Acute Malaria in Malawian Children and Adults is Characterized by Thrombocytopenia That Normalizes in Convalescence.

Background: Plasmodium falciparum malaria has been linked with significant perturbations of the peripheral cell-mediated immune system during acute phase. Some of these changes include lower than normal platelet counts. Although the exact mechanisms that drive thrombocytopenia in P. falciparum malaria are not fully known, a number of hypotheses have been proposed. We conducted two sets of studies with one aimed at determining platelet counts in Malawian children, and the other in adults during acute P. falciparum malaria and a month post treatment. Materials and Methods: We recruited a total of 113 HIV-uninfected children with acute malaria [n=54 with uncomplicated malaria (UCM), n=30 with severe malarial anemia (SMA), n=29 presenting with cerebral malaria (CM)]. We also recruited 42 HIV-uninfected healthy controls. Out of the 113 participants with malaria, 73 (65%) [n=34 (63%) UCM, n=21 (70%) SMA and n=18 (62%) CM] were successfully followed-up one month after treatment. A 5mL peripheral blood sample was collected for platelet count using HMX Haematological Analyzer analysis both at baseline (acute malaria) and at follow-up a month later. Platelet counts were also determined in blood samples of 106 HIV-uninfected adults, 47 of whom presented with UCM and 29 with severe malaria (SM) and these counts were compared to those of 30 healthy controls. Of the malaria cases, platelet counts for 44 UCM and 21 SM were determined again during follow-up a month after treatment. Results: In both children and adults, platelet counts were significantly lower during acute disease compared to the levels in the healthy controls with the lowest levels observed in CM (children) or SM (adults). These lower than normal levels increased close to normal levels a month post treatment. Conclusion: P. falciparum malaria in Malawian children and adults was characterized by profound thrombocytopenia which recovered during convalescence.

Optimization of stimulation and staining conditions for intracellular cytokine staining (ICS) for determination of cytokine-producing T cells and monocytes.

Cell-mediated responses to immunological stimuli are often localised in inflammatory sites and involve a number of cell types. These responses can be functionally characterised at the single-cell level on the basis of the types of cytokines expressed either in whole blood or PBMCs. The ability to measure antigen-specific cell responses at the single cell level is an important tool with a wide range of potential applications ranging from studies of disease pathogenesis to the evaluation of vaccines. A number of experiments were performed in this study in order to establish the optimal conditions for in vitro stimulation of cytokine production by T cells and monocytes in whole blood samples collected from healthy adult Malawian participants and the optimal staining conditions for various cytokine producing cells. Different stimulation methods and conditions, different culture tubes and incubators and different antibody labelling conditions were assessed in order to establish optimal conditions for detecting cytokine-producing cells in whole blood samples. The use of PMA plus Ionomycin produced highest cytokine-producing T cells whereas LPS was a better stimulant for cytokine producing monocytes. Stimulation of whole blood for 5 h was optimal for cytokine detection in T cells whereas 4 h was optimal for monocytes. BFA was found to be a better Golgi blocker than Monensin and the use of 15 ml Falcon-type polypropylene tubes while stationary resulted in the detection of the highest proportion of cytokine-producing cells. T cells were found to be producers of mainly TNF-α, IFN-γ and IL-2 whereas Monocytes were mainly producing TNF-α and IL-6. Anti-CD3-PerCP (used at a ratio of 1:25), anti-CD14-APC (used at a ratio of 1:50) and anti-cytokine-PE (used at a ratio of 1:12.5) resulted in the best results. The highest cytokine production monocytes were detected when 1 X FACS Lysing solution was used at a volume of 40X that of the whole blood sample compared to the other volumes. These optimal conditions are essential in determination of proportion of cytokine-producing cells using ICS in whole blood.

The Effect of Frequent Exposure to P. falciparum, HIV-Infection and Other Co-Morbidities on Development of Severe Malaria in Malawian Adults.

AIM: Plasmodium falciparum malaria predominantly affects children residing in endemic areas. However, recently a significant number of Malawian adults, who otherwise should have attained some malaria-specific immunity, have been observed to succumb to either uncomplicated malaria (UM) or severe malaria (SM). In addition, it is still unknown whether HIV is a contributing factor to SM in Malawian non-pregnant adults. We conducted this study to determine clinical and demographic features that characterize Malawian adults presenting with UM or SM. METHODS: Study participants were recruited when they presented at Queen Elizabeth Central Hospital (QECH), Blantyre, Malawi with UM or SM and had their demographic details recorded after consenting to participate in the study. Malaria infection was confirmed by Malaria Rapid Diagnostic Test (MRDT) and malaria slide microscopy. A venous blood sample was collected before treatment for various analyses. RESULTS: Both HIV-infected and HIV-uninfected participants were at risk of presenting with either UM or SM although there were more (37%) SM cases among those who were HIV-infected compared to those who were HIV-uninfected (28%) but the difference was not significant but similar numbers of UM cases (61% for HIV-uninfected and 60% for HIV-infected). Previous visit to areas of high malaria transmission was not associated with presenting with either UM or SM. HIV/malaria co-infected participants were more likely to be found with a positive blood culture result (Diphtheriods, Stenotophomonas maltophilia, Salmonella typhimurium and Staphylococcus aureus) and were at a higher risk of dying compared to HIV-uninfected malaria patients. CONCLUSION: Although HIV-infection was associated with high mortality in those co-infected with HIV and malaria, recurrent malaria episodes and having other diseases co-existing with malaria infection were the main factors associated with the development of SM in this study cohort. Further studies need to be done to determine how the host immunity is affected in those co-infected with HIV and malaria.

Geostatistical analysis of Malawi's changing malaria transmission from 2010 to 2017.

Background: The prevalence of malaria infection in time and space provides important information on the likely sub-national epidemiology of malaria burdens and how this has changed following intervention. Model-based geostatitics (MBG) allow national malaria control programmes to leverage multiple data sources to provide predictions of malaria prevalance by district over time. These methods are used to explore the possible changes in malaria prevalance in Malawi from 2010 to 2017.  Methods: Plasmodium falciparum parasite prevalence ( PfPR) surveys undertaken in Malawi between 2000 and 2017 were assembled. A spatio-temporal geostatistical model was fitted to predict annual malaria risk for children aged 2-10 years ( PfPR 2-10) at 1×1 km spatial resolutions. Parameter estimation was carried out using the Monte Carlo maximum likelihood methods. Population-adjusted prevalence and populations at risk by district were calculated for 2010 and 2017 to inform malaria control program priority setting. Results: 2,237 surveys at 1,834 communities undertaken between 2000 and 2017 were identified, geo-coded and used within the MBG framework to predict district malaria prevalence properties for 2010 and 2017. Nationally, there was a 47.2% reduction in the mean modelled PfPR 2-10 from 29.4% (95% confidence interval (CI) 26.6 to 32.3%) in 2010 to 15.2% (95% CI 13.3 to 18.0%) in 2017. Declining prevalence was not equal across the country, 25 of 27 districts showed a significant decline ranging from a 3.3% reduction to 79% reduction. By 2017, 16% of Malawi's population still lived in areas that support PfPR 2-10 ≥ 25%. Conclusions: Malawi has made substantial progress in reducing the prevalence of malaria over the last seven years. However, Malawi remains in meso-endemic malaria transmission risk. To sustain the gains made and continue reducing the transmission further, universal control interventions need to be maintained at a national level.

HIV infection has a profound effect on hematological factors but not on electrolyte profile of Malawian adults presenting with uncomplicated malaria and severe malaria.

AIM: Although malaria and HIV infections independently affect the electrolyte and hematologic profiles, little is known of how these profiles are affected in individuals coinfected with malaria and HIV. We therefore conducted this study to investigate the electrolyte and hematologic profiles of Malawian adults presenting with either uncomplicated malaria (UM), severe malaria (SM), and those presenting with HIV and UM or HIV and SM. METHODS: Study participants were recruited at Queen Elizabeth Central Hospital, and malaria infection was confirmed by rapid diagnostic test and malaria slides, and full blood count, HIV, and wet chemistries were analyzed. RESULTS: Sodium, potassium, calcium, and chloride levels of all 4 study groups were similar to those of healthy controls. Both HIV-infected groups (UM and SM) had lower red blood cell counts and lower hemoglobin concentration than the reference range. Platelet counts were lower in both HIV-uninfected SM cases (64×109/L) and in the HIV-infected SM cases (66×109/L) compared to the reference range (115-290×109/L). HIV- UM cases had higher proportion and absolute counts of neutrophils and white blood cell counts compared to the HIV+ UM cases. CONCLUSION: HIV infection did not affect the electrolyte profile of Malawian adults presenting with UM or SM but had an effect on red blood cells, Hb concentration, neutrophils, and platelet counts.