Malaria Journal

Thrombospondin-related anonymous protein (TRAP) is essential for sporozoite motility and for liver cell invasion. TRAP is a type 1 membrane protein that possesses multiple adhesive domains in its extracellular region. Plasmodium falciparum TRAP (PfTRAP) and its subdomains were expressed in a mammalian expression system, and eleven different mutants generated to study interaction of PfTRAP with liver cells. Binding studies between HepG2 cell extracts and PfTRAP were performed using co-immunoprecipitation protocols. Five different amino acid residues of PfTRAP that are involved in liver cell binding have been identified. These PfTRAP mutants bound to heparin like the wild type PfTRAP thereby suggesting a non-heparin mediated binding of PfTRAP to liver cells. Three Src family proteins -Lyn, Lck and CrkL which interact with PfTRAP are also identified. Liver cell extracts and immunoprecipitated Src family kinases phosphorylated PfTRAP at multiple sites. An analysis of multiple TRAP sequences revealed Src homology 3 domain (SH3) binding motifs. Binding of PfTRAP to SH3-domain containing proteins like Src-family kinases and their ability to phosphorylate PfTRAP suggests a novel role for PfTRAP in cell signaling during sporozoite invasion and homing inside the liver cells. These data shed new light on TRAP-liver cell interactions.

Malaria is one of the top-five contributors to under-5 deaths in Myanmar. Use of insecticide-treated nets (ITN) and receiving early appropriate care in case of fever are the core interventions to prevent malaria and its complications and thereby deaths. This study aimed to assess among the under-five children, (a) utilization of ITNs and its associated factors, (b) care-seeking behaviour among their caregivers and its associated factors and uptake of malaria testing among those with fever in the last 2 weeks. This was a cross sectional study using secondary analysis of Myanmar Demographic and Health Survey (MDHS) conducted in 2015–2016. Multivariable logistic regression was used to explore the factors associated with non-utilization of ITNs and not seeking care for fever. Effect sizes have been presented using odds ratios with 95% confidence intervals. Data analysis was done using svyset command in STATA to account for the multi-stage sampling design of the survey. Of 4597 alive under-five children, 80.5% did not sleep under an ITN last night. The factors significantly associated with non-utilization of ITNs were residing in malaria elimination regions (aOR = 2.0, 1.3–3.2), urban residence (aOR = 1.8, 1.2–2.9), staying in delta region (aOR = 8.7, 4.7–12.2), hilly region (aOR = 3.0, 2.0–4.6, and having highest wealth quintile (aOR = 1.8, 1.1–3.0). Around 16% had fever in the last 2 weeks, of whom 66.7% sought care for fever and 3% got tested for malaria. Nearly half (50.9%) of the caregivers sought care from a government health facility, followed by private hospital/doctor (27.8%), shop (8.0%), village health worker (4.4%) and pharmacy (3.1%). The factors associated with not seeking care for fever were residing in specific geographical locations (hilly, delta and central plains compared to coastal region) and having lowest wealth quintile (aOR = 2.3, 1.1–5.7). This study highlighted that ownership and utilization of ITNs was very poor among under-5children. Care-seeking behaviour of the caregivers of under-5 children in case of fever was dismal with two-thirds not seeking care. The programme should seriously consider addressing these barriers if Myanmar is to achieve zero malaria deaths by 2030.

According to the World Health Organization reports, billions of people around the world are at risk for malaria disease and it is important to consider the preventive strategies for protecting the people that are living in high risk areas. One of the main reasons of disease survival is diversity of vectors and parasites in different malaria regions that have their specific features, behaviour and biology. Therefore, specific regional strategies are necessary for successful control of malaria. One of the tools that needs to be developed for elimination and prevention of reintroduction of malaria is a vaccine that interrupt malaria transmission (VIMTs). VIMT is a broad concept that should be adjusted to the biological characteristics of the disease in each region. One type of VIMT is a vector-based vaccine that affects the sexual stage of Plasmodium life cycle. According to recent studies, the aminopeptidase N-1 of Anopheles gambiae (AgAPN-1) is as a potent vector-based VIMT with considerable inhibition activity against the sexual stage of Plasmodium parasite. Systems for rapid amplification of cDNA ends (3ʹ-RACE) and genome walking methods were used for sequence determination of apn-1 gene from Anopheles stephensi and distinct bioinformatics software were used for structural analysis. AsAPN-1 was expressed in Spodoptera frugiperda (Sf9) insect cell line using the baculovirus expression system. Recombinant AsAPN-1 was purified under the hybrid condition and its biological activity was assayed. Asapn-1 gene and its coded protein from An. stephensi were characterized for the first time in this study. Subsequently, the structural features and immunological properties of its coded protein were evaluated by in silico approaches. Enzymatic activity of the recombinant AsAPN-1, which was expressed in Sf9 insect cell line, was equal to 6 unit/μl. Results of this study revealed that AsAPN-1 is very similar to its counterpart in An. gambiae. In silico evaluation and fundamental data which are necessary for its evaluation as a VIMT-based vaccine in the next steps were acquired in this study and those could be useful for research groups that study on malaria vaccine for countries that An. stephensi is the main malaria vector there.

It has been documented that unplanned urbanization leads to the exposure of members of the Anopheles vectors to a range of water pollution in urban settings. Many surveys from African and Asian countries reported the presence of Anopheles larvae in polluted urban habitats. The present study documents an obvious tolerance of the melanic and normal forms of Anopheles arabiensis to urban polluted larval habitats accompanied by resistance to Temephos larvicide. A cross-sectional survey was carried out to inspect apparently polluted An. arabiensis larval habitats during the hot dry season of 2015. Larval specimens were collected from only apparently polluted habitats after visual inspection from 5 localities in Khartoum State. After morphological and molecular identification of random samples of larvae the magnitude of water pollution was determined using nine abiotic factors. The susceptibility status of An. arabiensis larval forms from normal and polluted habitats to Temephos was tested using the WHO standard diagnostic concentration doses. Morphological and PCR analysis of anopheline larvae revealed the presence of An. arabiensis, a member of the Anopheles gambiae complex. Seven out of 9 physiochemical parameters showed higher concentrations in polluted larval habitats in comparison to control site. Anopheles arabiensis larvae were found in water bodies characterized by high mean of conductivity (1857.8 ± 443.3 uS/cm), turbidity (189.4 ± 69.1 NTU) and nitrate (19.7 ± 16.7 mg/l). The range of mortality rates of An. arabiensis larvae collected from polluted habitats in comparison to An. arabiensis larvae collected from non-polluted habitats was 6.7–64% (LD50 = 1.682) and 67.6–96% (LD50 = 0.806), respectively. The present study reveals that minor populations of An. arabiensis larval forms are adapted to breed in polluted urban habitats, which further influenced susceptibility to Temephos, especially for the melanic larval forms. This could have further implications on the biology of the malaria vector and on the transmission and epidemiology of urban malaria in Sudan.

Recent increases in funding for malaria control have led to the reduction in transmission in many malaria endemic countries, prompting the national control programmes of 36 malaria endemic countries to set elimination targets. Accounting for human population movement (HPM) in planning for control, elimination and post-elimination surveillance is important, as evidenced by previous elimination attempts that were undermined by the reintroduction of malaria through HPM. Strategic control and elimination planning, therefore, requires quantitative information on HPM patterns and the translation of these into parasite dispersion. HPM patterns and the risk of malaria vary substantially across spatial and temporal scales, demographic and socioeconomic sub-groups, and motivation for travel, so multiple data sets are likely required for quantification of movement. While existing studies based on mobile phone call record data combined with malaria transmission maps have begun to address within-country HPM patterns, other aspects remain poorly quantified despite their importance in accurately gauging malaria movement patterns and building control and detection strategies, such as cross-border HPM, demographic and socioeconomic stratification of HPM patterns, forms of transport, personal malaria protection and other factors that modify malaria risk. A wealth of data exist to aid filling these gaps, which, when combined with spatial data on transport infrastructure, traffic and malaria transmission, can answer relevant questions to guide strategic planning. This review aims to (i) discuss relevant types of HPM across spatial and temporal scales, (ii) document where datasets exist to quantify HPM, (iii) highlight where data gaps remain and (iv) briefly put forward methods for integrating these datasets in a Geographic Information System (GIS) framework for analysing and modelling human population and Plasmodium falciparum malaria infection movements.

The survival of malaria parasites, under substantial haem-induced oxidative stress in the red blood cells (RBCs) is dependent on the pentose phosphate pathway (PPP). The PPP is the only source of NADPH in the RBC, essential for the production of reduced glutathione (GSH) and for protection from oxidative stress. Glucose-6-phosphate dehydrogenase (G6PD) deficiency, therefore, increases the vulnerability of erythrocytes to oxidative stress. In Plasmodium, G6PD is combined with the second enzyme of the PPP to create a unique bifunctional enzyme, named glucose-6-phosphate dehydrogenase–6-phosphogluconolactonase (G6PD-6PGL). RRx-001 is a novel, systemically non-toxic, epigenetic anticancer agent currently in Phase 2 clinical development for multiple tumour types, with activity mediated through increased nitric oxide (NO) production and PPP inhibition. The inhibition of G6PD and NO overproduction induced by RRx-001 suggested its application in cerebral malaria (CM). Plasmodium berghei ANKA (PbA) infection in C57BL/6 mice is an experimental model of cerebral malaria (ECM) with several similar pathological features to human CM. This study uses intravital microscopy methods with a closed cranial window model to quantify cerebral haemodynamic changes and leukocyte adhesion to endothelial cells in ECM. RRx-001 had both single agent anti-parasitic activity and significantly increased the efficacy of artemether. In addition, RRx-001 preserved cerebral perfusion and reduced inflammation alone or combined with artemether. RRx-001’s effects were associated with inhibition of PPP (G6PD and G6PD-6PGL) and by improvements in microcirculatory flow, which may be related to the NO donating properties of RRx-001. The results indicate that RRx-001 could be used to potentiate the anti-malarial action of artemisinin, particularly on resistant strains, and to prevent infection.

N,N-Diethyl-3-methylbenzamide (DEET) topical mosquito repellents are effective personal protection tools. However, DEET-based repellents tend to have low consumer acceptability because they are cosmetically unappealing. More attractive formulations are needed to encourage regular user compliance. This study evaluated the protective efficacy and protection duration of a new topical repellent ointment containing 15% DEET, MAÏA® compared to 20% DEET in ethanol using malaria and dengue mosquito vectors in Bagamoyo Tanzania. Fully balanced 3 × 3 Latin square design studies were conducted in large semi-field chambers using laboratory strains of Anopheles gambiae sensu stricto, Anopheles arabiensis and Aedes aegypti. Human volunteers applied either MAÏA® ointment, 20% DEET or ethanol to their lower limbs 6 h before the start of tests. Approximately 100 mosquitoes per strain per replicate were released inside each chamber, with 25 mosquitoes released at regular intervals during the collection period to maintain adequate biting pressure throughout the test. Volunteers recaptured mosquitoes landing on their lower limbs for 6 h over a period of 6 to 12-h post-application of repellents. Data analysis was conducted using mixed-effects logistic regression. The protective efficacy of MAÏA® and 20% DEET was not statistically different for each of the mosquito strains: 95.9% vs. 97.4% against An. gambiae (OR = 1.53 [95% CI 0.93–2.51] p = 0.091); 96.8% vs 97.2% against An. arabiensis (OR = 1.08 [95% CI 0.66–1.77] p = 0.757); 93.1% vs 94.6% against Ae. aegypti (OR = 0.76 [95% CI 0.20–2.80] p = 0.675). Average complete protection time (CPT) in minutes of MAÏA® and that of DEET was similar for each of the mosquito strains: 571.6 min (95% CI 558.3–584.8) vs 575.0 min (95% CI 562.1–587.9) against An. gambiae; 585.6 min (95% CI 571.4–599.8) vs 580.9 min (95% CI 571.1–590.7) against An. arabiensis; 444.1 min (95% CI 401.8–486.5) vs 436.9 min (95% CI 405.2–468.5) against Ae. aegypti. MAÏA® repellent ointment provides complete protection for 9 h against both An. gambiae and An. arabiensis, and 7 h against Ae. aegypti similar to 20% DEET (in ethanol). MAÏA® repellent ointment can be recommended as a tool for prevention against outdoor biting mosquitoes in tropical locations where the majority of the people spend an ample time outdoor before going to bed.

The detection of submicroscopic infections in low prevalence settings has become an increasingly important challenge for malaria elimination strategies. The current field rapid diagnostic tests (RDTs) for Plasmodium falciparum malaria are inadequate to detect low-density infections. Therefore, there is a need to develop more sensitive field diagnostic tools. In parallel, a highly sensitive laboratory reference assay will be essential to evaluate new diagnostic tools. Recently, the highly sensitive Alere™ Malaria Ag P.f ELISA (HS ELISA) was developed to detect P. falciparum histidine-rich protein 2 (HRP2) in clinical whole blood specimens. In this study, the analytical and clinical performance of the HS ELISA was determined using recombinant P. falciparum HRP2, P. falciparum native culture parasites, and archived highly pedigreed clinical whole blood specimens from Karen village, Myanmar and Nagongera, Uganda. The HS ELISA has an analytical sensitivity of less than 25 pg/mL and shows strong specificity for P. falciparum HRP2 when tested against P. falciparum native culture strains with pfhrp2 and pfhrp3 gene deletions. Additionally, the Z′-factor statistic of 0.862 indicates the HS ELISA as an excellent, reproducible assay, and the coefficients of variation for inter- and intra-plate testing, 11.76% and 2.51%, were acceptable. Against clinical whole blood specimens with concordant microscopic and PCR results, the HS ELISA showed 100% (95% CI 96.4–100) diagnostic sensitivity and 97.9% (95% CI 94.8–99.4) diagnostic specificity. For P. falciparum positive specimens with HRP2 concentrations below 400 pg/mL, the sensitivity and specificity were 100% (95% CI 88.4–100) and 88.9% (95% CI 70.8–97.6), respectively. The overall sensitivity and specificity for all 352 samples were 100% (CI 95% 96–100%) and 97.3% (CI 95% 94–99%). The HS ELISA is a robust and reproducible assay. The findings suggest that the HS ELISA may be a useful tool as an affordable reference assay for new ultra-sensitive HRP2-based RDTs.