The trial profile is shown in figure . At enrolment, mothers' mean age was 23 years; 15/103 (15%) were HIV-positive; 15/103 (15%) had malaria; 38/101 (38%) had hookworm, 23/101 (23%) Schistosoma mansoni, 13/101 (13%) Trichuris trichiura, 6/101 (6%) Ascaris lumbricoides, 13/84 (15%) Strongyloides stercoralis and 21/99 (22%) Mansonella perstans. Of those with hookworm, 28 had light (1–999 eggs/g), eight moderate (1000–3999 eggs/g) and two heavy (≥ 4000 eggs/g) infection. Maternal helminth infection showed expected associations with younger age (p = 0.027) and lower education (p = 0.002). Mothers in albendazole and placebo groups were similar in age, HIV, malaria and helminth prevalence, and baseline cytokine levels and responses. After delivery the prevalence of hookworm was 0/47 (0%) in the albendazole group and 14/42 (33%) in the placebo group; other helminths remained similar in the two groups. Among one-year-olds, 3/63 had hookworm (one whose mother received albendazole, two placebo), 1/63 had Trichuris (albendazole-mother) and 1/54 Mansonella (placebo-mother).
Kato-Katz stool results for mothers at enrolment and all cytokine results were available for 90/103 mothers at enrolment, 79/93 at delivery (72 with results at both time points), and 57/63 one-year-olds. Whole blood assay data were missing when assays were unsatisfactory, and samples insufficient for repeats. Analysis of cord blood samples was restricted to 48/52 samples where mixing with maternal blood was excluded using an alkaline denaturation test [13
]; serum cytokine results were complete for these 48 samples. Whole blood assay data were complete for only 33 of these cord blood samples: only five had a positive IFN-γ response, and one a positive IL-5 to CFP; therefore cord blood whole blood assay results are not presented further here.
Effects of maternal helminth infection on cytokine responses in mothers at enrolment and in their one-year-old infants
At enrolment, all helminth species were associated with lower maternal IFN-γ responses to CFP. This effect was most marked, and statistically significant, for hookworm (median 10 (interquartile range 0, 162) pg/ml compared to 173 (4, 471) pg/ml, p = 0.048) (Figure ). Five mothers had a positive IL-5 response to CFP; all had helminth infections.
Figure 2 Effects of maternal hookworm on responses mycobacterial antigens in mothers during pregnancy and their one-year-old infants. Effects of maternal hookworm on IFN-γ responses to culture filtrate proteins of Mycobacterium tuberculosis in (a) mothers (more ...)
Among one-year-old infants, the opposite effect was seen: IFN-γ responses to CFP were higher, for all species of helminth, among infants of mothers who had helminth infections in pregnancy. Despite effective treatment of maternal hookworm in approximately half the cases, this effect was also strongest for hookworm (median IFN-γ production 737 (220, 1805) pg/ml for infants of mothers with hookworm; 75 (24, 228) pg/ml for infants of mothers without helminths, p = 0.020) (Figure ). Nine infants had positive IL-5 responses to CFP; 6 had mothers with helminth infection.
It was considered possible that the associations between maternal hookworm and responses to CFP in the mothers, or in their infants, might be explained by confounding factors such as co-infection with other pathogens (maternal malaria or HIV), maternal age, socioeconomic factors, or maternal BCG immunisation. We investigated this by creating binary variables for IFN-γ production below and above the median level of response in mothers and infants. This allowed us to examine associations between such factors and responses to CFP (Table) and to adjust for their effects using logistic regression. The observed associations between maternal hookworm and IFN-γ responses to CFP in mothers and their infants were not explained by any of the variables examined: odds ratios for the associations between maternal hookworm and high IFN-γ responses to CFP, adjusted for maternal age, HIV status, malaria parasitaemia, education, BCG scar and schistosomiasis were, for mothers at enrolment: 0.14 (95% confidence interval 0.02–0.83, p = 0.021); for infants at one year of age: 17.65 (95% CI: 1.20–258.66, p = 0.013).
Analysis of the trial: effects of maternal treatment with albendazole on cytokine responses to CFP
Mothers who received albendazole had an increase in IFN-γ response to CFP (median increase +26 (-7, +293) pg/ml), whereas placebo recipients did not (0 (-279, +73) pg/ml) (p = 0.019)). Among hookworm-infected mothers, corresponding increases were +55 (0, +386) pg/ml for albendazole, and 0 (-44, +81) pg/ml for placebo recipients (p = 0.055). Thus, treatment of hookworm in the mothers resulted in a profile of CFP responses that was similar to the "no helminth" group (Figure ). No effects of albendazole were observed on maternal IL-5 responses.
Figure 3 Effects of maternal hookworm and albendazole treatment on responses mycobacterial antigens in mothers at delivery and their one-year-old infants. Effects of maternal hookworm, and maternal treatment with albendazole during pregnancy, on IFN-γ (more ...)
Conversely, IFN-γ responses to CFP were lower in infants of hookworm-infected mothers who received albendazole than in those whose mothers received placebo (Figure ) and IL-5 responses less frequent, although these effects were not statistically significant. The BCG scar diameter in one-year-old infants followed a similar pattern, being significantly smaller in infants of hookworm-infected mothers who received albendazole (2.0 (1.0, 3.0) mm) than in those whose mothers received placebo (3.5 (2.8, 4.0) mm; p = 0.026). BCG scar diameter showed a positive correlation with the IL-5 response to CFP (ρ = 0.32, p = 0.011) but not with the IFN-γ response.
Non-specific immune responses
In most analyses, maternal helminths and albendazole treatment showed no effect on PHA responses or serum cytokine levels. However, cord blood IL-10 was lower for albendazole than placebo recipients, and this effect was statistically significant among hookworm mothers, (median 3 (0, 5) pg/ml versus 12 (4, 18) pg/ml, p = 0.031). TGF-β was detected in cord blood only for three mothers, all placebo recipients, of whom two were hookworm-infected.