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ABSTRACT |
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
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INTRODUCTION |
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TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES |
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MATERIALS AND METHODS |
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TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES |
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Study design. All subjects were admitted to hospital and treated according to standard protocols, using oral or intravenous quinine followed by a second anti-malarial agent (usually sulfadoxine/pyrimethamine). Demographic and clinical data were recorded at admission and daily thereafter onto a standard proforma. Twenty milliliters of blood was taken before any anti-malarial treatment was given for routine hematologic (full blood count) and biochemical (urea and electrolytes and liver function tests) analyses, and repeated as clinically indicated. Sera were stored from this admission sample for immunoglobulin and cytokine assays. Malaria films were taken daily, stained with Field’s stain, and examined at 1,000x magnification with the density of asexual infection expressed as a percentage.
Outcome measures. There is no generally accepted definition of what constitutes immunity to malaria, nor is there any laboratory marker that consistently predicts protection from disease.4 As a result, we made no attempt to classify these subjects as semi-immune or naïve. Rather, we divided the study population into two groups, mild versus severe disease, using modified World Health Organization criteria1 and then looked for any differences between the groups in terms of ethnicity, residence in a malaria-endemic area, previous history of malaria, or immunologic parameters that might be expected to be associated with less severe disease.
Subjects were classified as having severe disease if any of the following were present at any time: asexual parasitemia > 2%, schizonts on the blood film, bilirubin > 20 IU/L, creatinine >130 mmol/L, hemoglobin < 9 g/dL, platelet count < 50 x 109/L, or disease of sufficient severity to warrant admission to the intensive therapy unit (ITU). The decision to admit to the ITU was taken by attending physicians who were not otherwise involved in the study. Patients with severe disease had an average of 3.8 markers of severity, and their median parasitemia was 3.9%. All patients were monitored for hypoglycemia while receiving quinine but hypoglycemia per se was not taken as a marker of severe disease. Blood gas and lactate measurements were not routinely made.
Axillary temperature was measured at admission and every four hours thereafter. The fever experienced by each subject during their admission was expressed as a cumulative sum around a value of 37.0°C.8 As an example, a temperature of 39°C scored +2 and a second reading of 36.5°C scored –0.5; the final figure therefore represents the summation of each temperature reading taken during the admission. Duration of fever was taken as the number of days from the first recorded fever to the beginning of the first 24-hour period without fever (axillary temperature 
37.5°C) irrespective of acetaminophen usage.
Immunologic parameters.
Pro-inflammatory and anti-inflammatory cytokines (tumor necrosis factor-
[TNF-], interleukin-10 [IL-10], interferon-
[IFN-], and transforming growth factor-ß [TGF-ß]) and antibodies against P. falciparum schizont extracts (Pfse) were measured by an enzyme-linked immunosorbent assay (ELISA). Plasmodium falciparum schizonts obtained from Percoll purification were used for preparing an antigen to detect antibody to P. falciparum by standard capture ELISA as previously described.9 Plasma concentrations of TNF- and TGF-ß were determined by DuoSet ELISA (R&D Systems, Abingdon, United Kingdom) according to manufacturers’ instructions. Monoclonal antibody pairs (Pharmingen-BD, Oxford, United Kingdom) were used for determining the levels of IL-10 and IFN- as previously described.10 Plasma samples were tested in duplicate at 1:12 dilution for TGF-ß and at a dilution of 1:5 or 1:10 in all other cytokine assays; cytokine concentrations were then calculated per milliliter of undiluted plasma. All samples were tested in duplicate and assays were repeated at least once on different aliquots of each sample.
Statistical methods.
The frequency and percentages for categorical variables were shown and differences between the mild and severe groups were tested using the chi-square distribution. For continuous variables, means and standard deviations were shown for normally distributed data, and for non-normal data (cytokine concentrations and duration data) geometric means and ranges were presented. For the comparison of continuous data across mild and severe patients, the T-test was used (on the log transformed data if necessary). With 25 patients with severe disease and 75 patients with mild disease, the study had 80% power to detect as significant an odds ratio of 4.0 for common categorical exposures, a 70% increase in IL-10 and IFN-, a 40% increase in TNF-, and a 30% increase in TGF-ß.
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RESULTS |
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TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES |
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). Twenty-one had an asexual parasitemia >2%, 17 had schizonts on the blood film, 24 had hyperbilirubinemia, 6 had an elevated creatinine levels (4 of whom required dialysis), 7 had a hemoglobin concentration < 9 g/dL, and 13 had a platelet count < 50 x 109/L. Six required admission to the ITU, four of whom had a history of previous exposure to malaria.
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As expected, the group with severe disease had a higher maximal temperature, were febrile for longer, and required a longer hospital stay, but there were few other differences between the two groups. There was no difference between those with severe disease compared with those with mild disease in terms of ethnicity, residence in a malaria-endemic area, or previous history of malaria (as shown either by the stated history of previous malaria or by the level of IgG antibody to Pfse). These results are shown in Table 2. A higher percentage of patients with mild disease reported taking anti-malarial prophylaxis, and this difference was even greater among those who reported taking prophylaxis in the last 28 days, although not reaching statistical significance (P = 0.10). Although IgG, IgM and IFN- levels tended to be lower in severe cases than mild cases, these differences were not statistically significant. However, as expected, TNF- concentrations were significantly higher in severe cases than mild cases; there was also a tendency (although not statistically significant) for IL-10 concentrations to be higher among severe cases. Conversely, there was a significant inverse relationship between the IgG antibody concentration to Pfse and highest recorded parasitemia (correlation = –0.24, P = 0.023) and highest recorded temperature (correlation = –0.26, P = 0.016). This suggested that individuals with previous exposure to malaria (i.e., those with anti-malarial antibodies) did have some degree of anti-malarial immunity. Furthermore, IL-10 concentrations were significantly correlated with highest recorded parasitemia (n = 41, correlation = 0.30, P = 0.05) and temperature (n = 39, correlation = 0.36, P = 0.025), and TNF- concentrations were significantly associated with highest recorded parasitemia (n = 90, correlation = 0.49, P < 0.001) but not temperature (n = 85, correlation = 0.04, P = 0.75).
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DISCUSSION |
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ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES |
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It is commonly believed that immunity to malaria is rapidly lost once an individual moves away from a malaria-endemic area, leaving previously immune individuals at high risk of severe disease should they become re-infected.4 The one death in this study, that of a Nigerian woman who was re-exposed to malaria after eight years in Europe, could be seen as an example of this. However, systematic surveys of malaria in immigrants and during epidemics where malaria was reintroduced after many years of effective control showed that previously immune individuals, although they do become sick, have significantly lower parasite densities and a lower risk of severe and fatal outcomes than comparable, previously naive individuals.11 Thus, our observation that a significant proportion of visitors and recent immigrants to the United Kingdom from malaria-endemic countries develop severe malaria suggests that they have (or had) little or no pre-existing functional immunity to malaria.
There is no marker that has reliably been shown to correlate with immunity to malaria, although there are inherent resistance traits such as sickle-cell trait or other hemoglobinopathies4 that are known to be associated with protection against severe disease. Precisely what constitutes immunity to malaria is still much debated, but however it is defined, it is mediated in part by pro-inflammatory and anti-inflammatory cytokines, and there is evidence that these cytokines have a prognostic role.12 In accordance with previous findings,13–16 high serum concentrations of TNF- and IL-10 were associated with higher parasite counts and more severe symptoms. However, although anti-malarial antibody levels (which can be viewed as a marker for previous exposure to malaria)17 were inversely correlated with maximal parasitemia, there was no clear indication that antibody levels differed between those with mild disease and those with severe disease. This is consistent with the notion that malarial symptoms are in part immune-mediated and that individuals with considerable previous exposure to malaria can still have severe disease.18
The prevalence of severe disease among malaria-exposed adults in this study (26% of those who had ever lived in a malaria-endemic area and 22% of those born in sub-Saharan Africa had severe rather than mild malaria) might be considered rather high, given that most non-pregnant adults who become infected with malaria in malaria-endemic areas develop only trivial clinical disease and many people in malaria-endemic areas have parasites in the blood without developing any symptoms. However, the patients included in the present study may not be representative of adult malaria infections in general. Our patients are self-selected by virtue of the fact that they have symptoms of sufficient severity to seek medical care; it is possible that some cases of imported malaria in immune individuals go unrecognized and unreported because they have few clinical symptoms. Furthermore, African and Asian patients who come to London with malaria will, by definition, have access to sufficient resources to have traveled to Europe and, by inference, may have previously lived in more affluent areas and may not have had as much exposure to the parasite as their less-advantaged compatriots. Both of these considerations support the notion that most malaria patients, wherever they originate from, who come to hospitals in non-endemic countries are likely to have little effective anti-malarial immunity.
This study raises some important points for the management of imported P. falciparum malaria. First, malaria is not a trivial disease. Six percent of our patients required admission to the ITU and one patient died after developing cerebral malaria complicated by ARDS and renal failure. Second and more unexpectedly, severe disease was no less common among patients who might be assumed to have a degree of protective immunity than among previously malaria-naive patients. Many physicians managing cases of P. falciparum malaria in temperate areas may assume that complications are less likely to arise if the patient has lived in a malaria-endemic region, as a result of some degree of acquired immunity. Our study suggests that this assumption may not be valid.
Received May 24, 2006. Accepted for publication August 2, 2006.
Acknowledgments: We are grateful to the following clinicians for recruiting patients to this study: Kazeem Aderogba, Jim Buckley, Alex Bunn, Anna Checkley, Rod Escombe, Matthew Hamill, Damien Hatton, Damien Mack, Fabiola Martin, Elinor Moore, Baz Nadjm, Richard Stümpfle, Sonali Sudarshi, and Jenny Whetham. We thank Professor Peter Chiodini and laboratory staff in the Department of Parasitology at the Hospital for Tropical Diseases for processing blood samples and examining malaria blood films. We are also grateful to Liz King for performing the cytokine assays and to Suzanna MacDonald for assisting with these assays.
Financial support: This study was supported by the Special Trustees of the Hospital for Tropical Diseases.
* Address correspondence to Justin F. Doherty, Hospital for Tropical Diseases, Mortimer Market, Capper Street, London WC1E 6AU, United Kingdom. E-mail: tom.doherty{at}uclh.org 
Authors’ addresses: Richard M. Jennings, Margaret Armstrong, and Katie L. Flanagan, Hospital for Tropical Diseases, Mortimer Market, Capper Street, London WC1E 6AU, United Kingdom. J. Brian de Souza, Jim E. Todd, and Eleanor M. Riley, London School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT, United Kingdom. Justin F. Doherty, Hospital for Tropical Diseases, Mortimer Market, Capper Street, London WC1E 6AU, United Kingdom and London School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT, United Kingdom.
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ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
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  A. Gonzalez, J. M. Nicolas, J. Munoz, P. Castro, J. Mas, M. E. Valls, J. R. Coma, J. Aibar, and J. Gascon Severe Imported Malaria in Adults: Retrospective Study of 20 Cases Am J Trop Med Hyg, October 1, 2009; 81(4): 595 - 599. [Abstract] [Full Text] [PDF]
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