed by developing larvae and represents an attractive target for vaccine production. Finally, one of the most intriguing questions in filariasis relates to the genesis of pathological reactions. Although this is a difficult problem, we are now beginning to compare the immune responses of individuals of differing clinical status to certain defined parasite antigens, in an attempt to correlate disease development with particular categories of immune response in infected patients. In this way there is hope to advance the basic understanding of filarial disease, while providing practical means for controlling filariasis at the individual and community levels.

Fibroblast growth factor-2 (FGF-2) and platelet-derived growth factor-A (PDGF-AA) are potent modulators of oligodendrocytes, the main responsible cells for myelin regeneration. We measured FGF-2 and PDGF-AA in the sera and cerebrospinal fluid (CSF) of patients with relapsing-remitting multiple sclerosis (RR-MS) and compared these values with control subjects.Twenty-three patients with RR-MS and 23 subjects without inflammatory and demyelinating diseases were included. Serum samples of the patients were collected in both relapse and remission phases and were analyzed with ELISA method. CSF was drawn during the relapse period. Blood and CSF were drawn from control subjects for comparison. Wilcoxon and Mann-Whitney U-test and Spearman's rank correlation were used for analysis. P values of0.05 were considered significant.Age and sex distribution were similar in both groups. Serum values of FGF-2 were higher in relapse phase compared with remission phase, with a trend toward significance (P=0.052). CSF PDGF-AA showed significant negative correlation with disease duration (correlation coefficient=-0.58, P=0.004). Serum levels of PDGF did not differ between two phases significantly. There was no difference in serum and CSF values of these factors between patients and controls. When we compared patients with prolonged disease with controls, a significant difference between the CSF levels of PDGF-AA was observed (mean±SEM 2.78±0.8 in controls vs. 0.55±0.29 in patients with MS≥2 years, P0.05).Our study was the first to show that CSF PDGF-AA is related to disease duration. Supporting previous findings we showed that serum and CSF levels of these factors are weak indicators of disease severity.

Eczema is a growing problem in Africa, particularly amongst children.To investigate the point-prevalences of eczema by physical examination in schoolchildren living in rural and urban areas and with different socioeconomic backgrounds in Ghana, Gabon and Rwanda. In Ghana period-prevalences were also estimated by questionnaire and compared with the point-prevalences.In total, 4839 schoolchildren in Ghana, Gabon and Rwanda were seen by at least one dermatologist. The point-prevalences of eczema were estimated on the basis of physical examination. Period-prevalences were measured in Ghana with questionnaire based-interviews adapted from the International Study of Asthma and Allergies in Childhood (ISAAC).The point-prevalences were 1.5% and 1.6% in the two Ghanaian studies; 4% in Gabon and 0.8% in Rwanda. The period-prevalences were 2.6% and 4.4% in the two Ghanaian studies. The prevalences of eczema were not significantly different when comparing the urban and rural groups as well as the different socioeconomic levels. The sensitivity and positive predictive value to identify eczema cases based on the questionnaires compared to the diagnoses by physical examination were only 33% and 22% in the first Ghanaian study and 10% and 4% in the second Ghanaian study respectively.The point-prevalences of eczema in the three African countries studied were low compared with industrialized countries. Physical examination by a dermatologist is still the gold standard to identify eczema cases because the sensitivity and the positive predictive value to identify eczema cases with questionnaires were low in the two Ghanaian studies.

Tinea capitis is endemic among schoolchildren in tropical Africa. The objective was to determine the prevalence of symptomatic tinea capitis in schoolchildren in Gabon. A cross-sectional study was conducted with 454 children aged 4-17 years, attending a rural school and an urban school. The diagnosis of tinea capitis was based on clinically manifest infection, direct microscopic examination using 20% potassium hydroxide (KOH) solution and fungal culture. Based on clinical examination, 105 (23.1%) of 454 children had tinea capitis. Seventy-four (16.3%) children were positive by direct examination (KOH) and/or fungal culture. The prevalence of tinea capitis depended on the school studied and ranged from 20.4% in the urban school with a higher socioeconomic status to 26.3% in the rural school with a lower socioeconomic status. Similarly, the spectrum of causative species varied between the different schools. Taken the schools together, Trichophyton soudanense (29.4%) was the most prominent species, followed by Trichophyton tonsurans (27.9%) and Microsporum audouinii (25.0%). Clinically manifest tinea capitis is endemic among schoolchildren in the Lambaréné region in Gabon. The prevalence of tinea capitis and the causative species depended on the type of school that was investigated.

Atopic eczema is an increasing clinical problem in Africa.To determine allergic characteristics and to identify possible risk factors for eczema among schoolchildren in an urbanized area in Ghana.Schoolchildren aged 3-16 years with eczema were recruited. For each patient, one to three age- and sex-matched controls were selected. All children completed a questionnaire and were skin prick tested with a panel of allergens. Blood was drawn to determine the total and allergen-specific IgE. Conditional logistic regression models with the matching factors included in the model were used to calculate the odds ratios and to adjust for possible confounders.A total of 52 children with eczema (27 boys and 25 girls) and 99 controls were included. Levels of total IgE were found to be 9.1 (1.1; 78.4) times more often elevated in children with eczema. This association was mainly driven by elevated IgE levels against cockroach antigen. Children with eczema were found to have 2.0 (0.87; 4.7) times more often positive skin prick tests (SPT), but this association diminished to 1.2 (0.40; 3.6) after adjustment for total IgE levels. Frequent washing with soap was identified as a risk factor for the development of eczema among these children.Schoolchildren with eczema in Ghana were characterized by elevated IgE levels especially against cockroach antigen. The association between eczema and positive SPT was much weaker suggesting immune hyporesponsiveness of the skin. After adjustment for IgE level, SPT were less suitable to distinguish children with and without eczema.

The field of infectious disease immunology is at an exciting intersection with new concepts in immune regulation meeting with the dynamics of infectious diseases. We discuss how the identification of regulatory mechanisms has already helped develop new models to understand helminth infections, which remain among the most prevalent chronic diseases in the world today. The epidemiological imbalance between helminth infections in developing countries, and intensifying allergies and autoimmune pathologies in the industrialised nations, seems to reflect a fundamental shift in regulation of immune responsiveness. Experimental studies have verified that helminths can downmodulate a range of immunopathological conditions, with the regulatory T cell being one of the most common mechanisms in play. We discuss further the context of host genetic predisposition, together with the impact of infection on the evolution of the human immune system, and suggest future strategies to harness our new understanding of helminth organisms in order to control both infectious and non-infectious immunological disorders.

Schistosomes carry lipid moieties that interact with the immune system. To understand the consequence of interactions in terms of polarizing the cytokine profiles, the effect of two Toll-like receptor-2 (TLR2) activating schistosomal lipid fractions was studied on whole blood from Gabonese children living in a schistosomiasis endemic area. One fraction contained lysophosphatidylserine [monoacylglycerophosphoserine (lysoGPSer)] plus diacylphosphatidylserine [diacylglycerophosphoserine (GPSer)] while the other contained lysoGPSer and only a trace of GPSer. The effect of these schistosomal lipid fractions was compared with the known bacterial TLR2 ligands PAM3CSK4 and MALP-2. PAM3CSK4 and MALP-2 had preferential IL-10-activating capacities, while the fraction containing lysoGPSer plus GPSer had a strong TNF-alpha-inducing capacity. The fraction containing lysoGPSer was neutral with respect to pro- vs. anti-inflammatory effects. When Th1 and Th2 cytokines were analysed, the schistosomal lipid fraction containing lysoGPSer plus GPSer showed a stronger Th2 response compared to PAM3CSK4, MALP-2 and lysoGPSer alone. Therefore, the study indicates that not only TLR2 ligands derived from bacteria or from parasites can generate distinct cytokine profiles but also that the composition of lipid entities reaching the immune system can be important in leading to different immune outcomes. This information may be important for exploitation of immune modulatory molecules.

Identification and characterization of risk and protective factors for allergy is important for developing strategies for prevention or treatment. The prevalence of allergy is clearly higher in affluent countries than in developing countries like, e.g. Africa. Especially in urban areas of developing countries, allergy is however on the increase. In Africa, we have the unique opportunity to investigate risk and protective factors and the influence of urbanization and westernization, i.e. almost to take a look at Europe, Australia or the USA as they were before their allergy epidemics. Moreover, migrants from developing to affluent countries experiencing an increased burden of allergy provide new insights into risk and protective factors. Allergen exposure, diet and infections are the major exogenous influences playing a role as risk and protective factors. Depending on the nature, timing, chronicity and level of exposure, each of them can promote or inhibit allergy. Perhaps with the exception of infections, availability of data from Africa on their role in the development of allergy is limited. Detailed epidemiological studies in rural and urban Africa combined with basic immunological research are needed to unravel mechanisms of increase in allergy and of protection. The maturation of the immune system at young age under influence of exogenous factors results in differences in T-cell-skewing (Th1/Th2/Treg) and humoral responses. It is essential to perform studies from a 'non-Eurocentric' angle (e.g. local allergens, locally validated questionnaires and diagnostic procedures). Such studies will provide the affluent countries with new leads to combat the allergy epidemic and more importantly help prevent it in Africa.

We have followed a population in an area endemic for Brugia malayi for three years after intensive treatment with diethylcarbamazine (DEC). Microfilariae were cleared from the circulation within four months in all eligible study participants (n = 60). There appeared to be a strong correlation between the maximum reduction in specific IgG4 and the number of days drug was taken under supervision (p = 0.41, P0.001), indicating that high total dosage of DEC is necessary for optimal reduction of active infection. In individuals with good compliance (at least 180 mg/kg of body weight, n = 34), we observed variable IgG4 patterns. All pre-treatment IgG4+ children (9-14 years old) and 40% of the IgG4+ adult population (or = 15 years old) showed a gradual decrease in anti-filarial IgG4; 53% of these showed complete clearance of worm burden by the end of the study. In contrast, another group of male IgG4+ adults showed IgG4 patterns that started to increase between nine months and two years after treatment, indicating either a partial efficacy of DEC that allowed recovery of resident adult worms or reinfection.

The last decades have seen a dramatic rise in the prevalence of allergic diseases throughout the industrialised world. The "hygiene hypothesis" postulates that this is due to a reduced exposure to infections during childhood. A cohort study in children from Gabon gave us the unique opportunity to examine the relationship between exposure to P. falciparum and atopy. 91 children, who had been closely followed for an average of 5 years and of whom the exact incidence of malaria attacks was known, underwent a skin-prick test with mite antigen. 16 children (18%) had a positive reaction. Gender or age had no effect on the outcome of the test. However, those tested positive had had less infections and a lower incidence of malaria than children tested negative (p = 0.017). Survival analysis shows that children with a high exposure to P. falciparum were at lower risk of an atopic skin reaction (p = 0.001). We postulate that the low exposure to the malaria parasite contributes to the development of an imbalanced immune system with a subsequent higher reactivity to the allergen tested. Immuno-suppression is commonly seen during a malaria attack and this correlates positively with the level of anti-inflammatory cytokines such as interleukin-10. High exposure to parasite antigens might counterbalance pro-inflammatory immune reactions and thus protect against allergic diseases. A better understanding of the relationship between parasitic infection and allergy will help us to develop strategies to prevent allergic disease without being exposed to infectious diseases.

This study reports reversals in microfilarial density and the accompanying changes in cellular immune responses to filarial antigens of 39 individuals (11 microfilaria-positives, 22 microfilaria-negatives and six converters) living in an area endemic for brugian filariasis. Microfilarial counts decreased from April, the end of the rainy season to July, middle of the dry season (g.m. 88 mf/ml and 38 mf/ml, respectively; P = 0.001) and subsequently increased in November, the beginning of the rainy season (P = 0.088). Whereas the proliferative responses remained low throughout the study period in microfilaraemic individuals, in amicrofilaraemics these responses changed in the opposite direction to that of microfilarial densities. In three converters, proliferation changed in the opposite direction to the presence or absence of microfilariae. Cytokine analysis in the converters revealed that interferon-gamma was most affected by the shifts in microfilarial densities. In contrast, interleukin-4 responses showed little correlation with changes in parasite densities.

To understand the intricate balance and the coordinate expression of the Th1 and Th2 cytokines following a natural mode of T cell triggering, antigen-stimulated IL-4, IL-13 and IFN-gamma production was studied in primary peripheral blood mononuclear cell cultures at a single-cell level. Cells from filariasis patients who respond to parasite antigen by producing not only IFN-gamma but also IL-4 and IL-13 were stimulated with Brugia malayi adult worm antigen and analyzed for co-expression of cytokines by intracellular staining. IL-4 and IL-13 were frequently co-expressed (54% of IL-4+ cells stained for IL-13 and 29% of IL-13+ cells expressed IL-4 at all time points), whereas IFN-gamma expression was totally segregated from both IL-4 and IL-13. These data indicate that in human peripheral T cells the co-expression of the dominant Th1 and Th2 cytokines within a single cell is a rare event and that IL-13 is clearly more frequently associated with a Th2 than a Th1 type response in primary T cell cultures.

Human infections with filarial nematodes such as Brugia malayi are accompanied by unusually high titres of parasite-specific IgG4 antibodies. We have compared the profile of antigens recognised by filarial-specific IgG1 and IgG4 isotypes by Western blotting. Serum samples were collected from 120 subjects exposed to Brugia malayi, divided into three groups of asymptomatic amicrofilaraemic (endemic normal), microfilaraemic, and elephantiasis patients. Antigen preparations were tested from both adult B. malayi parasites, and from microfilariae; 24 distinct bands were analysed from the former, and 19 from the latter. Both qualitative scoring for band reactivity, and densitometric scanning of major bands, were employed. The consistent result was one of high and preferential IgG4 reactivity to a set of low molecular weight bands, of 15, 17, 20, 31 and 33 kDa; most of the 19 other bands showed higher reactivity with IgG4. Analysis of Western blot patterns showed an overall tendency for stronger IgG4 responses in microfilaraemic cases, and higher IgG1 responses in elephantiasis patients, consistent with published studies using ELISA on unfractionated parasite extracts. This study has defined an array of filarial antigens from each stage, and relative levels of IgG4 recognition, which will be important in unravelling distinct immune responses to this complex parasite.

Parasitic worms of the genus Schistosoma excrete relatively large amounts of immunogenic glycoproteins (circulating cathodic antigen [CCA]) that contain polysaccharide side chains with the trisaccharide Lewis-x (L(ex)) as a repeating unit. These carbohydrates evoke high titers of specific IgM antibodies that cross-react with the repeating L(ex) units on the surface of granulocytes. Consequently this might lead, in the presence of complement, to lysis of the granulocytes. In the present study, this hypothesis was investigated using anti-CCA mouse monoclonal antibodies (MoAbs) and polyclonal antibodies purified from sera of infected humans. By flow cytometry, it was demonstrated that the mouse MoAbs directed against CCA strongly recognized the granulocytes. It could also be shown that these MoAbs, as well as anti- CCA IgM antibodies purified from infected human sera, caused lysis of granulocytes in a complement-dependent cytotoxicity assay. Sera from healthy controls or from patients with other helminth infections resulted in negligible granulocytotoxicity. These in vitro observed phenomena may explain the mild to moderate neutropenia that occurs in schistosomiasis patients.

CD30 is a member of the TNF receptor superfamily that is commonly used as a marker for Hodgkin and Reed-Sternberg cells in Hodgkin's disease. More recently, it has been proposed that CD30 is preferentially up-regulated on Th2-type human T cells. We analyzed regulation of CD30 expression on both peripheral blood T cells and T cell clones. In short-term culture, CD30 expression could be induced on T cells by Ags that elicit Th2-type responses (Schistosoma haematobium, adult worm Ag, and Toxocaria canis, excretory/secretory Ag) and Th0-type responses (tetanus toxoid), as well as Th1-type responses (tuberculin purified protein derivative). Moreover, simultaneous measurement of membrane phenotype and cytokine production showed that CD30-expressing cells can produce IFN-gamma. Finally, within panels of randomly generated as well as Ag-specific T cell clones, CD30 expression was found on Th0-, Th2-, and Th1-type clones. We conclude that induction of CD30 on activated T cells is not related to differentiation in Th0-, Th1-, or Th2-type cells.

A population of 164 adult individuals resident in an area endemic for Brugia malayi lymphatic filariasis has been studied for humoral immune responses to filarial parasites. Antibody levels to Ag extracted from adult worms were determined for each of the IgG subclasses, for IgM and for IgE. The dominant isotype of antifilarial antibody was IgG4, which represented 88% of total IgG in asymptomatic microfilaremics, most of whom possessed 100 to 1000 micrograms/ml of specific antibody of this subclass (geometric mean 762 micrograms/ml). Patients with chronic disease (elephantiasis), who were generally amicrofilaremic, had substantially higher levels of IgG1, IgG2, and IgG3, but a 3.4-fold lower geometric mean level of specific IgG4 (222 micrograms/ml) than asymptomatics with or without microfilaremia. In contrast, specific IgE antibody levels in cases of elephantiasis were on average 4.5 times higher than those found in the asymptomatic carrier state. The majority of microfilaremics were therefore typified by extremely high specific IgG4 concentrations and relatively low IgE reactivities, whereas clinical cases tended to show the reverse relationship. The possible roles of these isotypes and the implications of changing IgG4/IgE balances in disease are discussed.

To study the cytotoxic reactions responsible for mediating eosinophil damage to schistosomula of Schistosoma mansoni, we have used cytoplasts (eosinophil or neutrophil vesicles devoid of granules and nuclei, with an intact oxidase in their plasma membrane) in combination with purified eosinophil cationic protein (ECP) or major basic protein (MBP) in a cytotoxicity test toward schistosomula. Suboptimal concentrations of ECP (10(-6) M) or MBP (10(-6) M) resulting in less than 10% killing were used in combination with cytoplasts. Cytoplasts alone in the presence of immune serum tested over a wide range of cytoplast:schistosomula ratios generated superoxide and hydrogen peroxide, but were unable to damage schistosomula. However, when a suboptimal ECP concentration (10(-6) M) was combined with neutroplasts or eosinoplasts, 43.9% +/- 8.5 (n = 7) and 24.7% +/- 9.8 (n = 3), respectively, of the schistosomula were killed. Oxygen metabolites were responsible for the synergism, because cytoplasts from a patient with chronic granulomatous disease were unable to act in synergy with ECP. In contrast to ECP, no synergism was found between cytoplasts and MBP (10(-6) to 2 X 10(-5)M). These results show that oxygen metabolites are important for the killing of schistosomula by lowering the concentration of ECP needed to inflict damage.

After interaction with so-called priming agents, the respiratory burst in human granulocytes does not become activated, but is enhanced upon subsequent stimulation with the chemoattractant FMLP. Investigating the mechanism of the priming reaction, we found that a transient rise in the cytosolic free calcium concentration [( Ca2+]i) suffices to irreversibly prime human granulocytes. Thus, platelet-activating factor (PAF) induced a transient increase in [Ca2+]i and primed the cells to an enhanced respiratory burst upon subsequent interaction with FMLP. Artificially, the transient rise in [Ca2+]i was mimicked by addition and subsequent removal of the Ca2+ ionophore ionomycin; this treatment too, primed the respiratory burst of the granulocytes. The priming induced by ionomycin was completely abolished when [Ca2+]i changes were buffered during exposure of the cells to the ionophore. The priming induced by PAF was only partially inhibited under [Ca2+]i-buffering conditions during priming, indicating that multiple pathways exist in the priming of granulocytes by PAF.

Eosinophils from the blood of normal individuals were purified by centrifugation over discontinuous Percoll gradients. Eosinophil suspensions were obtained with a mean purity of 96% and a mean recovery of 64% (n = 19). When incubated with phorbol-myristate acetate, eosinophils consumed twice as much oxygen as did neutrophils from the same donors. With serum-treated zymosan, 70% and 100% of the maximal oxidative response (i.e. the response to phorbol-myristate acetate) was obtained with eosinophils and neutrophils, respectively. The calcium ionophore A23187 is a weak stimulus that triggered only 2.5% of the eosinophil and 10% of the neutrophil oxidative capacity. The response of both cell types to formyl-methionyl-leucyl-phenylalanine (fMLP) was rapid, with a maximum after 3 min. The magnitude of this eosinophil reaction was half that of neutrophils. Although the activities of the granule enzymes beta-glucuronidase and arylsulphatase were 2.5 and 6 times higher in eosinophils than in neutrophils, respectively, the exocytosis of these enzymes in response to various stimuli was lower in eosinophils. The high yield of eosinophils from our separation method enabled us to prepare eosinoplasts by centrifugation of eosinophils over discontinuous Ficoll gradients that contained cytochalasin B. Eosinoplasts are plasma membrane vesicles derived from eosinophils, filled with cytoplasm but devoid of granules and nucleus. The eosinoplasts contained 30% of the cytoplasm and plasma membrane present in intact eosinophils. Eosinoplasts still possessed a functionally intact oxidase enzyme that could be stimulated with various stimuli. Therefore, eosinoplasts may provide a valuable tool to study separately the role of the oxidase products and that of the granule contents in eosinophil functions.

Resealed erythrocyte membranes (ghosts) filled with (Fe3+)cytochrome c were used as an assay system to measure the release of superoxide (O-2) from human phagocytes into the incubation medium. Neutrophils, activated by either opsonized zymosan particles or the soluble stimulus phorbol myristate acetate, released O-2, which subsequently entered the ghosts and reduced (Fe3+)cytochrome c. This reaction was dependent on the time of incubation, the concentration of neutrophils, the concentration of stimulus, and the concentration of ghosts. The reaction was completely inhibited by superoxide dismutase and by 4,4'-diisothiocyano-2,2'-disulfonic acid, a specific blocker of anion channels in membranes. The reduction of (Fe3+)cytochrome c free in solution was about four times as fast as the reduction of (Fe3+)cytochrome c in the ghosts. Human eosinophils stimulated by phorbol myristate acetate reacted similarly to human neutrophils; the rate of O-2 production/cell was about twice as high for eosinophils as for neutrophils. In contrast, eosinophils stimulated with opsonized zymosan particles only reduced (Fe3+)cytochrome c free in solution, but not (Fe3+)cytochrome c in ghosts. This lack of reaction was not due to production of an inhibitor or below threshold generation of O-2 for the ghost assay. These results indicate: 1) activated human neutrophils and eosinophils can release O-2 or a similar product into the incubation medium; and 2) reduction of (Fe3+)cytochrome c free in solution is no proof for O-2 excretion by phagocytes.

Controlled human infection (CHI) models are an important research tool. Healthy volunteers are experimentally infected with a pathogen. In malaria research the model has been used for decades. Here, the model was used to test new Plasmodium falciparum strains, NF135.C10 an NF166.C8, and compare these with the commonly used strain NF54. In addition, a genetically modified malaria vaccine, PfSPZ-GA1, was tested. Unfortunately only few volunteers were protected against malaria.For schistosomiasis, a controlled human schistosomiasis infection (CoHSI) model was developed and hereafter a dose finding study with single-sex male only cercariae was performed. This study showed that in 80% of volunteers 20 cercariae were effective to induce an infection. Although the use of 30 cercariae resulted in 100% infection rate two out of three volunteers developed Katayama syndrome. These side effects were less after infection with 20 cercariae.At last suggestions were made to further improve the CHI model by the use of historical controls. Although this study design can only be used in CHI’s with well-know outcomes these study will be safer by reducing the cumulative risks as less volunteers can be used for these trials.