Technical assessment of the affymetrix yeast expression GeneChip YE6100 platform in a heterologous model of genes that confer resistance to antimalarial drugs in yeast. (73/2261)

The advent of high-density gene array technology has revolutionized approaches to drug design, development, and characterization. At the laboratory level, the efficient, consistent, and dependable exploitation of this complex technology requires the stringent standardization of protocols and data analysis platforms. The Affymetrix YE6100 expression GeneChip platform was evaluated for its performance in the analysis of both global (6,000 yeast genes) and targeted (three pleiotropic multidrug resistance genes of the ATP binding cassette transporter family) gene expression in a heterologous yeast model system in the presence and absence of the antimalarial drug chloroquine. Critical to the generation of consistent data from this platform are issues involving the preparation of the specimen, use of appropriate controls, accurate assessment of experiment variance, strict adherence to optimized enzymatic and hybridization protocols, and use of sophisticated bioinformatics tools for data analysis.  (+info)

Presence of a Na(+)/H(+) exchanger in acidocalcisomes of Leishmania donovani and their alkalization by anti-leishmanial drugs. (74/2261)

Acidocalcisomes are acidic vacuoles present in trypanosomatids that contain most of the cellular calcium. The data presented here demonstrate that Leishmania donovani acidocalcisomes possess a Na(+)/H(+) exchanger. 3,5-Dibutyl-4-hydroxytoluene, in the concentration range of 0-20 microM, inhibited the Na(+)/H(+) exchanger, and strongly stimulated the activity of the vacuolar H(+)-ATPase responsible for vacuolar acidification. As occurs with Na(+), the cationic anti-leishmanial drugs pentamidine, WR-6026, and chloroquine promoted a fast and extensive alkalization of the L. donovani acidocalcisomes.  (+info)

Economic impact of febrile morbidity and use of permethrin-impregnated bed-nets in a malarious area I. Study of demographics, morbidity, and household expenditures associated with febrile morbidity in the Republic of Benin. (75/2261)

In preparation for a study on the effect of bed net use on malaria, this article describes febrile morbidity and malaria expenditures in a sub-Saharan area (Benin) of hyperendemic malaria. The 325 randomly selected households were visited weekly between April 1994 and March 1995 to determine febrile morbidity and household expenditures for prevention and treatment. The results indicate that rural children had two febrile episodes annually compared with 0.3 episodes among children living in the city. There was no difference in mean annual febrile episodes between adults and children (adults = 1.5, children = 1.5; P = 0.48) and in the expenditures per febrile episode (adults = US$1.85, children = US$1.62; P = 0.45). Annual prevention expenditures were higher for adults than for children (US$1.73 and US$1.28, respectively; P < 0.001), although there was no significant difference in expenditures for annual treatment for adults and children (US$2.15 and US$2.34, respectively). These and other findings are analyzed further and discussed.  (+info)

Economic impact of febrile morbidity and use of permethrin-impregnated bed nets in a malarious area II. Determinants of febrile episodes and the cost of their treatment and malaria prevention. (76/2261)

The objective of this study is to determine the effect of permethrin insecticide-treated bed net (PITN) use on the incidence of febrile episodes and on household malaria expenses in Benin. Over the course of one year, 208 randomly selected PITN user and non-user households were visited weekly to determine expenditures on febrile morbidity and its treatment, and to monitor spending on malaria prevention. Multivariate analyses were performed to distinguish the effects of PITN use from other important determinants of morbidity, such as malaria-related beliefs and practices, income, and other socio-economic variables. Results from the logistic regression analysis show that the use of PITNs decreases the risk of febrile episodes by 34% in children living in the rural zone. Multiple regression analysis reveals that PITN use does not reduce prevention and treatment expenses. These expenses are significantly associated with women's income. This report also discusses other factors associated with febrile morbidity and malaria-related expenditures.  (+info)

Chloroquine-resistant isolates of Plasmodium falciparum with alternative CG2 omega repeat length polymorphisms. (77/2261)

A particular polymorphism in the cg2 gene has previously been linked to chloroquine resistance in reference isolates of Plasmodium falciparum. To assess the association of this polymorphism with chloroquine resistance in field specimens of P. falciparum, we analyzed the omega repeat region of the cg2 gene in 47 isolates of P. falciparum collected in the Ingwavuma District of northern KwaZulu-Natal, South Africa. Polymerase chain reaction (PCR) primers, which were designed to amplify the region of DNA surrounding the omega repeat, were used to obtain omega repeat PCR products from the field isolates. The PCR product for each isolate varied in length, depending on the number of cg2 omega repeats for that isolate. We found that several in vivo and in vitro chloroquine-resistant isolates of P. falciparum did not have the expected 16 omega repeats. These results suggest that the link between the cg2 polymorphism and chloroquine resistance identified previously may not apply in all malarious areas.  (+info)

Gametocytemia and infectivity to mosquitoes of patients with uncomplicated Plasmodium falciparum malaria attacks treated with chloroquine or sulfadoxine plus pyrimethamine. (78/2261)

Plasmodium falciparum gametocytemia and its related infectivity for mosquitoes was studied in 115 patients (median age = 18 years, range = 4-45) with simple malaria attacks who lived in the hypoendemic area of Dakar, Senegal. Patients were included in a 28-day in vivo sensitivity test after treatment with chloroquine (CQ, n = 82) or sulfadoxine plus pyrimethamine (SP, n = 33). The prevalence of resistant infections was 58.5% in those treated with CQ and 0% in those treated with SP. The gametocytemia peaked at day 7 after treatment. The maximal gametocyte prevalence was 38.2% in the CQ-sensitive infection group, 89.6% in the CQ-resistant group, and 97.0% in those treated with SP The maximal geometric mean gametocytemia was 2.19/microl in the CQ-sensitive infection group, 29.12/microl in the CQ-resistant group and 85.55/microl in those treated with SP. The period between appearance of the first clinical symptom and treatment was positively related to gametocyte prevalence at days 0 and 2. Experimental infection of wild Anopheles arabiensis using membrane feeders was performed at days 0 and 7, and mosquito infectivity was measured by oocyst detection on the midgut. At day 0, 14.1% of the patients had infected at least 1 mosquito, and at day 7, this value was 38.5%. The mean percentage of infected mosquitoes was 3.2% at day 0 and 12.6% at day 7. At day 7 after treatment with CQ, the relative risk for patients with resistant infections of infecting anophelines was 4.07 higher than in those with sensitive infections. No difference was observed in infectivity for mosquitoes between RI-type resistance and the RII + RIII-type resistance. A sporonticidal effect of SP was observed at day 7 after treatment. These data show that P. falciparum gametocytes and their infectivity for mosquitoes were differentiated according to the drug used, its efficacy, and the duration of symptoms before treatment; they were not dependent on the density of asexual stages. Prompt treatment of malaria cases performed at the beginning of symptoms could limit the spread of resistant parasites.  (+info)

Shigella flexneri IpaH(7.8) facilitates escape of virulent bacteria from the endocytic vacuoles of mouse and human macrophages. (79/2261)

The behavior of Shigella flexneri ipaH mutants was studied in human monocyte-derived macrophages (HMDM), in 1-day-old human monocytes, and in J774 mouse macrophage cell line. In HMDM, strain pWR700, an ipaH(7.8) deletion mutant of S. flexneri 2a strain 2457T, behaved like the wild-type strain 2457T. This strain caused rapid host cell death by oncosis, and few bacterial CFU were recovered after incubation in the presence of gentamicin as previously described for 2457T-infected HMDM. However, analysis of bacterial compartmentalization within endocytic vacuoles with gentamicin and chloroquine indicated that more pWR700 than 2457T was present within the endocytic vacuoles of HMDM, suggesting that ipaH(7.8) deletion mutant transited more slowly from the vacuoles to the cytoplasm. In contrast to findings with HMDM, CFU recovered from pWR700-infected mouse J774 cells were 2 to 3 logs higher than CFU from 2457T-infected J774 cells. These values exceeded CFU recovered after infection of J774 cells with plasmid-cured avirulent strain M4243A1. Incubation with gentamicin and chloroquine clearly showed that pWR700 within J774 cells was mostly present within the endocytic vacuoles. This distribution pattern was similar to that seen with M4243A1 and contrasted with the pattern seen with 2457T. Complementation of pWR700 with a recombinant clone expressing ipaH(7. 8) restored the intracellular distribution of bacteria to that seen with the wild-type strain. Strains with deletions in ipaH(4.5) or ipaH(9.8), however, behaved like 2457T in both HMDM and J774 cells. The distribution profile of pWR700 in 1-day-old monocytes was similar to that seen in J774 cells. Like infected J774 cells, 1-day-old human monocytes demonstrated apoptosis upon infection with virulent Shigella. These results suggest that a role of the ipaH(7. 8) gene product is to facilitate the escape of the virulent bacteria from the phagocytic vacuole of monocytes and macrophages.  (+info)

Perlecan heparan sulfate proteoglycan: a novel receptor that mediates a distinct pathway for ligand catabolism. (80/2261)

Cell surface heparan sulfate proteoglycans (HSPGs) participate in the catabolism of many physiologically important ligands. We previously reported that syndecan HSPGs directly mediate endocytosis, independent of coated pits. We now studied perlecan, a major cell surface HSPG genetically distinct from syndecans. Cells expressing perlecan but no other proteoglycans bound, internalized, and degraded atherogenic lipoproteins enriched in lipoprotein lipase. Binding was blocked by heparitinase, and degradation by chloroquine. Antibodies against beta(1) integrins reduced initial ligand binding, consistent with their roles as cell surface attachment sites for perlecan. By several criteria, catabolism via perlecan was distinct from either coated pits or the syndecan pathway. The kinetics of internalization (t(12) = 6 h) and degradation (t(12) approximately 18 h) were remarkably slow, unlike the other pathways. Blockade of the low density lipoprotein receptor-related protein did not slow perlecan-dependent internalization. Internalization via perlecan was inhibited by genistein but unaffected by cytochalasin D, a pattern distinct from coated pits or syndecan-mediated endocytosis. Finally, we examined cooperation between perlecan and low density lipoprotein receptors and found limited synergy. Our results demonstrate that perlecan mediates internalization and lysosomal delivery that is kinetically and biochemically distinct from other known uptake pathways and is consistent with a very slow component of HSPG-dependent ligand processing found in vitro and in vivo.  (+info)