Changes in protein levels as markers of severe disease: an investigation of severe malaria

Compounds directly involved in the pathogenesis of cerebral malaria (CM) remain unclear due to lack of robust methods of identifying and quantifying proteins expressed in low abundance. New developments in proteomics have now made it possible to identify low abundant proteins and provided new tools for studying host-parasite interactions. With these new tools, it may be possible to identify proteomic signatures for patients with various complications associated with severe malaria. A global proteomic strategy was used to identify differentially expressed proteins in archived plasma and CSF drawn from children diagnosed with cerebral malaria (CM) compared to those with acute bacterial meningitis (ABM) and slide negative encephalopathy (EN). Samples were first separated using two-dimensional gel electrophoresis (2-DE) or two-dimensional liquid chromatography (2D-LC) and analysed using mass spectrometry. The data collected was analyzed using various bio-informatics tools. Finally, a CM mass profile was created using MALDI-ToF mass spectrometry. Averages of about 150 spots per gel were resolved from CSF from CM and EN patients and 80 spots from ABM patients. In the gels from the CM and EN groups, 45 human proteins were found whilst 20 human proteins were unique to ABM compared to CM. For CSF, a total of 202 human proteins were identified using the 2D-LC system. Of these 13 were unique to CM, 124 to ABM and 32 to EN. 6 proteins were found in both CM and ABM and 18 were found in EN and ABM. 9 proteins were common to all 3 disease groups. A total of 66 P. falciparum proteins were identified but of these 48 were hypothetical proteins. Of the non-hypothetical proteins, 2 were found in both CM and ABM and the rest were found only in ABM. Results show that proteomics can be used to create protein profiles of different disease groups. Majority of the human proteins identified by 2-DE were found to be high abundant proteins found in CSF and plasma. The use of 2D-LC enabled the identification of more low abundant proteins but some of the P. falciparum proteins identified by 2-DE were not seen in the 2D-LC method. Majority of the human proteins found were acute phase response plasma proteins including common circulating proteins such as albumin and apolipoproteins, blood transporters and binding proteins, protease inhibitors, enzymes, cytokines and hormones, and channel and receptor-derived proteins. There seems to be a correlation between the number of proteins found in the CSF and the level of blood brain barrier break down

Global proteomic analysis of plasma from mice infected with Plasmodium berghei ANKA using two dimensional gel electrophoresis and matrix assisted laser desorption ionization-time of flight mass spectrometry

Malaria-infection in mice results in a wide perturbation of the host serum proteome involving a range of proteins and functions. Of particular interest is the increased secretion of anti-inflammatory and anti apoptotic proteins

Discovery and Validation of Biomarkers to Guide Clinical Management of Pneumonia in African Children

Background Pneumonia is the leading cause of death in children globally. Clinical algorithms remain suboptimal for distinguishing severe pneumonia from other causes of respiratory distress such as malaria or distinguishing bacterial pneumonia and pneumonia from others causes, such as viruses. Molecular tools could improve diagnosis and management. Methods We conducted a mass spectrometry–based proteomic study to identify and validate markers of severity in 390 Gambian children with pneumonia (n = 204) and age-, sex-, and neighborhood-matched controls (n = 186). Independent validation was conducted in 293 Kenyan children with respiratory distress (238 with pneumonia, 41 with Plasmodium falciparum malaria, and 14 with both). Predictive value was estimated by the area under the receiver operating characteristic curve (AUC). Results Lipocalin 2 (Lpc-2) was the best protein biomarker of severe pneumonia (AUC, 0.71 [95% confidence interval, .64–.79]) and highly predictive of bacteremia (78% [64%–92%]), pneumococcal bacteremia (84% [71%–98%]), and “probable bacterial etiology” (91% [84%–98%]). These results were validated in Kenyan children with severe malaria and respiratory distress who also met the World Health Organization definition of pneumonia. The combination of Lpc-2 and haptoglobin distinguished bacterial versus malaria origin of respiratory distress with high sensitivity and specificity in Gambian children (AUC, 99% [95% confidence interval, 99%–100%]) and Kenyan children (82% [74%–91%]). Conclusions Lpc-2 and haptoglobin can help discriminate the etiology of clinically defined pneumonia and could be used to improve clinical management. These biomarkers should be further evaluated in prospective clinical studies

Assessing Gender Identity and Sex in Diverse Low-, Middle-, and High-Income Settings: Findings from a WHO/HRP Consultative Process

Obtaining detailed data on gender identity and sex in population-based sexual health studies is important. We convened a group to develop consensus survey items. We identified 2 items to capture data on gender identity and sex that can be used in diverse settings

Strengthening capacity for community and public engagement (CPE): a mixed-methods evaluation of the ‘DELTAS Africa CPE seed fund’ pilot

Background: The ‘DELTAS Africa CPE seed fund’ was a pilot scheme designed to strengthen capacity in community and public engagement (CPE) via a ‘learn by doing’ approach. The scheme supported a total of 25 early career researchers and research support staff belonging to the DELTAS Africa network to design and implement a variety of CPE projects between August 2019 and February 2021. We examine recipient experiences of the DELTAS Africa CPE seed fund initiative, changes in their CPE attitudes, knowledge and proficiency and their CPE practice and/or practice intentions post-award. Methods: A mixed-methods process and performance evaluation drawing on three data sources: An anonymous, online knowledge, attitude and practice survey completed by CPE seed fund awardees pre- and post-project implementation (N=23); semi-structured interviews completed with a sub-sample of awardees and programme implementors (N=9); and ‘end-of-project’ reports completed by all seed fund awardees (N=25). Results: All awardees described their seed fund experience in positive terms, despite invariably finding it more challenging than originally anticipated. The combined survey, interview and end of project report data all uniformly revealed improvement in awardees’ self-reported CPE knowledge, attitudes and proficiency by completion of their respective projects. Commitment to continued CPE activity post award was evident in the survey data and all interviewees were adamant that they would integrate CPE within their respective research work going forward. Conclusion: The DELTAS Africa CPE seed fund appeared to work successfully as a CPE capacity strengthening platform and as a vehicle for fostering longer-term interest in CPE activities

The value proposition of coordinated population cohorts across Africa

Building longitudinal population cohorts in Africa for coordinated research and surveillance can influence the setting of national health priorities, lead to the introduction of appropriate interventions, and provide evidence for targeted treatment, leading to better health across the continent. However, compared to cohorts from the global north, longitudinal continental African population cohorts remain scarce, are relatively small in size, and lack data complexity. As infections and noncommunicable diseases disproportionately affect Africa's approximately 1.4 billion inhabitants, African cohorts present a unique opportunity for research and surveillance. High genetic diversity in African populations and multiomic research studies, together with detailed phenotyping and clinical profiling, will be a treasure trove for discovery. The outcomes, including novel drug targets, biological pathways for disease, and gene-environment interactions, will boost precision medicine approaches, not only in Africa but across the globe

Plasma and cerebrospinal proteomes from children with cerebral malaria differ from those of children with other encephalopathies.

Clinical signs and symptoms of cerebral malaria in children are nonspecific and are seen in other common encephalopathies in malaria-endemic areas. This makes accurate diagnosis difficult in resource-poor settings. Novel malaria-specific diagnostic and prognostic methods are needed. We have used 2 proteomic strategies to identify differentially expressed proteins in plasma and cerebrospinal fluid from children with a diagnosis of cerebral malaria, compared with those with a diagnosis of malaria-slide-negative acute bacterial meningitis and other nonspecific encephalopathies. Here we report the presence of differentially expressed proteins in cerebral malaria in both plasma and cerebrospinal fluid that could be used to better understand pathogenesis and help develop more-specific diagnostic methods. In particular, we report the expression of 2 spectrin proteins that have known Plasmodium falciparum-binding partners involved in the stability of the infected red blood cell, suppressing further invasion and possibly enhancing the red blood cell's ability to sequester in microvasculature

T-cell responses to the DBLα-tag, a short semi-conserved region of the Plasmodium falciparum membrane erythrocyte protein 1.

The Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) is a variant surface antigen expressed on mature forms of infected erythrocytes. It is considered an important target of naturally acquired immunity. Despite its extreme sequence heterogeneity, variants of PfEMP1 can be stratified into distinct groups. Group A PfEMP1 have been independently associated with low host immunity and severe disease in several studies and are now of potential interest as vaccine candidates. Although antigen-specific antibodies are considered the main effector mechanism in immunity to malaria, the induction of efficient and long-lasting antibody responses requires CD4+ T-cell help. To date, very little is known about CD4+ T-cell responses to PfEMP1 expressed on clinical isolates. The DBLα-tag is a small region from the DBLα-domain of PfEMP1 that can be amplified with universal primers and is accessible in clinical parasite isolates. We identified the dominant expressed PfEMP1 in 41 individual clinical parasite isolates and expressed the corresponding DBLα-tag as recombinant antigen. Individual DBLα-tags were then used to activate CD4+ T-cells from acute and convalescent blood samples in children who were infected with the respective clinical parasite isolate. Here we show that CD4+ T-cell responses to the homologous DBLα-tag were induced in almost all children during acute malaria and maintained in some for 4 months. Children infected with parasites that dominantly expressed group A-like PfEMP1 were more likely to maintain antigen-specific IFNγ-producing CD4+ T-cells than children infected with parasites dominantly expressing other PfEMP1. These results suggest that group A-like PfEMP1 may induce long-lasting effector memory T-cells that might be able to provide rapid help to variant-specific B cells. Furthermore, a number of children induced CD4+ T-cell responses to heterologous DBLα-tags, suggesting that CD4+ T-cells may recognise shared epitopes between several DBLα-tags