Growth of Plasmodium falciparum in response to a rotating magnetic field.

BACKGROUND: Plasmodium falciparum is the deadliest strain of malaria and the mortality rate is increasing because of pathogen drug resistance. Increasing knowledge of the parasite life cycle and mechanism of infection may provide new models for improved treatment paradigms. This study sought to investigate the paramagnetic nature of the parasite's haemozoin to inhibit parasite viability. RESULTS: Paramagnetic haemozoin crystals, a byproduct of the parasite's haemoglobin digestion, interact with a rotating magnetic field, which prevents their complete formation, causing the accumulation of free haem, which is lethal to the parasites. Plasmodium falciparum cultures of different stages of intraerythrocytic growth (rings, trophozoites, and schizonts) were exposed to a magnetic field of 0.46 T at frequencies of 0 Hz (static), 1, 5, and 10 Hz for 48 h. The numbers of parasites were counted over the course of one intraerythrocytic life cycle via flow cytometry. At 10 Hz the schizont life stage was most affected by the rotating magnetic fields (p = 0.0075) as compared to a static magnetic field of the same strength. Parasite growth in the presence of a static magnetic field appears to aid parasite growth. CONCLUSIONS: Sequestration of the toxic haem resulting from haemoglobin digestion is key for the parasites' survival and the focus of almost all existing anti-malarial drugs. Understanding how the parasites create the haemozoin molecule and the disruption of its creation aids in the development of drugs to combat this disease.

Long-term in vitro culture of Plasmodium vivax isolates from Madagascar maintained in Saimiri boliviensis blood.

BACKGROUND: Plasmodium vivax is the most prevalent human malaria parasite and is likely to increase proportionally as malaria control efforts more rapidly impact the prevalence of Plasmodium falciparum. Despite the prominence of P. vivax as a major human pathogen, vivax malaria qualifies as a neglected and under-studied tropical disease. Significant challenges bringing P. vivax into the laboratory, particularly the capacity for long-term propagation of well-characterized strains, have limited the study of this parasite's red blood cell (RBC) invasion mechanism, blood-stage development, gene expression, and genetic manipulation. METHODS AND RESULTS: Patient isolates of P. vivax have been collected and cryopreserved in the rural community of Ampasimpotsy, located in the Tsiroanomandidy Health District of Madagascar. Periodic, monthly overland transport of these cryopreserved isolates to the country's National Malaria Control Programme laboratory in Antananarivo preceded onward sample transfer to laboratories at Case Western Reserve University, USA. There, the P. vivax isolates have been cultured through propagation in the RBCs of Saimiri boliviensis. For the four patient isolates studied to-date, the median time interval between sample collection and in vitro culture has been 454 days (range 166-961 days). The median time in culture, continually documented by light microscopy, has been 159 days; isolate AMP2014.01 was continuously propagated for 233 days. Further studies show that the P. vivax parasites propagated in Saimiri RBCs retain their ability to invade human RBCs, and can be cryopreserved, thawed and successfully returned to productive in vitro culture. CONCLUSIONS/SIGNIFICANCE: Long-term culture of P. vivax is possible in the RBCs of Saimiri boliviensis. These studies provide an alternative to propagation of P. vivax in live animals that are becoming more restricted. In vitro culture of P. vivax in Saimiri RBCs provides an opening to stabilize patient isolates, which would serve as precious resources to apply new strategies for investigating the molecular and cellular biology of this important malaria parasite.

Protective Effect of Chronic Schistosomiasis in Baboons Coinfected with Schistosoma mansoni and Plasmodium knowlesi.

Malaria and schistosomiasis coinfections are common, and chronic schistosomiasis has been implicated in affecting the severity of acute malaria. However, whether it enhances or attenuates malaria has been controversial due the lack of appropriately controlled human studies and relevant animal models. To examine this interaction, we conducted a randomized controlled study using the baboon (Papio anubis) to analyze the effect of chronic schistosomiasis on severe malaria. Two groups of baboons (n = 8 each) and a schistosomiasis control group (n = 3) were infected with 500 Schistosoma mansoni cercariae. At 14 and 15 weeks postinfection, one group was given praziquantel to treat schistosomiasis infection. Four weeks later, the two groups plus a new malaria control group (n = 8) were intravenously inoculated with 10(5) Plasmodium knowlesi parasites and monitored daily for development of severe malaria. A total of 81% of baboons exposed to chronic S. mansoni infection with or without praziquantel treatment survived malaria, compared to only 25% of animals infected with P. knowlesi only (P = 0.01). Schistosome-infected animals also had significantly lower parasite burdens (P = 0.004) than the baboons in the P. knowlesi-only group and were protected from severe anemia. Coinfection was associated with increased spontaneous production of interleukin-6 (IL-6), suggesting an enhanced innate immune response, whereas animals infected with P. knowlesi alone failed to develop mitogen-driven tumor necrosis factor alpha and IL-10, indicating the inability to generate adequate protective and balancing immunoregulatory responses. These results indicate that chronic S. mansoni attenuates the severity of P. knowlesi coinfection in baboons by mechanisms that may enhance innate immunity to malaria.

Application of magnetic cytosmear for the estimation of Plasmodium falciparum gametocyte density and detection of asexual stages in asymptomatic children.

BACKGROUND: Conventional malaria parasite detection methods, such as rapid diagnostic tests (RDT) and light microscopy (LM), are not sensitive enough to detect low level parasites and identification of gametocytes in the peripheral blood. A modified and sensitive laboratory prototype, Magnetic Deposition Microscopy (MDM) was developed to increase the detection of sub-microscopic parasitaemia and estimation of gametocytes density in asymptomatic school children. METHODS: Blood samples were collected from 303 asymptomatic school children from seven villages in Bagamoyo district in Tanzania. Participants were screened for presence of malaria parasites in the field using RDT and MDM whereas further examination of malaria parasites was done in the laboratory by LM. LM and MDM readings were used to calculate densities and estimate prevalence of asexual and sexual stages of the parasite. RESULTS: Plasmodium falciparum parasites (asexual and sexual stages) were detected in 23 (7.6 %), 52 (17.2 %), and 59 (19.5 %) out of 303 samples by LM, RDT and MDM respectively. Gametocytes were detected in 4 (1.3 %) and 12 (4.0 %) out of the same numbers of samples by LM, and MDM, respectively. Likewise, in vitro results conducted on two laboratory strains of P. falciparum, 3D7 and NF54 to assess MDM sensitivity on gametocytes detection and its application on concentrating gametocytes indicated that gametocytes were enriched by MDM by 10-fold higher than LM. Late stages of the parasite strains, 3D7 and NF54 were enriched by MDM by a factor of 20.5 and 35.6, respectively. MDM was more specific than LM and RDT by 87.5 % (95 %, CI 71.2-89.6 %) and 89.0 % (95 % CI 82.9-91.4) respectively. It was also found that MDM sensitivity was 62.5 % (95 % CI 49.5-71.8) when compared with RDT while with LM was 36.5 % (95 % CI 32.2-60.5). CONCLUSIONS: These findings provide strong evidence that MDM enhanced detection of sub-microscopic P. falciparum infections and estimation of gametocyte density compared to current malaria diagnostic tools. In addition, MDM is superior to LM in detecting sub-microscopic gametocytaemia. Therefore, MDM is a potential tool for low-level parasitaemia identification and quantification with possible application in malaria transmission research.

Oral pioglitazone reduces infarction volume and improves neurologic function following MCAO in rats.

Thiazolidinediones (TZDs) are neuroprotective in rodent stroke models using intra-peritoneal, intra-venous and inter-ventricular routes of administration.We tested if oral pioglitazone at doses similar to those used by humans to treat diabetes reduces infarction volume following middle cerebral artery occlusion (MCAO) in the rat. Rats were fed DMSO or pioglitazone (0.65 mg/kg equivalent to a 45 mg dose in a 70 kg man, 0.40 mg/kg equivalent to a 30 mg dose or 0.20mg/kg to a 15 mg dose) dissolved in DMSO daily for five days prior to 2 hour MCAO. Animals underwent serial functional analysis using the modified neurologic stroke scale (mNSS), the adhesive sticker test and the inclined plane, all of which test motor sensory function. Twenty one days later, MCAO rats were sacrificed and infarct volumes determined. We found significant reductions in the infarct volume using the 0.65 and 0.40 mg/kg dose. Furthermore, these rats had improved performance on behavioural assays. The 0.20mg/kg dose did not significantly reduce infarction volume or improve behaviour.

Myristicyclins A and B: antimalarial procyanidins from Horsfieldia spicata from Papua New Guinea.

An antimalarial screen for plants collected from Papua New Guinea identified an extract of Horsfieldia spicata as having activity. Isolation of the active constituents led to the identification of two new compounds: myristicyclins A (1) and B (2). Both compounds are procyanidin-like congeners of myristinins lacking a pendant aromatic ring. Myristicyclin A was found to inhibit the ring, trophozoite, and schizont stages of Plasmodium falciparum at similar concentrations in the mid-µM range.

The PPARγ agonist pioglitazone crosses the blood-brain barrier and reduces tumor growth in a human xenograft model.

PURPOSE: The peroxisome proliferator-activated receptor gamma (PPARγ), a member of the nuclear hormone receptor family, represents a target in glioma therapy due to its antineoplastic effects in vitro on human glioma cell lines. We investigate the antineoplastic effects of the PPARγ agonist pioglitazone (pio) in a human glioma xenograft model to define the minimal required dose to induce antineoplastic effects. Additionally, we assess the ability of pio to cross the blood-brain barrier by measuring brain parenchymal concentration after oral administration. METHODS: Human LN-229 cells were injected into the striatum of Balb/cJHanHsd-Prkdc-scid mice. Tumor volumes, invasion, proliferation and parenchymal pio concentrations were measured in this xenograft model after continuous intracerebral drug administration through an osmotic pump or after oral administration. RESULTS: Continuous intracerebral or oral administration of pio reduced tumor volumes, invasion, and proliferation in vivo. To achieve a significant antineoplastic effect, pio needed to be dosed at 240 PPM in the oral group and >1 µM when delivered intracerebrally. After oral pio administration, the drug reached >1 nM levels in brain parenchyma. CONCLUSIONS: These data indicate that pioglitazone crosses the blood-brain barrier and has antineoplastic effects in this glioma xenograft model and may be of potential use in treatment of malignant gliomas.