Host-parasite interaction and morbidity in malaria endemic areas.

Severe morbidity due to Plasmodium falciparum is a major health problem in African children. The patterns of morbidity in endemic areas are modified by the immune response, and vary markedly with transmission intensity. Severe disease falls into three overlapping syndromes: coma, respiratory distress, and severe anaemia. Recently, it has become clear that metabolic acidosis plays a major role in the pathogenesis of severe disease and is particularly important in the overlap between the different clinical syndromes. We propose that the different manifestations of severe malarial morbidity arise from the interaction of a limited number of pathogenic processes: red cell destruction, toxin-mediated activation of cytokine cascades, and infected cell sequestration in tissue microvascular beds. The pattern of severe morbidity varies with age within any one endemic area, with severe anaemia predominating in the youngest children and coma having its highest incidence in older children. Between endemic areas there is a marked variation in mean age of children with severe malaria, and therefore in the importance of different clinical syndromes. The shift in mean age is due to a combination of increased challenge and more rapid development of immunity at higher levels of transmission. Recent comparative studies indicate that at higher levels of transmission the net effect of these shifts may be a paradoxical reduction in total severe malarial morbidity.     

Anti-malarial drugs and the prevention of malaria in the population of malaria endemic areas

Anti-malarial drugs can make a significant contribution to the control of malaria in endemic areas when used for prevention as well as for treatment. Chemoprophylaxis is effective in preventing deaths and morbidity from malaria, but it is difficult to sustain for prolonged periods, may interfere with the development of naturally acquired immunity and will facilitate the emergence and spread of drug resistant strains if applied to a whole community. However, chemoprophylaxis targeted to groups at high risk, such as pregnant women, or to periods of the year when the risk from malaria is greatest, can be an effective and cost effective malaria control tool and has fewer drawbacks. Intermittent preventive treatment, which involves administration of anti-malarials at fixed time points, usually when a subject is already in contact with the health services, for example attendance at an antenatal or vaccination clinic, is less demanding of resources than chemoprophylaxis and is now recommended for the prevention of malaria in pregnant women and infants resident in areas with medium or high levels of malaria transmission. Intermittent preventive treatment in older children, probably equivalent to targeted chemoprophylaxis, is also highly effective but requires the establishment of a specific delivery system. Recent studies have shown that community volunteers can effectively fill this role. Mass drug administration probably has little role to play in control of mortality and morbidity from malaria but may have an important role in the final stages of an elimination campaign.     

Genes,nitric oxide and malaria in African children

The unresolved and complex relationship between nitric oxide and falciparum malaria is reflected in recent genetic and immunohistochemical studies in African children. Different genetic associations, perhaps geographically distinctive, are seen between genetic variants of the inducible nitric oxide gene and various disease manifestations in African populations. The picture might not be complete without considering the emerging roles of carbon monoxide, another endogenous gaseous mediator with similar effects to those of nitric oxide. Only when genetic comparisons from across tropical Africa are examined, in conjunction with the newly recognized complexities in the events of systemic inflammation, will this relationship be understood.     

Community knowledge and practices regarding malaria and long-lasting insecticidal nets during malaria elimination programme in an endemic area in Iran

A literature review for operational research on malaria control and elimination was conducted using the term 'malaria' and the definition of operational research (OR). A total of 15 886 articles related to malaria were searched between January 2008 and June 2013. Of these, 582 (3.7%) met the definition of operational research. These OR projects had been carried out in 83 different countries. Most OR studies (77%) were implemented in Africa south of the Sahara. Only 5 (1%) of the OR studies were implemented in countries in the pre-elimination or elimination phase. The vast majority of OR projects (92%) were led by international or local research institutions, while projects led by National Malaria Control Programmes (NMCP) accounted for 7.8%. With regards to the topic under investigation, the largest percentage of papers was related to vector control (25%), followed by epidemiology/transmission (16.5%) and treatment (16.3%). Only 19 (3.8%) of the OR projects were related to malaria surveillance. Strengthening the capacity of NMCPs to conduct operational research and publish its findings, and improving linkages between NMCPs and research institutes may aid progress towards malaria elimination and eventual eradication world-wide.     

Diversity of Anopheles species and trophic behavior of putative malaria vectors in two malaria endemic areas of northwestern Thailand

We determined the species diversity, blood‐feeding behavior, and host preference of Anopheles mosquitoes in two malaria endemic areas of Tak (Mae Sot District) and Mae Hong Son (Sop Moei District) Provinces, located along the Thai border with Myanmar, during a consecutive two‐year period. Anopheline mosquitoes were collected using indoor and outdoor human‐landing captures and outdoor cow‐baited collections. Mosquitoes were initially identified using morphological characters, followed by the appropriate multiplex AS‐PCR assay for the identification of sibling species within Anopheles (Cellia) complexes and groups present. Real‐time PCR was performed for parasite‐specific detection in mosquitoes (Plasmodium spp. and Wuchereria bancrofti). A total of 7,129 Anopheles females were captured, 3,939 from Mae Sot and 3,190 from Sop Moei, with 58.6% and 37% of all anophelines identified as An. minimus, respectively. All three malaria vector complexes were detected in both areas. One species within the Minimus Complex (An. minimus) was present along with two related species in the Funestus Group, (An. aconitus, An. varuna), two species within the Dirus Complex (An. dirus, An. baimaii), and four species within the Maculatus Group (An. maculatus, An. sawadwongporni, An. pseudowillmori, and An. dravidicus). The trophic behavior of An. minimus, An. dirus, An. baimaii, An. maculatus, and An. sawadwongporni are described herein. The highest An. minimus densities were detected from February through April of both years. One specimen of An. minimus from Mae Sot was found positive for Plasmodium vivax.     

Cross-talk between nitric oxide and transforming growth factor-beta1 in malaria

Malaria has re-emerged as a global health problem, leading to an increased focus on the cellular and molecular biology of the mosquito Anopheles and the parasite Plasmodium with the goal of identifying novel points of intervention in the parasite life cycle. Anti-parasite defenses mounted by both mammalian hosts and Anopheles can suppress the growth of Plasmodium. Nonetheless, the parasite is able to escape complete elimination in vivo, perhaps by thwarting or co-opting these mechanisms for its own survival, as do numerous other pathogens. Among the defense systems used by the mammalian host against Plasmodium is the synthesis of nitric oxide (NO), catalyzed by an inducible NO synthase (iNOS). Nitric oxide produced by the action of an inducible Anopheles stephensi NO synthase (AsNOS) may be central to the anti-parasitic arsenal of this mosquito. In mammals, iNOS can be modulated by members of the transforming growth factor-beta (TGF-beta) cytokine superfamily. Transforming growth factor-beta is produced as an inactive precursor that is activated following dissociation of certain inhibitory proteins, a process that can be promoted by reaction products of NO as well as by hemin. Ingestion by Anopheles of blood containing Plasmodium initiates parasite development, blood digestion which results in the accumulation of hematin (hemin) in the insect midgut, and induction of both AsNOS and TGF-beta-like (As60A) gene expression in the midgut epithelium. Active mammalian TGF-beta1 can be detected in the A. stephensi midgut up to 48h post-ingestion and latent TGF-beta1 can be activated by midgut components in vitro, a process that is potentiated by NO and that may involve hematin. Further, mammalian TGF-beta1 is perceived as a cytokine by A. stephensi cells in vitro and can alter Plasmodium development in vivo. Bloodfeeding by Anopheles, therefore, results in a juxtaposition of evolutionarily conserved mosquito and mammalian TGF-beta superfamily homologs that may influence transmission dynamics of Plasmodium in endemic regions.     

Transgenic expression of human C-reactive protein suppresses endothelial nitric oxide synthase expression and bioactivity after vascular injury

C-reactive protein (CRP) is a risk marker and a potential modulator of vascular disease. Whether CRP modulates nitric oxide (NO) synthase (NOS) activity and NO metabolism remains unclear. We studied the effect of CRP on NO metabolism in transgenic mice that express human CRP (CRPtg). CRPtg and wild-type mice were subjected to controlled femoral artery wire injury. CRP serum levels at baseline and 6 and 24 h after injury were 12.4 +/- 9, 18.6 +/- 6.9, and 58.4 +/- 13 mg/l, respectively, in CRPtg mice but were undetectable at all time points in wild-type mice. Endothelial NOS protein and mRNA expression were significantly suppressed in the injured arteries of CRPtg mice (n = 5, P < 0.05). A similar reduction in eNOS expression was observed in the distant lung and heart. NO release after injury was significantly lower in CRPtg mice, as measured by nitrate and nitrite breakdown products, with a concomitant suppression of cGMP NO signaling after injury. Endothelial NOS and NO expression after vascular injury are locally and systemically suppressed in mice that express human CRP. These in vivo observations support the hypothesis that CRP modulates NO metabolism and may have implications regarding the mechanisms by which CRP modulates vascular disease.     

Effects of treatment and drug resistance on the transmission dynamics of malaria in endemic areas

We present a mathematical model for malaria treatment and spread of drug resistance in an endemic population. The model considers treated humans that remain infectious for some time and partially immune humans who are also infectious to mosquitoes although their infectiousness is always less than their non immune counterparts. The model is formulated by considering delays in the latent periods in both mosquito and human populations and in the period within which partial immunity is lost. Qualitative analysis of the model including positivity and boundedness of solutions is performed. Analysis of the reproductive numbers shows that if the treated humans become immediately uninfectious to mosquitoes then treatment will always reduce the number of sensitive infections. If however treated humans are infectious then for treatment to effectively reduce the number of sensitive infections, the ratio of the infectious period of the treated humans to the infectious period of the untreated humans multiplied by the ratio of the transmission rate from a treated human to the transmission rate of an untreated human should be less than one. Our results show that the spread of drug resistance with treatment as a control strategy depends on the ratio of the infectious periods of treated and untreated humans and on the transmission rates from infectious humans with resistant and sensitive infections. Numerical analysis is performed to assess the effects of treatment on the spread of resistance and infection. The study provides insight into the possible intervention strategies to be employed in malaria endemic populations with resistant parasites by identifying important parameters.     

Proposed link between cytokines, nitric oxide and human cerebral malaria

Nitric oxide (NO), also known as endothelial-derived relaxing factor (EDRF), is generated by a range of cell types including endothelial cells, smooth muscle cells and neurons, and mediates a range of different physiological functions, such as maintenance of vascular tone and neuro-transmission. In this article, Ian Clark, Kirk Rockett and Bill Cowden propose that when vascular generation of NO is particularly high (for example, if local intravascular levels of tumour necrosis factor (TNF) are markedly increased) this mediator could diffuse to nearby neurons, be misinterpreted as being of synaptic origin and thus interfere with orderly neuro-transmission. NO of vascular origin could also, through vasodilation of cerebral vessels, contribute to increased intracranial pressure and thus to certain of the clinical signs seen in cerebral malaria. As well as contributing to cerebral malaria, these phenomena could also lead to the neurological changes observed in certain other systemic diseases.     

The role of nitric oxide and lipid peroxidation in patients with Plasmodium vivax malaria

In this study, we investigated the role of nitric oxide metabolism and lipid peroxidation in patients with P. vivax malaria. The levels of nitrite and nitrate were analyzed using a procedure based on the Griess reaction and malondialdehyde levels which index of lipid peroxidation was determined by thiobarbituric acid reaction. The levels of nitrite/nitrate and malondialdehyde in patients were higher than controls and found to be statistically significant (p < 0.001). We performed this study to determine whether nitric oxide and lipid peroxidation is produced during blood-stage P. vivax malaria. This present study shows that lipid peroxidation occurs in P. vivax malaria. The levels of nitric oxide are associated with lipid peroxidation in this disease.     

Long‐term survival and diversification of an endemic Melitaea species in mountains of Iran and adjacent areas

Disjunct distribution patterns regularly resulted in the separation of different genetic lineages in glacial refugia. Recent patterns of survival and expansion have been often revealed by climatic niche modelling. We used the combination of genetic markers, geometric morphometry and climatic niche modelling to clear up the taxonomy and reconstruct the potential range of an endemic Iranian, taxonomically disputed Melitaea species in climatically different epochs. Our results show that this species (Melitaea abbas Gross and Ebert 1975, comb. n. = M. zagrosi Tóth and Varga syn. nova) is clearly separated from all taxa of the Melitaea phoebe species group and only occurs in Iran and Azerbaijan but were also predicted for some adjacent regions. Molecular markers and distribution modelling show consistently that this species should have had a long‐term survival in this area, and its range could have been slightly larger during the last glacial maximum than currently. Based on the studied molecular markers, three main groups in M. abbas can be recognized: those of steppic area of Azerbaijan, western Iran and northeastern Iran. Each group is characterized by own mitochondrial haplotypes, but also a high level of genetic diversity appears in the central part of the distribution area (Zagros Mts.).     

C-reactive protein down-regulates endothelial nitric oxide synthase expression and promotes apoptosis in endothelial progenitor cells through receptor for advanced glycation end-products

Objective

C-reactive protein (CRP), the prototypic marker of inflammation, has been shown to be an independent predictor of atherosclerosis. CRP can regulate receptor for advanced glycation end-products (RAGE) expression in endothelial progenitor cells (EPCs). Endothelial nitric oxide synthase (eNOS) deficiency is a pivotal event in atherogenesis. It is believed that decreased eNOS bioactivity occurs early in atherogenesis. Therefore, we tested the hypothesis that CRP can alter eNOS expression and promote apoptosis in EPCs through RAGE.

Methods and results

EPCs, isolated from bone marrow, were cultured in the presence or absence of LPS-free CRP (5, 10, 15, 20, and50 μg/ml). RAGE protein expression and siRNA were measured by flow cytometric analysis. PCR was used to detect eNOS mRNA expression. eNOS protein expression was measured by Western blot analysis. A spectrophotometer was used to assess eNOS activity. A modified Boyden's chamber was used to assess the migration of EPCs and the number of recultured EPCs was counted to measure adhesiveness. A MTT assay was used to determine proliferation. Apoptosis was evaluated by annexin V immunostaining and TUNEL staining. Co-culturing with CRP caused a significant down-regulation of eNOS expression, inhibited the proliferation, migration, and adhesion of EPCs, and induced EPC apoptosis. In addition, these effects were attenuated during RAGE protein expression blockade by siRNA.

Conclusions

CRP, at concentrations known to predict cardiovascular event, directly quenches the expression of eNOS and diminishes NO production, and may serve to impair EPC function and promote EPC apoptosis through RAGE. These data further support a direct role of CRP in the development and/or progression of atherosclerosis and indicate a new pathophysiologic mechanism of disturbed vascular adaptation in atherosclerosis.     

Inflammatory status and preerythrocytic stages of malaria: role of the C-reactive protein

In the acquisition of protection against malaria, the role played by nonspecific factors, some being part of the cascade effect of cytokines, has to be considered. The C-reactive protein, a major acute phase reactant secreted by interleukin-1 stimulated hepatocytes, has an effect on the hepatic development of Plasmodia, both by preventing penetration of the sporozoite into the hepatocyte and by blocking parasite division through an antibody-like effect. This latter effect confirms the potential interest of targeting the uninuclear form of the parasite. Nevertheless, C-reactive Protein alone does not account for all the effects of the inflammatory response, other reactants from both serum and hepatocytes are also involved.     

Low nitric oxide bioavailability contributes to the genesis of experimental cerebral malaria

The role of nitric oxide (NO) in the genesis of cerebral malaria is controversial. Most investigators propose that the unfortunate consequence of the high concentrations of NO produced to kill the parasite is the development of cerebral malaria. Here we have tested this high NO bioavailability hypothesis in the setting of experimental cerebral malaria (ECM), but find instead that low NO bioavailability contributes to the genesis of ECM. Specifically, mice deficient in vascular NO synthase showed parasitemia and mortality similar to that observed in control mice. Exogenous NO did not affect parasitemia but provided marked protection against ECM; in fact, mice treated with exogenous NO were clinically indistinguishable from uninfected mice at a stage when control infected mice were moribund. Administration of exogenous NO restored NO-mediated signaling in the brain, decreased proinflammatory biomarkers in the blood, and markedly reduced vascular leak and petechial hemorrhage into the brain. Low NO bioavailability in the vasculature during ECM was caused in part by an increase in NO-scavenging free hemoglobin in the blood, by hypoargininemia, and by low blood and erythrocyte nitrite concentrations. Exogenous NO inactivated NO-scavenging free hemoglobin in the plasma and restored nitrite to concentrations observed in uninfected mice. We therefore conclude that low rather than high NO bioavailability contributes to the genesis of ECM.     

Multimodality imaging of nitric oxide and nitric oxide synthases

Nitric oxide (NO) and NO synthases (NOSs) are crucial factors in many pathophysiological processes such as inflammation, vascular/neurological function, and many types of cancer. Noninvasive imaging of NO or NOS can provide new insights in understanding these diseases and facilitate the development of novel therapeutic strategies. In this review, we will summarize the current state-of-the-art multimodality imaging in detecting NO and NOSs, including optical (fluorescence, chemiluminescence, and bioluminescence), electron paramagnetic resonance (EPR), magnetic resonance (MR), and positron emission tomography (PET). With continued effort over the last several years, these noninvasive imaging techniques can now reveal the biodistribution of NO or NOS in living subjects with high fidelity which will greatly facilitate scientists/clinicians in the development of new drugs and/or patient management. Lastly, we will also discuss future directions/applications of NO/NOS imaging. Successful development of novel NO/NOS imaging agents with optimal in vivo stability and desirable pharmacokinetics for clinical translation will enable the maximum benefit in patient management.     

Lipid peroxides, nitric oxide and essential fatty acids in patients with Plasmodium falciparum malaria

Long chain polyunsaturated fatty acids derived from essential fatty acids have been shown to be toxic to Plasmodium falciparum both in vitro and in vivo. Here, we present evidence to suggest that in patients with Plasmodium falciparum malaria the levels of lipid peroxides (a marker of free radical generation) nitric oxide (a potent free radical with immunomodulatory actions), and concentrations of linoleic acid (LA) and alpha-linolenic acid (ALA) are low, whereas those of eicosapentaenoic acid (EPA) are high. The ability of the fatty acids to kill P. falciparum is dependent on their capacity to stimulate free radical generation in neutrophils and macrophages. EPA is more potent than LA in killing the parasite. In view of this, the results of the present study suggest that in patients with P. falciparum malaria the decreased levels of lipid peroxides and nitric oxide may contribute to the persistence of the infection, whereas elevated levels of EPA may be a feeble attempt to overcome this defect.     

von Willebrand factor, C-reactive protein, nitric oxide, and vascular endothelial growth factor in a dietary reversal model of hypercholesterolemia in rabbit

Aims: Endothelial dysfunction is considered a sign of the early vascular changes preceding atherosclerosis. We studied the alteration of von Willebrand Factor (vWF), C - reactive protein (CRP), nitrite and Vascular Endothelial Growth Factor (VEGF) in a dietary reversal model of hypercholesterolemia in rabbit. Methods: This project was designed in two phases. In phase I, male rabbits (n = 11) were fed a 1% high cholesterol diet for 30 days. Then the diet was replaced with normal rabbit chow for other 30 days (cholesterol withdrawal phase, phase II). To compare the fatty streak formation with normal condition, a control group (n = 6) received normal diet during the study. The serum lipid levels, vWF, CRP, nitrite, and VEGF were measured before the experiment and by the end of each phase. Fatty streak formation in the walls of the aortas in both groups (high cholesterol diet and control group) was determined using intima thickness/media thickness (IMT) ratio. Results: The results indicate that the level of cholesterol, Low Density Lipoproteins (LDL), vWF and CRP increased significantly in phase I, and decreased after hypercholesterolemic diet withdrawal (p < 0.05). No statistically significant changes were found in VEGF levels but the serum level of nitrite increased significantly during both phases of the study (p < 0.05). The IMT ratio in the walls of aortas was significantly different between the groups in both phases of studies (p < 0.05). There was a significant correlation between nitrite and cholesterol levels in both phases (r = 0.62 and r = 0.98, p < 0.05). Nitrite concentration also correlated with IMT ratio in both phases of the study (r = 0.75 and r = -0.99, p < 0.05). vWF did not correlate with cholesterol but it correlated with IMT ratio in both phases of the study (r = 0.87 and r = 0.84, p < 0.05). CRP only correlated with cholesterol in the first phase (r = 0.91, p < 0.05). Conclusions: Among the endothelial biomarkers, vWF was found to be a biological marker for identifying the risk of developing atherosclerosis; however a single biomarker may not provide appropriate information.