Heritability of human hookworm infection in Papua New Guinea.

Hookworms infect approximately 740 million humans worldwide and are an important cause of morbidity. The present study examines the role of additive genetic effects in determining the intensity of hookworm infection in humans, and whether these effects vary according to the sex of the host. Parasitological and epidemiological data for a population of 704 subjects in Papua New Guinea were used in variance components analysis. The 'narrow-sense' heritability of hookworm infection was estimated as 0.15+/-0.04 (P<0.001), and remained significant when controlling for shared environmental (household) effects. Allowing the variance components to vary between the sexes of the human host consistently revealed larger additive genetic effects in females than in males, reflected by heritabilities of 0.18 in females and 0.08 in males in a conservative model. Household effects were also higher in females than males, although the overall household effect was not significant. The results indicate that additive genetic effects are an important determinant of the intensity of human hookworm infection in this population. However, despite similar mean and variance of intensity in each sex, the factors responsible for generating variation in intensity differ markedly between males and females.

Alpha+ -thalassaemia and pregnancy in a malaria endemic region of Papua New Guinea.

The effect of maternal alpha+ -thalassaemia on pregnancy was assessed in the north coastal region of Papua New Guinea (PNG), where malaria is hyperendemic and alpha+ -thalassaemia is extremely common. In a prospective study of 987 singleton hospital deliveries, we correlated maternal alpha-globin genotype with markers of reproductive fitness (age in primigravidae, gravidity, pregnancy interval and the number of miscarriages and stillbirths), Plasmodium falciparum(P. falciparum) infection of the mother and placenta, maternal haemoglobin, preterm delivery and birthweight. The frequency of the -alpha genotype in mothers was 0.61. Markers of reproductive fitness were similar in women with and without alpha+ -thalassaemia. Median haemoglobin concentration during pregnancy and after delivery was about 1.0 g/dl lower in homozygous alpha+ -thalassaemia than in women with a normal alpha- globin genotype (P < or = 0.001). The frequency of placental P. falciparum infection and systemic malaria infection after delivery showed no consistent relationship to alpha-globin genotype. The frequency of preterm delivery and low birthweight did not vary significantly according to maternal alpha-globin genotype. Maternal alpha+ -thalassaemia does not affect reproductive fitness or susceptibility to malaria during pregnancy. Although median haemoglobin concentration was significantly lower in mothers homozygous for alpha+ -thalassaemia than those with a normal alpha-globin genotype, this did not result in an adverse outcome of pregnancy.

Predisposition to hookworm infection in Papua New Guinea.

Reinfection with hookworm (Necator americanus) following chemotherapy was studied over 8 years in a rural village in Madang Province, Papua New Guinea. Faecal egg counts were performed on up to 202 individuals in July 1988, August 1990 and November 1996; the study population was treated after sampling in 1988 and 1990. Reinfection burdens in 1996 did not differ significantly from pretreatment burdens (in 1988), and were significantly higher than burdens in 1990. However, the prevalence of hookworm infection was significantly lower in 1996 than in either 1988 or 1990. There was significant predisposition to high or low hookworm burden between 1990 and 1996; this predisposition was stronger in children than adults. However, there was no detectable predisposition between 1988 and 1996 in individuals who had been treated 2 or more times between surveys. The mean weight of adult hookworms in individual hosts was measured in 1988 and 1990 using worms expelled after chemotherapy. There was a significant positive correlation between mean male hookworm weight in the 2 years, suggesting that individual hosts are predisposed to infection with heavy or light hookworms. These data suggest that differences in host susceptibility are involved in generating predisposition, but that longer-term variation in either exposure or susceptibility limits the period over which significant predisposition can be detected.

Causes of preterm delivery and intrauterine growth retardation in a malaria endemic region of Papua New Guinea.

AIM: To identify causes of preterm delivery and intrauterine growth retardation (IUGR) in a malaria endemic region of Papua New Guinea. METHODS: Independent predictors of preterm delivery and birthweight in term infants were identified using multiple regression analysis in a prospective study of 987 singleton live births delivered in Madang Hospital. RESULTS: Overall, Plasmodium falciparum infection of the placenta was associated with a reduction in birthweight of 130 g. Malaria was significantly more common in primigravidae than multigravidae and probably contributed to both preterm delivery and IUGR. Maternal haemoglobin concentrations were significantly lower in malaria infected than noninfected women and reduced haemoglobin was the main determinant of preterm delivery. Poorer maternal nutritional status and smoking were associated with both prematurity and IUGR. Greater antenatal clinic attendance predicted increased birthweight in term infants. CONCLUSIONS: Protection against malaria during pregnancy, especially in primigravidae, improved nutrition in women and discouragement of smoking would probably reduce both preterm delivery and IUGR. Greater use of existing antenatal clinics might increase birthweight in term infants.

Resistance of Plasmodium falciparum malaria to amodiaquine, chloroquine and quinine in the Madang Province of Papua New Guinea, 1990-1993.

The in vivo response of Plasmodium falciparum parasites to amodiaquine or chloroquine was assessed in children with symptomatic malaria attending different health facilities in the Madang area. Among the 27 subjects who were completely followed up, 4 (15%) were infected with parasites fully susceptible and 23 (85%) with parasites exhibiting some degree of resistance. Out of the latter group, 52% were of RI level, 26% RII and 22% RIII. 14 subjects out of 42 (33%) failed to clear their parasitaemia by day 7 and 92 out of 134 (69%) had persistent or recrudescent parasitaemia at day 21. The level of in vivo resistance was similar for amodiaquine and chloroquine. 86% of the isolates tested in vitro showed resistance to amodiaquine, 86% to chloroquine and 7% to quinine. In ten years the prevalence of resistant isolates in vivo has increased from 47% to 85%. Of more concern is the shift from RI level of resistance to RII and RIII: the proportion of resistant strains that were RI dropped from 90% to 52% over the ten-year period. To determine if the standard antimalarial regimens are still appropriate, there is a need not only to assess the level of parasite resistance but also the prevalence of treatment failure in different parts of Papua New Guinea.

PIP: The in vivo response of Plasmodium falciparum parasites to amodiaquine and chloroquine was assessed in children 1-9 years of age with symptomatic malaria recruited from health centers in Papua New Guinea's Madang area. Among the 27 children who were completely followed up, 4 (15%) were infected with fully susceptible parasites; in the remaining 23 cases (85%), there was some degree of resistance. 52% of parasites in the latter group were RI level, 26% RII, and 22% RIII. There was no correlation between level of resistance and age. 14 out of 42 children (33%) failed to clear their parasitemia by day 7 and 92 out of 134 (69%) had persistent or recrudescent parasitemia at day 21. Both amodiaquine and chloroquine had similar levels of in vivo resistance. 86% of isolates tested in vitro showed resistance to amodiaquine, 86% to chloroquine, and 7% to quinine. In 10 years, the prevalence of resistant isolates in vivo has increased from 47% to 85%. Of particular concern is the finding that the proportion of resistant strains that were RI dropped from 90% to 52% over this decade. The increase of resistance is attributed to indiscriminate use of 4-aminoquinolines in all cases of fever. Needed, to assess whether standard antimalarial regimens are still appropriate, is a review of treatment failure in different parts of Papua New Guinea.

Human cerebral malaria: lack of significant association between erythrocyte rosetting and disease severity.

The ability of Plasmodium falciparum isolates from 103 Papua New Guinea children with cerebral malaria and 158 children with uncomplicated malaria to form rosettes in vitro was studied. Of these, 81 isolates from cerebral malaria and 151 isolates from uncomplicated malaria grew to schizogony and were included in the rosetting analysis. Wide variation occurred in the level of rosette formation, with all isolates from both cerebral and uncomplicated malaria patients being able to form rosettes. No statistically significant difference existed between the geometric mean rosetting rate of isolates obtained from cerebral malaria and those from uncomplicated malaria (9% versus 8.6%, P = 0.27). The ability of acute sera to inhibit rosette formation was not significantly different between 18 cerebral malaria cases and 20 controls tested [mean reduction in rosetting rate 6.1% (SD 11.5) versus 8.4% (SD 12.3), P = 0.57]. The rosetting rate of cerebral malaria cases was not associated with the clinical outcome. Among the clinical and laboratory variables tested, only blood group and parasite density were significantly associated with rosetting. These data do not support the hypothesis that rosette formation is associated with cerebral malaria in Papua New Guinea, but indicate that rosetting is an intrinsic property of parasites occurring in all manifestations of the disease.

Dynamics of malaria parasitaemia associated with febrile illness in children from a rural area of Madang, Papua New Guinea.

Active community and self-reporting surveillance techniques have been used to describe the dynamics of febrile illness and associated malaria infection in children aged 2 to 15 years from a rural area of Madang Province, Papua New Guinea (PNG). Both history of fever and fever in association with parasitaemia appeared to be reliable indicators of malaria morbidity in this endemic area. Parasite density was observed to be a major determinant of mild malarial disease at both the population level and within an individual. Age-specific prevalence of febrile illness correlated with age-specific patterns of parasite density but not of parasite prevalence. Seasonal changes in fever incidence correlated with parasite density. The transition from afebrile to febrile state within an individual was generally associated with an increase in parasite density. Surveillance and self-reported febrile cases (which differ in severity on the basis of the perceived need for treatment) could be distinguished on the basis of parasite density. Thus surveillance techniques divide clinical malaria in rural PNG into 'mild' and 'very mild' forms. The age-specific pattern of decline of prevalence of malaria-associated febrile illness and parasite density is best explained by induction of strain-specific anti-disease immunity upon infection with a given strain of Plasmodium falciparum. The fever threshold in self-reporting febrile cases was seen to decrease with age and can be explained by an age-specific decline in anti-toxic immunity.

Resistance of Plasmodium falciparum to amodiaquine in Papua New Guinea.

The sensitivity of Plasmodium falciparum to amodiaquine was assessed in children in the Madang region of Papua New Guinea. Fifty-four tests in vivo were carried out and 19 (35%) of these showed resistance at the RII or RIII level. Twenty-two of the 43 isolates tested showed some degree of resistance to amodiaquine in vitro. A negative correlation between the age of the patient and apparent resistance of the parasite in vivo was observed. There was no correlation between resistance in vivo and in vitro.

Isotypic variation in antibody responses in a community in Papua New Guinea to larval and adult antigens during infection, and following reinfection, with the hookworm Necator americanus.

The natural infection of a community with the hookworm Necator americanus induces a vigorous humoral response to both larval and adult parasite antigens. This response occurs in all five human antibody isotypes, and data are presented to show that, at the population level, isotypes respond differently, following chemotherapy and during reinfection, to changes in antigen stimulation. The differential response probably reflects the fact that the parasite, during the course of its life cycle, presents different amounts of antigens at different anatomical locations. It is suggested that IgG and IgM responses against adult excretory-secretory (ES) products most accurately reflect the efficacy of chemotherapy, and the load of resident adult infection, while IgG responses against larval somatic antigens reflect continuous exposure to infection. These hypotheses should now be tested, at the level of the individual, in a longitudinal manner using more closely spaced sampling intervals. This repetitive sampling, and the inclusion of a measure of the exposure of the population to infective stages, will allow more definitive conclusions to be made about the role of the immune response in controlling worm burdens.

Antigenic cross-reactivity between Necator americanus and Ascaris lumbricoides in a community in Papua New Guinea infected predominantly with hookworm.

Sero-epidemiological data are presented in which antigenic cross-reactivity between Necator americanus and Ascaris lumbricoides has been investigated in a community in Papua New Guinea infected predominantly with N. americanus. It is our contention that the antigenic cross-reactivity which undoubtedly exists between these species accounted for (i) a peak in antibody levels against N. americanus in 10-13 years old children (driven by infection with A. lumbricoides), and (ii) the maintenance of apparent antibody levels against A. lumbricoides in older age groups (driven by infection with N. americanus in the absence of overt infection with A. lumbricoides). Cross-reactivity was analysed further, and apparently N. americanus-specific epitopes identified, by immunoblotting. These observations could have considerable bearing on the interpretation of data from sero-epidemiological studies which failed to take account of concurrent infection with these parasites.

Serological evaluation of the macrofilaricidal effects of diethylcarbamazine treatment in bancroftian filariasis.

An Mr 200,000 phosphorylcholine-containing antigen (PC-Ag) of predominantly adult worm origin was found in the sera of humans infected with Wuchereria bancrofti. This paper describes results of a longitudinal study of changes in levels of PC-Ag in response to diethylcarbamazine (DEC) therapy as measured by two-site immunoradiometric assay (IRMA) and Western blotting. One hundred thirty-two residents of a bancroftian filariasis-endemic area of Papua New Guinea (PNG) were treated with a 72 mg/kg dose of DEC. A macrofilaricidal effect was seen with this dose of DEC as 34% of the treated subjects had localized side effects and long-term decreases in microfilariae (mf) counts were observed 12 months after treatment. The PC-Ag levels were reduced to 72%, 52%, and 51% of pretreatment values at 21 days and at six and 12 months after treatment. These decreases, observed by IRMA, were specifically associated with loss of the Mr 200,000 PC-Ag detected by immunoadsorption and Western blotting. From drug treatment data, the maximum half-life of PC-Ag in circulation was calculated to be 50 days, assuming a first-order decay process. This maximum half-life indicates that persistent antigenemia observed in the majority of treated subjects could only result from the survival of adult worms. In the absence of methods to directly demonstrate W. bancrofti adult worms, detection of serum PC-Ag levels provides a sensitive indirect measure of the dynamics of adult worm populations. This serological measurement may be useful in optimizing the macrofilaricidal and therapeutic effects of DEC and in assessing the macrofilaricidal action of new antifilarial drugs and immunological interventions.

Hookworm (Necator americanus) infection and storage iron depletion.

The relationship between iron status and the intensity of infection with hookworm was investigated in a rural population on Karkar Island, Mandang Province, Papua New Guinea. There was a significant negative correlation between plasma ferritin level and hookworm burden, which was strongest in males. In contrast, there was no correlation between plasma ferritin and hookworm egg count, and no consistent correlation between haemoglobin level or haematocrit and either measure of hookworm intensity. The results suggest that the role of hookworm in the aetiology of anaemia may be difficult to assess without the accurate measurement of hookworm burden.

Epidemiology and immunology of Necator americanus infection in a community in Papua New Guinea: humoral responses to excretory-secretory and cuticular collagen antigens.

Baseline data from an immuno-epidemiological study of hookworm infection in a rural village in Madang Province, Papua New Guinea are reported. Necator americanus was found to be the commonest helminth infection, with a prevalence of near 100% and intensity of 40 worms per host in adults. Enterobius vermicularis, Ascaris lumbricoides and Trichuris trichiura were also present, at prevalences of 53, 10 and 3% respectively; Ancylostoma duodenale was absent. The frequency distribution of N. americanus was highly over-dispersed, and was well described by a negative binomial distribution with aggregation parameter, k, of 0.370. Intensity of infection was significantly related to host age, but did not differ between the sexes. Haemoglobin levels and haematocrit values were indicative of anaemia in the community, but were unrelated to hookworm infection. Levels of antibodies (IgG, IgA and IgM combined) against adult Necator cuticular collagen and excretory-secretory (ES) products were determined. Serum concentrations of the two types of antibody were significantly correlated with each other. Significant positive correlations were found between anti-ES antibody levels and hookworm egg production, and between anti-collagen antibody levels and host age. It is suggested that the level of anti-collagen antibodies may reflect cumulative exposure to infection, whereas levels of anti-ES antibodies may be more dependent on current worm burden. No evidence was found to suggest that either antibody response is important in regulating parasite population growth. Similarly, the presence of a positive correlation between eosinophil concentration and infection intensity in adults indicates that eosinophilia reflects, rather than determines, the host's worm burden.