Both Glycolipid and Protein Components are Required for Plasmodium falciparum induced TNF-α and IL-1β Production in Human Monocytic Cells

Tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) are the endogenous pyrogens which mediate fever in malaria. The excessive production of TNF-α is associated with pathology of human malaria. The nature and properties of malaria antigens, which stimulate monocyte to secrete these cytokines were studied in vitro using human monocytic cell line THP-1. THP-1 cells produced the cytokines in response to Plasmodium falciparum malaria antigens similar to the response of peripheral blood monocytes. Malaria parasite components of infected erythrocytes and their culture supernatant were separately analyzed. Soluble and insoluble components of the infected erythrocytes and their culture supernatant stimulated cytokine production by THP-1 cells. Acid, base and pronase treatments of malaria culture supernatant greatly reduced the cytokine inducing activity, suggesting that both glycolipid and protein components are essential for cell stimulation. Considering the ultrafiltration results together, we assume that a complex of glycolipid and protein stimulates host cells to induce cytokine secretion. Application of Triton X-114 solubilization and phase separation procedures to the infected erythrocytes revealed that the membrane-free hemozoin pellet did not have any stimulation activity, whereas the hydrophobic components seemed to contribute to TNF-α and IL-1β production

Coexistence of GP195 Alleles of Plasmodium Falciparum in a Small Endemic Area

Dimorphic variations in the genotype of the precursor to Plasmodium falciparum major merozoite surface antigens or gp195, among wild isolates in a small malaria parasite population were examined using Southern blot hybridization techniques. Hybridization, with DNA fragment probes and oligonucleotide probes derived from variable blocks of known gp195 alleles against 18 wild isolates from Mae Sod district in Thailand, revealed the existence of seven gp195 alleles, two of which were newly identified in this study. In four out of 17 patients, two different alleles coexisted in the circulation. It was furthermore noted that the seven alleles did not occur at the same frequency, but rather several alleles predominated in the population of P. falciparum in this small malaria field

Loss of Tumor Necrosis Factor Production by Human Monocytes in Falciparum Malaria after Their Maturation in Vitro

In Plasmodium-infected mammals, phagocytosis and production of tumor necrosis factor (TNF) by monocytes and macrophages are prominent features. The present work aimed at clarifying the relationship between the maturation of human monocytes to macrophages and their TNF productivity and phagocytic ability in the presence of Plasmodium falciparum-infected erythrocytes. Fresh monocytes produced a significantly higher quantity of TNF in the presence of schizont-infected erythrocytes than macrophages obtained by in vitro monocyte maturation on autologous serum, whereas phagocytic activity of macrophages was much higher than that of fresh monocytes. This indicated that the TNF-inducing factors from P. falciparum-infected erythrocytes could stimulate fresh monocytes, but not macrophages, to release TNF, regardless of their development of phagocytosis. Activation of macrophages by interferon-{gamma} could not recover their TNF productivity in the presence of P. falciparum-infected erythrocytes, but it enhanced their TNF productivity in the presence of lipopolysaccharide(s). The TNF-inducing factors were contained mainly in erythrocytes infected with mature schizonts but not in erythrocytes infected with the younger stages of the parasites. Fractionation of infected erythrocytes revealed that both soluble and insoluble components almost equally contained those factors

Polymorphism of Plasmodium Falciparum Dihydrofolate Reductase and Dihydropteroate Synthase Genes Among Pregnant Women with Falciparum Malaria in Banjar District, South Kalimantan Province, Indonesia

Pregnant women are highly vulnerable to malaria infection in its endemic areas, particularly infection by Plasmodium falciparum that can cause premature, low birth weight, severe anemia in pregnant women, and death. Sulfadoxine-pyrimethamine (SP) for Intermittent Preventive Treatment for pregnant (IPTp) is used for malaria control in pregnancy recommended by the World Health Organization that has already been implemented in Africa. The P. falciparum resistance to SP has been reported in several malarial endemic areas, and mutations in the genes of Plasmodium falciparum Dihydrofolate Reductase (Pfdhfr) and Dihydropteroate Synthase (Pfdhps) are shown to be associated with parasite resistance to SP treatment. Genetic analysis of Pfdhfr and Pfdhps genes in pregnant women infected with P. falciparum has not yet been examined in Indonesia. The cross-sectional study was conducted at two subdistricts, Sungai Pinang and Peramasan, in Banjar district of South Kalimantan Province, where 127 pregnant women were recruited from 2008 to April 2010. Two important mutations in Pfdhfr gene (amino acid positions at N51 and S108) and three in Pfdhps gene (A437, K540 and A581) were analyzed by nested PCR-RFLP method. All of the seven pregnant women samples infected with P. falciparum presented PfDHFR 108N and PfDHPS 437G mutations. One of the samples had the additional mutation at PfDHPS 540, in which Lys is substituted by Glu. These results suggested that P. falciparum might present only some resistance to SP at Sungai Pinang and Peramasan subdistricts, Banjar District, South Kalimantan province, Indonesia. Although there were limited number of samples, this study showed only few mutations of Pfdhfr and Pfdhps genes in P. falciparum at Banjar district, South Kalimantan Province, that suggests SP might be effective for IPTp in this area. Thus, further analysis of the other mutation sites in Pfdhfr and Pfdhps genes and in vivo efficacy study of SP with more sufficient sample numbers will be necessary to confirm this preliminarily result

Two novel mutations of pfdhps K540T and I588F, affecting sulphadoxine-pyrimethamine-resistant response in uncomplicated falciparum malaria at Banjar district, South Kalimantan Province, Indonesia

Background: Mutations in pfdhfr and pfdhps genes have been shown to associate with sulphadoxine-pyrimethamine (SP) resistance of Plasmodium falciparum parasites. However, pfdhfr, pfdhps genotypes and the correlations to SP treatment outcome in Indonesia has not yet been well analysed. Methods. After obtaining informed consent, 61 uncomplicated falciparum malaria patients were recruited in Banjar district, South Kalimantan Province, Indonesia, from October 2009 to August 2010. They were treated by a single oral dose of SP and its effects on clinical and parasitological status were followed until day 28 after treatment. Occasionally, a thick smear blood film for microscopy observation and blood spot on a filter paper for pfdhfr and pfdhps genotype analysis were collected. Results: Pfdhfr and pfdhps genotypes from 24 P. falciparum-infected patients consisting of adequate clinical parasitological response (ACPR) (n = 6; 25.0%) and early treatment failure (ETF) (n = 10; 41.7%) or late parasitological failure (LPF) (n = 8; 33.3%) were obtained by sequencing. Two novel mutations of pfdhps gene, K540T and I588F, were determined in ten and five isolates, respectively. These mutations were present in the pfdhfr/pfdhps combined haplotypes of ANRNI/SGTGA (n = 6), ANRNL/SGTGA (n = 4), and ANRNI/SGEAA(588F) (n = 5), (mutation codons are bold typed); these haplotypes were mostly belonging to parasitological failure (ETF or LPF). The parasites acquiring five mutations in pfdhfr/pfdhps haplotypes and four mutations with additional I588F did not respond adequately to SP treatment. Conclusion: Many of Plasmodium falciparum infected patients in Banjar district, South Kalimantan, Indonesia did not respond adequately to SP treatment and these low ineffectiveness of SP in this area was associated with two novel mutations of pfdhps, K540T and I588F

Extracellular cyclophilin A possesses chemotaxic activity in cattle

International audienceCyclophilin A (CyPA) was originally discovered in bovine thymocytes as a cytosolic binding protein of the immunosuppressive drug cyclosporine A. Recent studies have revealed that in mice and humans, CyPA is secreted from cells in injured or infected tissues and plays a role in recruiting inflammatory cells in those tissues. Here we found that in cattle abundant level of extracellular CyPA was observed in tissues with inflammation. To aid in investigating the role of extracellular CyPA in cattle, we generated recombinant bovine CyPA (rbCyPA) and tested its biological activity as an inflammatory mediator. When bovine peripheral blood cells were treated with rbCyPA in vitro, we observed that rbCyPA reacts with the membranous surface of granulocytes, monocytes and lymphocytes. Chemotaxis analysis showed that the granulocytes migrate toward rbCyPA and the migration is inhibited by pre-treatment with an anti-bovine CyPA antibody. These results indicate that, as for mice and humans, extracellular CyPA possesses chemotactic activity to recruit inflammatory cells (e.g., granulocytes) in cattle, and could thus be a potential therapeutic target for the treatment of inflammation

Diagnosis of visceral leishmaniasis by polymerase chain reaction of DNA extracted from Giemsa\u27s solution-stained slides.

Visceral leishmaniasis (VL) is caused by the protozoan parasite Leishmania donovani and is a potentially fatal disease in endemic areas of the world. Nepal is an endemic area in which VL causes major public health problems in the lowland areas of the southeast regions. The aim of the present study was to evaluate the sensitivity of polymerase chain reaction (PCR) amplification for the detection of Leishmania DNA from Giemsa\u27s solution-stained bone marrow slides. Bone marrow samples were aspirated from a total of 115 VL suspected patients and used to prepare smears on glass slides and for the initiation of in vitro culture. Bone marrow slides were used for microscopic observation, DNA extraction, and subsequent PCR amplification. PCR analysis showed that all the positive samples were of Leishmania parasites. The PCR assay also showed a higher sensitivity (69%) than microscopic examination (57%) and culture (21%). In addition, PCR was able to detect VL in 12% of samples which were negative by microscopy. PCR of DNA extracted from Giemsa\u27s solution-stained bone marrow slides is a suitable tool for confirming diagnosis in patients with VL and may also be useful in the diagnosis of difficult cases. Bone marrow smears are easily stored and can be easily sent to research centers where PCR is available. This makes PCR a good option for diagnosis in the field

Stent-Jack Technique for Ruptured Vertebral Artery Dissecting Aneurysm Involving the Origin of Posterior Inferior Cerebellar Artery

We herein report a case of a ruptured vertebral artery dissecting aneurysm involving the origin of the posterior inferior cerebellar artery that was treated using the stent-jack technique. After parent artery occlusion of the distal vertebral artery, stenting of the posterior inferior cerebellar artery was performed. Further coiling was needed because distal vertebral artery recanalization occurred due to transformation of the coil mass. The stent-jack technique for a ruptured vertebral artery dissecting aneurysm involving the origin of the posterior inferior cerebellar artery is effective; however, careful attention to recanalization after stenting is needed due to transformation of the coil mass

Longitudinal survey of Plasmodium falciparum infection in Vietnam: characteristics of antimalarial resistance and their associated factors.

Plasmodium falciparum is the main cause of human malaria and is one of the important pathogens causing high rates of morbidity and mortality. The total number of malaria patients in Vietnam has gradually decreased over the last decade. However, the spread of pathogens with drug resistance remains a significant problem. Defining the trend in genotypes related to drug resistance is essential for the control of malaria in Vietnam. We undertook a longitudinal survey of Plasmodium falciparum malaria in 2001, 2002, and 2005 to 2007. The pfcrt, pfmdr1, pfdhfr, and pfdhps genes were analyzed by sequencing; and correlations by study year, age, gender, and genotype were identified statistically. The ratio of the chloroquine resistance genotype pfcrt 76T was found to have decreased rapidly after 2002. High numbers of mutations in the pfdhfr and pfdhps genes were observed only in 2001 and 2002, while the emergence of parasites with a new K540Y mutation in the P. falciparum dihydropteroate synthetase (PfDHPS) was observed in 2002. For males and those in younger age brackets, a correlation between vulnerability to P. falciparum infection and strains with pfcrt 76K or strains with decreased numbers of mutations in pfdhfr and pfdhps was demonstrated. The parasites with pfcrt 76T exhibited a greater number of mutations in pfdhfr and pfdhps

Novel Mutations in K13 Propeller Gene of Artemisinin-Resistant Plasmodium falciparum

We looked for mutations in the Plasmodium falciparum K13 propeller gene of an artemisinin-resistant parasite on islands in Lake Victoria, Kenya, where transmission in 2012-2013 was high. The 4 new types of nonsynonymous, and 5 of synonymous, mutations we detected among 539 samples analyzed provide clues to understanding artemis- inin-resistant parasites