Chemical Mapping of <i>Leishmania</i> Infection in Live Cells by SERS Microscopy

We report the direct probing of the molecular composition of <i>Leishmania</i>-infected macrophage cells in vitro by surface-enhanced Raman scattering (SERS). The microscopic mapping data indicate local abundance and distribution of molecular species that are very characteristic of the infection and that are observed here simultaneously. As revealed by electron microscopy, the gold nanoprobes used for SERS microspectrosopy have access to the parasitophorous vacuoles (PV) through the endosomal system. SERS nanoprobes located in the direct proximity to the parasite, in the greater volume of the PV, and in endolysosomal compartments in other cellular regions, respectively, report a characteristic chemical composition for each respective location. The data enable assessment of the distribution of ergosterol and cholesterol in the amastigote stage of the parasite and its immediate surroundings in the vacuole. Proteophosphoglycans of parasite origin, an important hallmark of the infection, are identified throughout the PV

Suicides in the Nenets Autonomous Okrug, Russia.

This is a study of suicides in the Nenets Autonomous Okrug (NAO), a region with a large proportion of indigenous Nenets. To our knowledge, this is the first study investigating the problem of suicide in the indigenous and non-indigenous populations of the Russian Arctic. Our study aim was to assess suicide rates in the indigenous and non-indigenous populations of the NAO, as well as the socio-demographic characteristics, differences in suicide methods, seasonal variations, and the potential role of alcohol in suicides in these two populations. We conducted a retrospective, population-based mortality study of suicides in the NAO, using data from the autopsy reports of suicide victims in the region in 2002-2012. Socio-demographic data were obtained from passports and medical records, and then linked to total population data from the 2002 and 2010 censuses. Suicide rates for indigenous Nenets and the non-indigenous population were calculated according to different socio-demographic characteristics, and corresponding relative risks for these two populations were compared. Variations in suicide methods, seasonal variations, and variations in the day of the week suicides occurred in the NAO were compared with national data from the Russian Federal Statistics Service (Rosstat). Data on the presence of alcohol in the blood and blood alcohol content in suicide cases from the NAO were compared with data from the neighboring Arkhangelsk Oblast. Suicide rates in the NAO were higher than corresponding national figures. Suicide rates were higher among the indigenous Nenets than the non-indigenous population, and were associated with different socio-demographic characteristics. We showed different relative frequencies of suicide by hanging, cutting, and firearm, as well as differences in suicide occurrence by month and day of the week in the NAO when compared with Russia as a whole. The study results and conclusions may be useful to create suicide prevention programs that are targeted to different population groups in the Russian Arctic. Key words: Nenets Autonomous Okrug (NAO), Arkhangelsk Oblast (AO), suicide rates, relative risks, person-years, indigenous Nenets, suicide methods, seasonality, alcohol

Establishment of a resazurin-based <i>G. duodenalis</i> growth assay.

<p>Various numbers of trophozoites (WB-C6 strain, 0.5×10³–10⁴ trophozoites per well) were seeded into 96-well plates. After 24 h, 48 h or 72 h of culture, the medium was replaced by PBS and resazurin solution was added. Trophozoites were further incubated for 4 h and the fluorescence intensity was measured fluorometrically using an excitation wavelength of 550 nm and an emission wavelength of 595 nm. Results are presented as fluorescence intensity (RFU) in relation to the initially inoculated number of trophozoites; Data are expressed as mean ± SD of three independent experiments. The correlation factor (r²) was calculated by linear regression using the software program Prism 5.01 (GraphPad).</p

No interaction effect of MTZ and orlistat in combination on the growth of <i>G. duodenalis</i> trophozoites.

<p>Trophozoites of <i>G. duodenalis</i>, (A) WB-C6 (n = 4) and (B) 14-03/F7 (n = 3), were incubated in the presence of various concentrations of MTZ (0.33–81 µM) or orlistat (0.33–27 µM) alone or in combination. Combination indices (CIs) of MTZ and orlistat were calculated for given drug ratios (MTZ:orlistat) of 1∶1 (filled circle), 3∶1 (open squares), or 1∶3 (open triangle). Results are plotted against the fraction of parasites affected (normalized growth inhibition). Data are presented as mean ± SD of independent experiments (each performed in triplicates). CI values between 0.8–1.2 (shaded area) indicate no interaction of the drugs regarding growth inhibition (i.e. drugs act independently).</p

Orlistat inhibits growth of <i>G. duodenalis</i> trophozoites.

<p><b><i> </i></b> Anti-<i>G. duodenalis</i> activity of MTZ (filled circle) and orlistat (blank squares) was determined in a resazurin assay and results are presented as relative percentage of growth of the untreated trophozoites as a control. In addition, the half maximal inhibitory concentrations (IC50_24h) are shown. (A) Growth inhibition curve of the laboratory strain WB-C6; the IC50_24h of orlistat (IC50_24h = 4.3 µM; n = 7) was significantly lower than the IC50_24h of MTZ (IC50_24h = 11.0 µM; n = 10), p = 0.0034. (B) Growth inhibition curve of the <i>G. duodenalis</i> patient isolate 14-03/F7; the IC50_24h of orlistat (IC50_24h = of 2.8 µM, n = 4) was significantly lower than the IC50 of MTZ (IC50_24h = 6.2 µM; n = 5), p = 0.0079. Data are expressed as mean ± SD. The Mann-Whitney-test was used to evaluate the statistical significance of the differences in IC50 values.</p

Scanning electron microscopy of orlistat-treated <i>Giardia duodenalis</i> trophozoites (WB-C6 strain) showing altered morphology.

<p>(A) Solvent control (DMSO, 0.1% in medium). (B) MTZ, 11 µM (C) Solvent control (ethanol, 0.12% in medium). (D, E, F) Orlistat, 5 µM. Controls (A, C) reveal the typical appearance of trophozoites with their pear-shaped cell body and the characteristic arrangement of the eight flagella. MTZ treated trophozoites (B) possess shorter flagella than the corresponding control trophozoites (A). Orlistat treated trophozoites (D, E, F) appear shrunken in comparison to corresponding control trophozoites (C) and show blebs on their dorsal surface (arrowheads) as well as at the tip of the flagella (arrow) (D). (E, F) In some of the trophozoites orlistat mediated effects (blebs at flagella and dorsal cell surface) are more prominent. Scale bar = 5 µm.</p

Heterologous expression of the filarial nematode gene products reveals their potential to inhibit immune function-1

<p><b>Copyright information:</b></p><p>Taken from "Heterologous expression of the filarial nematode gene products reveals their potential to inhibit immune function"</p><p>BMC Biology 2005;3():8-8.</p><p>Published online 23 Mar 2005</p><p>PMCID:PMC555940.</p><p>Copyright © 2005 Gomez-Escobar et al; licensee BioMed Central Ltd.</p>ith anti-serum or anti-ALT-1 antibody. Cloned transfected and transfected fixed parasites were incubated with an anti-ALT-1 antibody, which binds to both ALT-1 and ALT-2, and detected with an anti-mouse-FITC secondary antibody. FACS analysis of transfected promastigotes of . Fixed promastigotes were stained, without permeabilization, with anti-ALT-1 antibody and analysed by FACS. -transfected ; -transfected ; wild-type . Positive cells are those within the gate (M1) set above the threshold defined by reactivity of transfected parasites stained with normal mouse serum . Multiplication of transfectants within macrophages. -transformed wild-type amastigotes or transfected amastigotes were added to bone marrow-derived macrophages from CBA mice at a ratio of 10 parasites per macrophage. After 24 hours at 34°C the infected macrophages were observed by microscopy

Primer pairs utilized for characterization of the entire <i>FCN2</i> gene in 40 Syrian Arabs.

*<p>used only for sequencing.</p

Haploview plot illustrating the linkage disequilibrium of the <i>FCN2</i> functional variants in controls (2A) and patients (2B).

<p>At the top the SNPs are shown according to their succession from the start of translation of the <i>FCN2</i> gene. <i>Empty squares</i> indicate a high degree of LD (LD coefficient D′ = 1) between pairs of markers. Numbers indicate the D′ value expressed as a percentile. <i>Red squares</i> indicate pairs in strong LD with LOD scores for LD ≥70; purple squares, D′ = 1 with LOD >1; <i>white squares</i>, D′ <1.0 and LOD ≤2.</p

<i>FCN2</i> haplotypes in Cutaneous Leishmaniasis patients and healthy controls.

<p><i>FCN2</i> haplotypes in Cutaneous Leishmaniasis patients and healthy controls.</p