Prothrombotic autoantibodies targeting platelet factor 4/polyanion are associated with pediatric cerebral malaria.

BACKGROUNDFeatures of consumptive coagulopathy and thromboinflammation are prominent in cerebral malaria (CM). We hypothesized that thrombogenic autoantibodies contribute to a procoagulant state in CM.METHODSPlasma from children with uncomplicated malaria (UM) (n = 124) and CM (n = 136) was analyzed by ELISA for a panel of 8 autoantibodies including anti-platelet factor 4/polyanion (anti-PF4/P), anti-phospholipid, anti-phosphatidylserine, anti-myeloperoxidase, anti-proteinase 3, anti-dsDNA, anti-ß-2-glycoprotein I, and anti-cardiolipin. Plasma samples from individuals with nonmalarial coma (NMC) (n = 49) and healthy controls (HCs) (n = 56) were assayed for comparison. Associations with clinical and immune biomarkers were determined using univariate and logistic regression analyses.RESULTSMedian anti-PF4/P and anti-PS IgG levels were elevated in individuals with malaria infection relative to levels in HCs (P < 0.001) and patients with NMC (PF4/P: P < 0.001). Anti-PF4/P IgG levels were elevated in children with CM (median = 0.27, IQR: 0.19-0.41) compared with those with UM (median = 0.19, IQR: 0.14-0.22, P < 0.0001). Anti-PS IgG levels did not differ between patients with UM and those with CM (P = 0.39). When patients with CM were stratified by malaria retinopathy (Ret) status, the levels of anti-PF4/P IgG correlated negatively with the peripheral platelet count in patients with Ret+ CM (Spearman's rho [Rs] = 0.201, P = 0.04) and associated positively with mortality (OR = 15.2, 95% CI: 1.02-275, P = 0.048). Plasma from patients with CM induced greater platelet activation in an ex vivo assay relative to plasma from patients with UM (P = 0.02), and the observed platelet activation was associated with anti-PF4/P IgG levels (Rs= 0.293, P = 0.035).CONCLUSIONSThrombosis mediated by elevated anti-PF4/P autoantibodies may be one mechanism contributing to the clinical complications of CM.

The ability of Interleukin-10 to negate haemozoin-related pro-inflammatory effects has the potential to restore impaired macrophage function associated with malaria infection.

BACKGROUND: Although pro-inflammatory cytokines are involved in the clearance of Plasmodium falciparum during the early stages of the infection, increased levels of these cytokines have been implicated in the pathogenesis of severe malaria. Amongst various parasite-derived inducers of inflammation, the malarial pigment haemozoin (Hz), which accumulates in monocytes, macrophages and other immune cells during infection, has been shown to significantly contribute to dysregulation of the normal inflammatory cascades. METHODS: The direct effect of Hz-loading on cytokine production by monocytes and the indirect effect of Hz on cytokine production by myeloid cells was investigated during acute malaria and convalescence using archived plasma samples from studies investigating P. falciparum malaria pathogenesis in Malawian subjects. Further, the possible inhibitory effect of IL-10 on Hz-loaded cells was examined, and the proportion of cytokine-producing T-cells and monocytes during acute malaria and in convalescence was characterized. RESULTS: Hz contributed towards an increase in the production of inflammatory cytokines, such as Interferon Gamma (IFN-γ), Tumor Necrosis Factor (TNF) and Interleukin 2 (IL-2) by various cells. In contrast, the cytokine IL-10 was observed to have a dose-dependent suppressive effect on the production of TNF among other cytokines. Cerebral malaria (CM) was characterized by impaired monocyte functions, which normalized in convalescence. CM was also characterized by reduced levels of IFN-γ-producing T cell subsets, and reduced expression of immune recognition receptors HLA-DR and CD 86, which also normalized in convalescence. However, CM and other clinical malaria groups were characterized by significantly higher plasma levels of pro-inflammatory cytokines than healthy controls, implicating anti-inflammatory cytokines in balancing the immune response. CONCLUSIONS: Acute CM was characterized by elevated plasma levels of pro-inflammatory cytokines and chemokines but lower proportions of cytokine-producing T-cells and monocytes that normalize during convalescence. IL-10 is also shown to have the potential to indirectly prevent excessive inflammation. Cytokine production dysregulated by the accumulation of Hz appears to impair the balance of the immune response to malaria and exacerbates pathology.

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.

Characterization of Lymphocyte Subsets in Lymph Node and Spleen Sections in Fatal Pediatric Malaria.

Secondary lymphoid tissues play a major role in the human immune response to P. falciparum infection. Previous studies have shown that acute falciparum malaria is associated with marked perturbations of the cellular immune system characterized by lowered frequency and absolute number of circulating T cell subsets. A temporary relocation of T cells, possibly by infiltration to secondary lymphoid tissue, or their permanent loss through apoptosis, are two proposed explanations for this observation. We conducted the present study to determine the phenotype of lymphocyte subsets that accumulate in the lymph node and spleen during acute stages of falciparum malaria infection in Malawian children, and to test the hypothesis that lymphocytes are relocated to lymphoid tissues during acute infection. We stained tissue sections from children who had died of the two common clinical forms of severe malaria in Malawi, namely severe malarial anemia (SMA, n = 1) and cerebral malaria (CM, n = 3), and used tissue sections from pediatric patients who had died of non-malaria sepsis (n = 2) as controls. Both lymph node and spleen tissue (red pulp) sections from CM patients had higher percentages of T cells (CD4+ and CD8+) compared to the SMA patient. In the latter, we observed a higher percentage of CD20+ B cells in the lymph nodes compared to CM patients, whereas the opposite was observed in the spleen. Both lymph node and spleen sections from CM patients had increased percentages of CD69+ and CD45RO+ cells compared to tissue sections from the SMA patient. These results support the hypothesis that the relocation of lymphocytes to spleen and lymph node may contribute to the pan-lymphopenia observed in acute CM.

Expression of CD11a, CD11b, CD11c, and CD18 on Neutrophils from Different Clinical Types of Malaria in Malawian Children.

BACKGROUND: Malaria in individuals who have never had an infection before is usually characterized by an inflammatory response that is linked to the expression of specific activation markers on cells of the innate immune system. METHODS: This study investigated absolute white blood cell (WBC) and neutrophil counts and expression of several adhesion markers on neutrophils from HIV-uninfected children who were suffering from cerebral malaria (n=35), severe malarial anemia (SMA, n=39), and uncomplicated malaria (n=49) and healthy aparasitemic children (n=33) in Blantyre, Malawi. RESULTS: All clinical malaria groups had higher WBC and neutrophil counts compared to healthy controls, with the acute SMA group having significantly (p<0.0001) higher WBC counts than the controls. These elevated counts normalized during recovery. Surprisingly, in all clinical malaria groups, the surface expression of CD11b, CD11c, and CD18 on neutrophils was lower than in healthy controls, again normalizing during convalescence. CONCLUSION: In areas where Plasmodium falciparum malaria is hyperendemic, such as where this study was conducted, neutrophils have reduced expression of adhesion molecules and activation markers during acute stages of the infection, regardless of the clinical type of malaria. This reduced expression could be due to an individual's past exposure to P. falciparum or other parasite-related factors that manifest during active malaria that still need to be investigated.

SARS-CoV-2 and HIV-1: Should HIV-1-Infected Individuals in Sub-Saharan Africa Be Considered a Priority Group for the COVID-19 Vaccines?

Since its emergence in 2019 SARS-CoV-2 has proven to have a higher level of morbidity and mortality compared to the other prevailing coronaviruses. Although initially most African countries were spared from the devastating effect of SARS-CoV-2, at present almost every country has been affected. Although no association has been established between being HIV-1-infected and being more vulnerable to contracting COVID-19, HIV-1-infected individuals have a greater risk of developing severe COVID-19 and of COVID-19 related mortality. The rapid development of the various types of COVID-19 vaccines has gone a long way in mitigating the devastating effects of the virus and has controlled its spread. However, global vaccine deployment has been uneven particularly in Africa. The emergence of SARS-CoV-2 variants, such as Beta and Delta, which seem to show some subtle resistance to the existing vaccines, suggests COVID-19 will still be a high-risk infection for years. In this review we report on the current impact of COVID-19 on HIV-1-infected individuals from an immunological perspective and attempt to make a case for prioritising COVID-19 vaccination for those living with HIV-1 in Sub-Saharan Africa (SSA) countries like Malawi as one way of minimising the impact of COVID-19 in these countries.

The role of different components of the immune system against Plasmodium falciparum malaria: Possible contribution towards malaria vaccine development.

Plasmodium falciparum malaria still remains a major global public health challenge with over 220 million new cases and well over 400,000 deaths annually. Most of the deaths occur in sub-Saharan Africa which bears 90 % of the malaria cases. Such high P. falciparum malaria-related morbidity and mortality rates pose a huge burden on the health and economic wellbeing of the countries affected. Lately, substantial gains have been made in reducing malaria morbidity and mortality through intense malaria control initiatives such as use of effective antimalarials, intensive distribution and use of insecticide-treated nets (ITNs), and implementation of massive indoor residual spraying (IRS) campaigns. However, these gains are being threatened by widespread resistance of the parasite to antimalarials, and the vector to insecticides. Over the years the use of vaccines has proven to be the most reliable, cost-effective and efficient method for controlling the burden and spread of many infectious diseases, especially in resource poor settings with limited public health infrastructure. Nonetheless, this had not been the case with malaria until the most promising malaria vaccine candidate, RTS,S/AS01, was approved for pilot implementation programme in three African countries in 2015. This was regarded as the most important breakthrough in the fight against malaria. However, RTS,S/AS01 has been found to have some limitations, the main ones being low efficacy in certain age groups, poor immunogenicity and need for almost three boosters to attain a reasonable efficacy. Thus, the search for a more robust and effective malaria vaccine still continues and a better understanding of naturally acquired immune responses to the various stages, including the transmissible stages of the parasite, could be crucial in rational vaccine design. This review therefore compiles what is currently known about the basic biology of P. falciparum and the natural malaria immune response against malaria and progress made towards vaccine development.

Acceptability of Vegetable Fortified Ugali in Sub-Saharan Africa.

Corn flour-based porridge like dough, ugali, is the staple food of low-income population in sub-Saharan Africa. Lack of vitamin A, carotenoids, and dietary fibers brings about serious health issues to this population. In this study, vegetables including bok choy, broccoli, cabbage, carrot, Chinese onion stalk (C_onion), mushroom, are added during the cooking of ugali, as nutritional supplements. The freeze-dried powder of each vegetable was used for its long storage, stable nutrients, and similar particle size. Sub-Saharan African assessors were trained and sensory evaluated the six different vegetable fortified ugali with the plain, unfortified as the control on five attributes. The plain ugali was indistinguishable with the C_onion stalk fortified in color, with the carrot and C_onion stalk fortified in odor, with all vegetables (except broccoli and mushroom) fortified ugali in taste, with carrot and C_onion stalk fortified in granularity, and with cabbage, carrot, C_onion stalk fortified in viscosity. Preference ranking analysis showed that the C_onion stalk fortified ugali is even more favorably preferred than the plain, unfortified ugali, probably due to the umami components in C_onion that serve as the taste enhancer. This study indicates that Chinese onion stalk is a potential vegetable supplement to population in the sub-Saharan Africa.

Determinants of brain swelling in pediatric and adult cerebral malaria.

Cerebral malaria (CM) affects children and adults, but brain swelling is more severe in children. To investigate features associated with brain swelling in malaria, we performed blood profiling and brain MRI in a cohort of pediatric and adult patients with CM in Rourkela, India, and compared them with an African pediatric CM cohort in Malawi. We determined that higher plasma Plasmodium falciparum histidine rich protein 2 (PfHRP2) levels and elevated var transcripts that encode for binding to endothelial protein C receptor (EPCR) were linked to CM at both sites. Machine learning models trained on the African pediatric cohort could classify brain swelling in Indian children CM cases but had weaker performance for adult classification, due to overall lower parasite var transcript levels in this age group and more severe thrombocytopenia in Rourkela adults. Subgrouping of patients with CM revealed higher parasite biomass linked to severe thrombocytopenia and higher Group A-EPCR var transcripts in mild thrombocytopenia. Overall, these findings provide evidence that higher parasite biomass and a subset of Group A-EPCR binding variants are common features in children and adult CM cases, despite age differences in brain swelling.

Variation of B cell subsets with age in healthy Malawians.

Although a number of previous studies have shown that different lymphocyte subsets, including B cells, vary with age, how different B cell subsets vary with age in Malawian population has not been shown before. We recruited Malawian participants of different ages and analyzed their venous blood samples for different B cell subsets. We found that both percentage and absolute counts of B cells varied with age peaking in the 7 to 12 months age group. Proportion of naïve B cells was highest in neonates and decreased with age whereas the percentage of memory B cells was lowest in neonates and increased with age. When we zeroed in on the age band within which the proportion of B cells was highest, both classical and activated memory B cells increased with age and the naïve followed the opposite trend. These results provide additional knowledge in our understanding of the dynamics of B cell subsets in individuals of a specific ethnicity as they age.

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.

Characterization of DNA methylation in Malawian Mycobacterium tuberculosis clinical isolates.

BACKGROUND: Although Mycobacterium tuberculosis (Mtb) strains exhibit genomic homology of >99%, there is considerable variation in the phenotype. The underlying mechanisms of phenotypic heterogeneity in Mtb are not well understood but epigenetic variation is thought to contribute. At present the methylome of Mtb has not been completely characterized. METHODS: We completed methylomes of 18 Mycobacterium tuberculosis (Mtb) clinical isolates from Malawi representing the largest number of Mtb genomes to be completed in a single study using Single Molecule Real Time (SMRT) sequencing to date. RESULTS: We replicate and confirm four methylation disrupting mutations in 4 lineages of Mtb. For the first time we report complete loss of methylation courtesy of C758T (S253L) mutation in the MamB gene of Indo-oceanic lineage of Mtb. Additionally, we report a novel missense mutation G454A (G152S) in the MamA gene of the Euro-American lineage which could potentially be attributed to total disruption of methylation in the CCCAG motif but partial loss in a partner motif. Through a genomic and methylome comparative analysis with a global sample of sixteen, we report previously unknown mutations affecting the pks15/1 locus in L6 isolates. We confirm that methylation in Mtb is lineage specific although some unresolved issues still remain.

Plasma cell-free DNA predicts pediatric cerebral malaria severity.

BACKGROUNDPrediction of adverse outcomes in cerebral malaria (CM) is difficult. We hypothesized that cell-free DNA (cfDNA) levels would facilitate identification of severe and potentially fatal CM cases.METHODSIn this retrospective study, plasma from Malawian children with CM (n = 134), uncomplicated malaria (UM, n = 77), and healthy controls (HC, n = 60) was assayed for cfDNA using a fluorescence assay. Host and parasite cfDNA was measured by quantitative PCR. Immune markers were determined by ELISA, Luminex, or cytometric bead array.RESULTSTotal cfDNA increased with malaria severity (HC versus UM, P < 0.001; HC versus CM, P < 0.0001; UM versus CM, P < 0.0001), was elevated in retinopathy-positive (Ret+) CM relative to Ret- CM (7.66 versus 5.47 ng/µL, P = 0.027), and differentiated Ret+ fatal cases from survivors (AUC 0.779; P < 0.001). cfDNA levels in patients with non-malarial febrile illness (NMF, P = 0.25) and non-malarial coma (NMC, P = 0.99) were comparable with UM. Host DNA, rather than parasite DNA, was the major cfDNA contributor (UM, 268 versus 67 pg/µL; CM, 2824 versus 463 pg/µL). Host and parasite cfDNA distinguished CM by retinopathy (host, AUC 0.715, P = 0.0001; parasite, AUC 0.745, P = 0.0001), but only host cfDNA distinguished fatal cases (AUC 0.715, P = 0.0001). Total cfDNA correlated with neutrophil markers IL-8 (rs = 0.433, P < 0.0001) and myeloperoxidase (rs = 0.683, P < 0.0001).CONCLUSIONQuantifying plasma cfDNA is a simple assay useful in identifying children at risk for fatal outcome and has promise as a point-of-care assay. Elevated cfDNA suggests a link with host inflammatory pathways in fatal CM.FUNDINGNIH NCATS (AK), Burroughs-Wellcome (AK), and National Health and Medical Research Council of Australia (SJR).

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.

Performance of molecular methods for the detection of Salmonella in human stool specimens.

Background: The relationship between asymptomatic Salmonella exposure within the gastrointestinal tract and Salmonella bacteraemia is poorly understood, in part due to the low sensitivity of stool culture, and the lack of validated molecular diagnostic tests for the detection of Salmonella in stool. The study aimed to determine a reliable molecular diagnostic test for Salmonella in stool specimens. Methods: We optimized an in-house monoplex real time polymerase chain reaction (PCR) for the detection of Salmonella TTR and InvA genes in stool by including a selenite broth pre-culture step for Salmonella before DNA extraction, and validated their specificity against other local common pathogens. Then we assessed their performance  against a well-validated multiplex PCR targeting the same TTR and InvA genes, and against stool culture using clinical stool specimens collected from a cohort of 50 asymptomatic healthy Malawian children that were sampled at 1-month intervals over a period of 12 months. We employed a latent Markov model to estimate the specificities and sensitivities of PCR methods. Results: TTR and InvA primers were both able to detect all the different Salmonella serovars tested, and had superior limits of detection if DNA was extracted after selenite pre-culture. TTR sensitivity and specificity for monoplex-PCR were (99.53%, 95.46%) and for multiplex-PCR (90.30%, 99.30%) respectively. InvA specificity and specificity for using monoplex-PCR was (95.06%, 90.31%) and multiplex-PCRs (89.41%, 98.00%) respectively. Sensitivity and specificity for standard stool culture were 62.88% and 99.99% respectively. Culture showed the highest PPV (99.73%) and mono-TTR had the highest NPV (99.67%). Conclusion: Test methods demonstrated high concordance although stool culture and monoplexed TTR primers had superior specificity and sensitivity respectively. The use of selenite pre-enrichment step increased Salmonella detection rate. Taken together, molecular detection methods used here could be used to reveal the true extent of both asymptomatic and symptomatic Salmonella exposure events.

Variation in competent and respectful delivery care in Kenya and Malawi: a retrospective analysis of national facility surveys.

OBJECTIVE: Although substantial progress has been made in increasing access to care during childbirth, reductions in maternal and neonatal mortality have been slower. Poor-quality care may be to blame. In this study, we measure the quality of labour and delivery services in Kenya and Malawi using data from observations of deliveries and explore factors associated with levels of competent and respectful care. METHODS: We used data from nationally representative health facility assessment surveys. A total of 1100 deliveries in 392 facilities across Kenya and Malawi were observed and quality was assessed using two indices: the quality of the process of intrapartum and immediate postpartum care (QoPIIPC) index and a previously validated index of respectful maternity care. Data from standardised observations of care were analysed using descriptive statistics and multivariable random-intercept regression models to examine factors associated with variation in quality of care. We also quantified the variance in quality explained by each domain of covariates (patient-, provider- and facility-level and subnational divisions). RESULTS: Only 61-66% of basic elements of competent and respectful care were performed. In adjusted models, better-staffed facilities, private hospitals and morning deliveries were associated with higher levels of competent and respectful care. In Malawi, younger, primipara and HIV-positive women received higher-quality care. Quality also differed substantially across regions in Kenya, with a 25 percentage-point gap between Nairobi and the Coast region. Quality was also higher in higher-volume facilities and those with caesarean section capacity. Most of the explained variance in quality was due to regions in Kenya and to facility, and patient-level characteristics in Malawi. CONCLUSIONS: Our findings suggest considerable scope for improvement in quality. Increasing staffing and shifting births to higher-volume facilities - along with promotion of respectful care in these facilities - should be considered in sub-Saharan Africa to improve outcomes for mothers and newborns.

OBJECTIF: Bien que des progrès substantiels aient été accomplis dans l'amélioration de l'accès aux soins pendant l'accouchement, les réductions de la mortalité maternelle et néonatale ont été plus lentes. Des soins de mauvaise qualité peuvent être à blâmer. Dans cette étude, nous mesurons la qualité de la main-d'œuvre et des services d'accouchement au Kenya et au Malawi en utilisant les données des observations des accouchements et explorons les facteurs associés aux niveaux de la compétence et du respect dans les soins. MÉTHODES: Nous avons utilisé les données d'enquêtes d'évaluation des établissements de santé représentatives au niveau national. 1100 accouchements dans 392 établissements au Kenya et au Malawi ont été observés et la qualité a été évaluée à l'aide de deux indices: l'indice de qualité du processus de soins intra-partum et postpartum immédiat (QoPIIPC) et un indice précédemment validé de soins maternels respectueux. Les données des observations normalisées des soins ont été analysées à l'aide de statistiques descriptives et de modèles de régression à interceptions aléatoires multivariables pour examiner les facteurs associés à la variation de la qualité des soins. Nous avons également quantifié la variance de la qualité expliquée par chaque domaine de covariables (divisions au niveau des patients, des prestataires et des établissements, et infranationales). RÉSULTATS: Seuls 61% à 66% des éléments de base de soins compétents et respectueux ont été réalisés. Dans les modèles ajustés, des établissements mieux dotés en personnel, des hôpitaux privés et des accouchements le matin étaient associés à des niveaux plus élevés de soins compétents et respectueux. Au Malawi, les femmes plus jeunes, primipares et VIH positives ont reçu des soins de meilleure qualité. La qualité différait également considérablement d'une région à l'autre au Kenya, avec un écart de 25 points de pourcentage entre Nairobi et la région côtière. La qualité était également plus élevée dans les établissements avec un volume plus élevé et ceux ayant une capacité de césarienne. La majeure partie des raisons de la variance dans la qualité était liée aux régions du Kenya et à l'établissement et aux caractéristiques des patients au Malawi. CONCLUSIONS: Nos résultats suggèrent une marge considérable pour l'amélioration de la qualité. L'augmentation du personnel et le déplacement des naissances vers des établissements de plus grand volume - ainsi que la promotion de soins respectueux dans ces établissements - devraient être envisagés en Afrique subsaharienne pour améliorer les résultats pour les mères et les nouveau-nés.

Genetic diversity of Mycobacterium tuberculosis clinical isolates in Blantyre, Malawi.

Despite the high burden of tuberculosis (TB) worldwide, specific factors influencing disease transmission remain elusive. Long term epidemiological studies and in vitro experimental models provide evidence of variable relative fitness of Mycobacterium tuberculosis (Mtb) strains but few such studies are available. Large sequence polymorphisms (LSP) are a robust molecular marker and are feasible as an epidemiological investigative tool. Few Mtb molecular epidemiological studies have been reported in Malawi owing to lack of laboratories with molecular tools. We characterized the genetic diversity of Mtb clinical isolates amongst TB patients in Blantyre, Malawi. We genotyped 64 Mtb clinical isolates using LSP-PCR, assigned specific lineages and confirmed 18 of the isolates using SMRT sequencing. The 64 isolates clustered into 4 lineages (L1-L4) with L4 predominating. There were 10/64 (16%) isolates belonging to L1, 6/64 (9%) belonging to L2, 2/64 (3%) belonging to L3 and 46/64 (72%) belonging to L4. Comparison with a previous study done in Karonga revealed concordance in L1 and L4 but discodance in L2 and L3. The phylogenetic tree constructed, comprised of 3/4 lineages present in Blantyre with 3/18 belonging to L1, 3/18 belonging to L2 and 12/18 belonging to L4. Four Mtb lineages were present in Blantyre with L4 predominating. Larger studies are needed to understand the molecular epidemiology of TB in Blantyre in light of increased bi-directional migration with South Africa.

Malawian children with uncomplicated and cerebral malaria have decreased activated Vγ9Vδ2 γδ T cells which increase in convalescence.

Malaria is responsible for almost half a million deaths annually. The role of Vγ9Vδ2 γδ T cells in malaria is still unclear. Studies have reported an association between this cell subset and malaria symptoms and severity. Profiles of Vγ9Vδ2 γδ T cells in bigger cohorts with different levels of clinical severity have not been described. Proportion, numbers, and activation status of Vγ9Vδ2 γδ T cells were measured by flow cytometry in 59 healthy controls (HCs), 58 children with uncomplicated malaria (UM) and 67 with cerebral malaria (CM,) during acute malaria and in convalescence 28 days later. Vγ9Vδ2 γδ T cell were lower in children presenting with UM and CM than in HCs. Cell counts did not vary with malaria severity (CM median counts 40 x 103 cells/µL, IQR [23-103]; UM median counts 30 x 103 cells/µL [10-90], P = 0.224). Vγ9Vδ2 γδ T cell counts increased during convalescence for UM (70 [40-60] x 103 cells/µL and CM (90 [60-140] x 103 cells/µL), to levels similar to those in HCs (70 [50-140] x 103 cells/µL), p = 0.70 and p = 0.40 respectively. Expression of the activation markers CD69 and HLA-DR on Vγ9Vδ2 γδ T cells was higher in malaria cases than in controls (HCs vs UM or CM, p < 0.0001) but was similar between UM and CM. HLA-DR expression remained elevated at 28 days, suggesting sustained activation of Vγ9Vδ2 γδ T cells during recovery. Vγ9Vδ2 γδ T cell proportions and cells counts were suppressed in acute disease and normalized in convalescence, a phenomenon previously hypothesized to be due to transient migration of the cells to secondary lymphoid tissue. The presence of highly activated Vγ9Vδ2 γδ T cells suggests that this T cell subset plays a specific role in response to malaria infection.

Distinct Biomarker Profiles Distinguish Malawian Children with Malarial and Non-malarial Sepsis.

Presently, it is difficult to accurately diagnose sepsis, a common cause of childhood death in sub-Saharan Africa, in malaria-endemic areas, given the clinical and pathophysiological overlap between malarial and non-malarial sepsis. Host biomarkers can distinguish sepsis from uncomplicated fever, but are often abnormal in malaria in the absence of sepsis. To identify biomarkers that predict sepsis in a malaria-endemic setting, we retrospectively analyzed data and sera from a case-control study of febrile Malawian children (aged 6-60 months) with and without malaria who presented to a community health center in Blantyre (January-August 2016). We characterized biomarkers for 29 children with uncomplicated malaria without sepsis, 25 without malaria or sepsis, 17 with malaria and sepsis, and 16 without malaria but with sepsis. Sepsis was defined using systemic inflammatory response criteria; biomarkers (interleukin-6 [IL-6], tumor necrosis factor receptor-1, interleukin-1 ß [IL-1ß], interleukin-10 [IL-10], von Willebrand factor antigen-2, intercellular adhesion molecule-1, and angiopoietin-2 [Ang-2]) were measured with multiplex magnetic bead assays. IL-6, IL-1ß, and IL-10 were elevated, and Ang-2 was decreased in children with malaria compared with non-malarial fever. Children with non-malarial sepsis had greatly increased IL-1ß compared with the other subgroups. IL-1ß best predicted sepsis, with an area under the receiver operating characteristic (AUROC) of 0.71 (95% CI: 0.57-0.85); a combined biomarker-clinical characteristics model improved prediction (AUROC of 0.77, 95% CI: 0.67-0.85). We identified a distinct biomarker profile for non-malarial sepsis and developed a sepsis prediction model. Additional clinical and biological data are necessary to further explore sepsis pathophysiology in malaria-endemic regions.