Francis K. Nkrumah

Low plasma concentrations of interleukin 10 in severe malarial anaemia compared with cerebral and uncomplicated malaria

BACKGROUND Severe anaemia is a major complication of malaria but little is known about its pathogenesis. Experimental models have implicated tumour necrosis factor (TNF) in induction of bone-marrow suppression and eythrophagocytosis. Conversely, interleukin 10 (IL-10), which mediates feed-back regulation of TNF, stimulates bone-marrow function in vitro and counteracts anaemia in mice. We investigated the associations of these cytokines with malarial anaemia. METHODS We enrolled 175 African children with malaria into two studies in 1995 and 1996. In the first study, children were classified as having severe anaemia (n=10), uncomplicated malaria (n=26), or cerebral anaemia (n=41). In the second study, patients were classified as having cerebral malaria (n=33) or being fully conscious (n=65), and the two groups were subdivided by measured haemoglobin as normal (>110 g/L), moderate anaemia (60-90 g/L), and severe anaemia (<50 g/L). IL-10 and TNF concentrations were measured by ELISA in plasma samples from all patients. FINDINGS IL-10 concentrations were significantly lower in patients with severe anaemia than in all other groups. In 1995, geometric mean plasma IL-10 in patients with severe anaemia was 270 pg/mL (95% CI 152-482) compared with 725 pg/mL (465-1129) in uncomplicated malaria and 966 pg/mL (612-1526) in cerebral malaria (p<0.03). In 1996, fully conscious patients with severe anaemia also had significantly lower IL-10 concentrations than all other groups, including cerebral-malaria patients with severe anaemia and all patients with moderate anaemia (p<0.001). In both studies, TNF concentrations were significantly higher in cerebral malaria than in fully conscious patients (p<0.01). By contrast, the ratio of TNF to IL-10 was significantly higher in fully conscious patients with severe anaemia than in all other groups (p<0.001). INTERPRETATION Our findings identify severe malarial anaemia as a distinct disorder in which insufficient IL-10 response to high TNF concentrations may have a central role.

Reversible suppression of bone marrow response to erythropoietin in plasmodium falciparum malaria.

To study the importance of bone marrow inhibition in the pathogenesis of malarial anaemia, haematological and parasitological parameters were followed in patients with acute malaria. Three patient categories were studied, severe malarial anaemia (SA), cerebral malaria (CM) and uncomplicated malaria (UM). Red cell distribution width (RDW) was used as a surrogate marker of release of young erythrocytes and reticulocytes. Initially RDW was low in all patients in spite of markedly increased concentrations of erythropoietin (EPO). 3 d after institution of treatment and coinciding with parasite clearance RDW increased dramatically, reaching the highest levels 1–2 weeks later. Although severe anaemia was corrected by blood transfusion during the first 3 d of treatment, the peak RDW correlated significantly with the initial EPO levels. This suggests that Plasmodium falciparum infection causes a rapidly reversible suppression of the bone marrow response to EPO. Furthermore, the inhibition of bone marrow response was a general finding irrespective of initial haemoglobin levels suggesting that the severity of anaemia depends upon the degree of peripheral erythrocyte destruction in patients with suppressed bone marrow response to EPO.

Malaria transmission dynamics at a site in northern Ghana proposed for testing malaria vaccines.

We studied the malaria transmission dynamics in Kassena Nankana district (KND), a site in northern Ghana proposed for testing malaria vaccines. Intensive mosquito sampling for 1 year using human landing catches in three micro‐ecological sites (irrigated, lowland and rocky highland) yielded 18 228 mosquitoes. Anopheles gambiae s.l. and Anopheles funestus constituted 94.3% of the total collection with 76.8% captured from the irrigated communities. Other species collected but in relatively few numbers were Anopheles pharoensis (5.4%) and Anopheles rufipes (0.3%). Molecular analysis of 728 An. gambiae.s.l. identified Anopheles gambiae s.s. as the most dominant sibling species (97.7%) of the An. gambiae complex from the three ecological sites. Biting rates of the vectors (36.7 bites per man per night) were significantly higher (P < 0.05) in the irrigated area than in the non‐irrigated lowland (5.2) and rocky highlands (5.9). Plasmodium falciparum sporozoite rates of 7.2% (295/4075) and 7.1% (269/3773) were estimated for An. gambiae s.s. and An. funestus, respectively. Transmission was highly seasonal, and the heaviest transmission occurred from June to October. The intensity of transmission was higher for people in the irrigated communities than the non‐irrigated ones. An overall annual entomological inoculation rate (EIR) of 418 infective bites was estimated in KND. There were micro‐ecological variations in the EIRs, with values of 228 infective bites in the rocky highlands, 360 in the lowlands and 630 in the irrigated area. Approximately 60% of malaria transmission in KND occurred indoors during the second half of the night, peaking at daybreak between 04.00 and 06.00 hours. Vaccine trials could be conducted in this district, with timing dependent on the seasonal patterns and intensity of transmission taking into consideration the micro‐geographical differences and vaccine trial objectives.

Antibodies to Variant Antigens on the Surfaces of Infected Erythrocytes Are Associated with Protection from Malaria in Ghanaian Children

ABSTRACT Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) is a variant antigen expressed on the surface of infected erythrocytes. Each parasite genome contains about 40 PfEMP1 genes, but only 1 PfEMP1 gene is expressed at a given time. PfEMP1 serves as a parasite-sequestering ligand to endothelial cells and enables the parasites to avoid splenic passage. PfEMP1 antibodies may protect from disease by inhibiting sequestration, thus facilitating the destruction of infected erythrocytes in the spleen. In this study, we have measured antibodies in Ghanaian children to a conserved region of PfEMP1 by enzyme-linked immunosorbent assay and antibodies to variant molecules on erythrocytes infected with field isolates of P. falciparum by flow cytometry. Based on close clinical monitoring, the children were grouped into those who did (susceptible) and those who did not (protected) have malaria during the season. The prevalences of antibodies to both the conserved PfEMP1 peptide and the variant epitopes were greater than 50%, and the levels of immunoglobulin G (IgG) correlated with age. The levels of antibodies to both the conserved peptide and the variant epitopes were higher in protected than in susceptible children. After correcting for the effect of age, the levels of IgG to variant antigens on a Sudanese and a Ghanaian parasite isolate remained significantly higher in protected than in susceptible children. Thus, the levels of IgG to variant antigens expressed on the surface of infected erythrocytes correlated with protection from clinical malaria. In contrast, the levels of IgG to a peptide derived from a conserved part of PfEMP1 did not correlate with protection from malaria.

Naturally Acquired Antibodies to the Glutamate-Rich Protein Are Associated with Protection against Plasmodium falciparum Malaria

The development of effective malaria vaccines depends on the identification of targets of well-defined protective immune responses. Data and samples from a longitudinal study of a cohort of children from coastal Ghana were used to investigate the role of antibody responses to 3 regions of the Plasmodium falciparum glutamate-rich protein (GLURP). The data show that levels of the GLURP-specific IgG that occurs in the nonrepeat region of the antigen are significantly correlated with clinical protection from P. falciparum malaria, after correction for the confounding effect of age. Furthermore, levels of cytophilic antibodies were found to be of particular importance for protection, lending support to the hypothesis that antibody-dependent cellular inhibition is the important element in GLURP-specific protective immunity.

Lack of association between maternal antibody and protection of African infants from malaria infection.

ABSTRACT Maternally derived antibodies are believed to protect infants against infection, but there is little direct evidence for a protective role of passively acquired antibodies against malaria. A longitudinal study of malaria infection in 143 infants was conducted in a region of southern Ghana where Plasmodium falciparum is endemic. Infants born in the high-transmission season were less likely to become infected in the first 20 weeks of life than children born in the low-transmission season. Plasma, obtained at birth, was tested for immunoglobulin G (IgG) and IgG subclasses to P. falciparumschizonts and recombinant circumsporozoite antigen, MSP-119, MSP-2, AMA-1, and Pf155 (also called ring-infected erytrocyte surface antigen). Antibody levels at birth were not associated with resistance to malaria infection. On the contrary, antibodies at birth were positively associated with infection, indicating that high levels of maternally derived antibodies represent a marker for intensity of exposure to malaria infection in infants. However, all five children who experienced high-density infections (>100 parasites/μl of blood) were seronegative for MSP-119 at the time of infection.

Lymphatic filariasis on the coast of Ghana.

Parasitological, clinical and entomological surveys for lymphatic filariasis were carried out in 6 villages and 3 towns on the coast of Ghana. Few or no filarial infections were observed in the towns or in the villages east of Accra. However, Wuchereria bancrofti microfilaraemia was common in the 4 western villages, with overall prevalences of 9.2%-25.4% and overall microfilariae (mf) geometric mean intensities of 321-1172 mf/mL of blood. In the same villages, hydrocele affected 8.5%-27.9% of adult males (aged > or = 20 years), and 5.6%-6.6% of adult individuals had elephantiasis (mainly of the legs). In general, the patterns of filarial infection and disease in the endemic villages resembled those observed in endemic villages in the coastal part of East Africa, with the exception that in the Ghanaian focus more females than males were affected by elephantiasis. Entomological surveys revealed that Anopheles gambiae s.l. and A. funestus were vectors of filariasis in the endemic villages. Only negligible prevalences of microfilaraemia were observed in town communities located close to highly endemic villages. Control of filariasis in this area is difficult with presently available measures, and new control tools, especially development of new drug regimens for mass treatment, are greatly needed.

Seasonal profiles of malaria infection, anaemia, and bednet use among age groups and communities in northern Ghana

Summary We conducted all‐age point prevalence surveys to profile the severity and seasonality of malaria and anaemia in Kassena‐Nankana District of northern Ghana. Random cross‐sectional surveys were timed to coincide with the end of low (May 2001) and high (November 2001) malaria transmission seasons and to yield information as to the potential value of haemoglobin (Hb) levels and parasitaemia as markers of malaria morbidity and/or malaria vaccine effect. Parasitaemia was found in 22% (515 of 2286) screened in May (dry‐low transmission), and in 61% of the general population (1026 of 1676) screened in November (wet‐high transmission). Malaria prevalence in May ranged from 4% (infants <6 months and adults 50–60 years) to 54% (children 5–10 years). Age‐specific malaria prevalence in November ranged from 38% (adults 50–60 years) to 82% (children 5–10 years). Differences between low‐ and high‐transmission periods in the prevalence of severe anaemia (SA) among young children (6–24 months) were unexpectedly comparable (low, 3.9%vs. high, 5.4%; P = 0.52) and greatly reduced from levels measured in this same community and age group in November 2000 (12.5%) and November 1996 (22.0%). Despite the lower frequency of anaemia/SA in young children surveyed in 2001, it was still clear that this condition was strongly associated with parasitaemia and that children under 5 years of age experienced a significant drop in their mean Hb levels by the end of the high transmission season. Prevalence of parasitaemia was significantly lower (P < 0.01) among infants and young children (<2 years) whose parents reported the use of bednets. There was a significantly lower risk of parasitaemia among infants [odds ratio (OR) 6–8] and young children (OR 3–4) living in the central, more urbanized sector of the study area.

Anaemia caused by asymptomatic Plasmodium falciparum infection in semi-immune African schoolchildren

A cohort of 250 Ghanaian schoolchildren aged 5-15 years was followed clinically and parasitologically for 4 months in 1997/98 in order to study the effect of asymptomatic Plasmodium falciparum infections on haematological indices and bone-marrow responses. Of the 250 children 65 met the predefined study criteria. Thus, 14 children were parasite-free throughout (group 1), 44 had P. falciparum in all blood samples collected but no symptoms of malaria (group 2), and 7 had 1 malaria attack during the study period (group 3). At the end of the study the mean haemoglobin (Hb) level in group 1 was 123 g/L, significantly higher than the value of 114 g/L in groups 2 and 3 (P < 0.02, adjusted for age and splenomegaly). The low Hb in group 2 was associated with subnormal plasma iron. Low Hb was associated with elevated erythropoietin (EPO) levels, and there was a positive correlation between EPO and reticulocyte counts. However, the reticulocyte response to EPO was more pronounced in uninfected than in infected children, suggesting a partial interference with erythropoiesis in asymptomatic infections. Children with asymptomatic infections had significantly higher plasma levels of tumour necrosis factor than uninfected children (geometric means 50 ng/L and 27 ng/L, respectively, P < 0.001) and this cytokine may contribute to bone-marrow suppression and disturbed iron metabolism. We suggest that asymptomatic malaria leads to a homeostatic imbalance in which erythrocyte loss due to parasite replication is only partially compensated for by increased erythropoiesis. The consequences of the reduced Hb levels on the development and cognitive abilities of children with asymptomatic infections, and the risk of precipitation of iron deficiency, deserve further study and should be considered in malaria control programmes that aim at reducing morbidity rather than transmission.

Burkitt's lymphoma. A clinical study of 110 patients

One hundred and ten previously untreated patients with Burkitts lymphoma were studied prospectively over a period ranging from over 1 year to 5 years. Of 103 patients who were treated with cyclophosphamide as a single agent, 79 (77%) achieved complete remission. Vincristine plus methotrexate or cytosine arabinoside induced complete remissions in only two of 24 patients who failed to respond to cyclophosphamide. Fifty‐two percent of patients who entered complete remission subsequently relapsed with tumor. Relapse was significantly higher in patients who presented with disseminated disease (Stage III‐IV) than in patients with localized disease (Stage I‐II). Patients who relapsed early (remission duration < 12 weeks) had a significantly worse prognosis than patients who relapsed late (remission duration > 12 weeks). Actuarial calculated 2‐ and 4‐year survival for all patients was 44% and 38%, respectively. Factors that adversely affected survival were primary resistance to cyclophosphamide, early tumor relapse, central nervous system disease, and involvement of abdominal organs.