Species-specific identification of Leishmania in naturally infected sand flies captured in Mato Grosso do Sul State, Brazil

An increase in cutaneous and visceral leishmaniasis cases has been reported in recent years in the state of Mato Grosso do Sul, Brazil, and little is known to date about their etiological agents. An investigation into natural Leishmania infection of sand flies captured in this state between December 2003 and August 2004 was carried out. Mini-exon sequences were used as targets to identify Leishmania, and an RFLP technique was employed for those identified as belonging to the Viannia subgenus. Calculation of the minimal infection rate (MR) revealed that 1.6% of sand flies captured in the forest, peridomicile and intradomicile were positive. Six species were found to be infected by Leishmania (V.) braziliensis. Interestingly, two of the six species. Lutzomyia longipalpis and Nyssomyia whitmani, were captured in anthropic environments. The findings of this study constitute a useful tool for planning control measures against this disease in the State of Mato Grosso do Sul. (C) 2010 Elsevier B.V. All rights reserved.

Saturated Fatty Acids Produce an Inflammatory Response Predominantly through the Activation of TLR4 Signaling in Hypothalamus: Implications for the Pathogenesis of Obesity

In animal models of diet-induced obesity, the activation of an inflammatory response in the hypothalamus produces molecular and functional resistance to the anorexigenic hormones insulin and leptin. The primary events triggered by dietary fats that ultimately lead to hypothalamic cytokine expression and inflammatory signaling are unknown. Here, we test the hypothesis that dietary fats act through the activation of toll-like receptors 2/4 and endoplasmic reticulum stress to induce cytokine expression in the hypothalamus of rodents. According to our results, long-chain saturated fatty acids activate predominantly toll-like receptor 4 signaling, which determines not only the induction of local cytokine expression but also promotes endoplasmic reticulum stress. Rats fed on a monounsaturated fat-rich diet do not develop hypothalamic leptin resistance, whereas toll-like receptor 4 loss-of-function mutation and immunopharmacological inhibition of toll-like receptor 4 protects mice from diet-induced obesity. Thus, toll-like receptor 4 acts as a predominant molecular target for saturated fatty acids in the hypothalamus, triggering the intracellular signaling network that induces an inflammatory response, and determines the resistance to anorexigenic signals.

Molecular Mechanisms by Which Saturated Fatty Acids Modulate TNF-alpha Expression in Mouse Macrophage Lineage

Many macrophage functions are modulated by fatty acids (FAs), including cytokine release, such as tumor necrosis factor-alpha (TNF-alpha). TNF-alpha is of great interest due to its role in the inflammation process observed in several diseases such as rheumatoid arthritis, atherosclerosis, and obesity. However, the mechanisms by which FA effects occur have not been completely elucidated yet. In this study, we used a mouse monocyte lineage (J774 cells) to evaluate the effect of 50 and 100 mu M of saturated (palmitic and stearic acids), monounsaturated (oleic acid) and polyunsaturated (linoleic acid) FAs on TNF-alpha production. Alterations in gene expression, poly(A) tail length and activation of transcription factors were evaluated. Oleic and linoleic acids, usually known as neutral or pro-inflammatory FA, inhibited LPS-induced TNF-alpha secretion by the cells. Saturated FAs were potent inducers of TNF-alpha expression and secretion under basal and inflammatory conditions (in the presence of LPS). Although the effect of the saturated FA was similar, the mechanism involved in each case seem to be distinct, as palmitic acid increased EGR-1 and CREB binding activity and stearic acid increased mRNA poly(A) tail. These results may contribute to the understanding of the molecular mechanisms by which saturated FAs modulate the inflammatory response and may lead to design of associations of dietary and pharmacological strategies to counteract the pathological effects of TNF-alpha.

Saturated Fatty Acid-Induced Insulin Resistance Is Associated With Mitochondrial Dysfunction in Skeletal Muscle Cells

Increased plasma levels of free fatty acids (FFA) occur in states of insulin resistance such as obesity and type 2 diabetes mellitus. These high levels of plasma FFA are proposed to play an important role for the development of insulin resistance but the mechanisms involved are still unclear. This study investigated the effects of saturated and unsaturated FFA on insulin sensitivity in parallel with mitochondrial function. C2C12 myotubes were treated for 24 h with 0.1 mM of saturated (palmitic and stearic) and unsaturated (oleic, linoleic, eicosapentaenoic, and docosahexaenoic) FFA. After this period, basal and insulin-stimulated glucose metabolism and mitochondrial function were evaluated. Saturated palmitic and stearic acids decreased insulin-induced glycogen synthesis, glucose oxidation, and lactate production. Basal glucose oxidation was also reduced. Palmitic and stearic acids impaired mitochondrial function as demonstrated by decrease of both mitochondrial hyperpolarization and ATP generation. These FFA also decreased Akt activation by insulin. As opposed to saturated FFA, unsaturated FFA did not impair glucose metabolism and mitochondrial function. Primary cultures of rat skeletal muscle cells exhibited similar responses to saturated FFA as compared to C2C12 cells. These results show that in muscle cells saturated FFA-induced mitochondrial dysfunction associated with impaired insulin-induced glucose metabolism. J. Cell. Physiol. 222: 187-194, 2010. (C) 2009 Wiley-Liss, Inc.

A Phylogenetic Lineage of Closely Related Trypanosomes (Trypanosomatidae, Kinetoplastida) of Anurans and Sand Flies (Psychodidae, Diptera) Sharing the Same Ecotopes in Brazilian Amazonia

Analysis of the phylogenetic relationships among trypanosomes from vertebrates and invertebrates disclosed a new lineage of trypanosomes circulating among anurans and sand flies that share the same ecotopes in Brazilian Amazonia. This assemblage of closely related trypanosomes was determined by comparing whole SSU rDNA sequences of anuran trypanosomes from the Brazilian biomes of Amazonia, the Pantanal, and the Atlantic Forest and from Europe, North America, and Africa, and from trypanosomes of sand flies from Amazonia. Phylogenetic trees based on maximum likelihood and parsimony corroborated the positioning of all new anuran trypanosomes in the aquatic clade but did not support the monophyly of anuran trypanosomes. However, all analyses always supported four major clades (An01-04) of anuran trypanosomes. Clade An04 is composed of trypanosomes from exotic anurans. Isolates in clades An01 and An02 were from Brazilian frogs and toads captured in the three biomes studied, Amazonia, the Pantanal and the Atlantic Forest. Clade An01 contains mostly isolates from Hylidae whereas clade An02 comprises mostly isolates from Bufonidae; and clade An03 contains trypanosomes from sand flies and anurans of Bufonidae, Leptodactylidae, and Leiuperidae exclusively from Amazonia. To our knowledge, this is the first study describing morphological and growth features, and molecular phylogenetic affiliation of trypanosomes from anurans and phlebotomines, incriminating these flies as invertebrate hosts and probably also as important vectors of Amazonian terrestrial anuran trypanosomes.

Spatial and temporal patterns of occurrence of Lutzomyia sand fly species in an endemic area for cutaneous leishmaniasis in the Atlantic Forest region of northeast Brazil

Sand fly populations of different ecological niches in the Amaraji endemic American Cutaneous Leishmaniasis (ACL) focus of the Pernambuco Atlantic Forest region of northeastern Brazil were monitored spatiotemporally. Lutzomyia whitmani was dominant in all niches but occurred in smaller numbers in forested locations. L. whitmani was significantly less seasonal than the other species, being present throughout the year while other species were more abundant between February and April. These results suggest that L. whitmani may potentially be the principal vector of ACL in the region, even though the sand fly fauna was diverse: 88% were L.whitmani and 12% belonged to 11 other species. Two other species, L. complexa (1.3%) and L. migonei (0.8%), considered to be ACL vectors in other regions, were also present. This detailed picture of the sand fly population`s abundance and spatiotemporal distribution provides a basis for future modeling studies of forecasting sand fly activity patterns and ACL occurence.

Structural Characterization of Photopolymerizable Binary Liposomes Containing Diacetylenic and Saturated Phospholipids

The use of liposomes to encapsulate materials has received widespread attention for drug delivery, transfection, diagnostic reagent, and as immunoadjuvants. Phospholipid polymers form a new class of biomaterials with many potential applications in medicine and research. Of interest are polymeric phospholipids containing a diacetylene moiety along their acyl chain since these kinds of lipids can be polymerized by Ultra-Violet (UV) irradiation to form chains of covalently linked lipids in the bilayer. In particular the diacetylenic phosphatidylcholine 1,2-bis(10,12-tricosadiynoyl)- sn-glycero-3-phosphocholine (DC8,9PC) can form intermolecular cross-linking through the diacetylenic group to produce a conjugated polymer within the hydrocarbon region of the bilayer. As knowledge of liposome structures is certainly fundamental for system design improvement for new and better applications, this work focuses on the structural properties of polymerized DC8,9PC:1,2-dimyristoyl-sn-glycero-3-phusphocholine (DMPC) liposomes. Liposomes containing mixtures of DC8,9PC and DMPC, at different molar ratios, and exposed to different polymerization cycles, were studied through the analysis of the electron spin resonance (ESR) spectra of a spin label incorporated into the bilayer, and the calorimetric data obtained from differential scanning calorimetry (DSC) studies. Upon irradiation, if all lipids had been polymerized, no gel-fluid transition would be expected. However, even samples that went through 20 cycles of UV irradiation presented a DSC band, showing that around 80% of the DC8,9PC molecules were not polymerized. Both DSC and ESR indicated that the two different lipids scarcely mix at low temperatures, however few molecules of DMPC are present in DC8,9PC rich domains and vice versa. UV irradiation was found to affect the gel fluid transition of both DMPC and DC8,9PC rich regions, indicating the presence of polymeric units of DC8,9PC in both areas, A model explaining lipids rearrangement is proposed for this partially polymerized system.

Basic oxygen furnace slag as a treatment material for pathogens: Contribution of inactivation and attachment in virus attenuation

Basic oxygen furnace (BOF) slag media were studied as a potential treatment material in on-site sanitation systems. Batch and column studies were conducted to evaluate attenuation of the bacteriophage PR772 and 0.190 mu m diameter microspheres by BOF media, and to delineate the relative contributions of two principle processes of virus attenuation: inactivation and attachment. In the batch studies, conducted at 4 degrees C, substantial inactivation of PR772 did not occur in the pH 7.6 and 9.5 suspensions. At pH 11.4, bimodal inactivation of PR772 was observed, at an initial rate of 2.1 log C/C(0) day(-1) for the first two days, followed by a much slower rate of 0.124 log C/C(0) day(-1) over the following 10 days. Two column studies were conducted at 4 degrees C at a flow rate of 1 pore volume day(-1) using two slag sources (Stelco, Ontario; Tubarao, Brazil) combined with sand and pea gravel. In both column experiments, the effluent microsphere concentration approached input concentrations over time (reductions of 0.1-0.2 log C/C(0)), suggesting attachment processes for microspheres were negligible. Removal of PR772 virus was more pronounced both during the early stages of the experiments, but also after longer transport times (0.5-1.0 log C/C(0)). PR772 reduction appeared to be primarily as a result of virus inactivation in response to the elevated pH conditions generated by the BOF mixture (10.6-11.4). On-site sanitation systems using BOF media should be designed to maintain sufficient contact time between the BOF media and the wastewater to allow sufficient residence time of pathogens at elevated pH conditions. (C) 2009 Published by Elsevier Ltd.

Direct determination of iron in sand using solid sampling graphite furnace atomic absorption spectrometry

A fast and reliable method for the direct determination of iron in sand by solid sampling graphite furnace atomic absorption spectrometry was developed. A Zeeman-effect 3-field background corrector was used to decrease the sensitivity of spectrometer measurements. This strategy allowed working with up to 200 mu g of samples, thus improving the representativity. Using samples with small particle sizes (1-50 mu m) and adding 5 mu g Pd as chemical modifier, it was possible to obtain suitable calibration curves with aqueous reference solutions. The pyrolysis and atomization temperatures for the optimized heating program were 1400 and 2500 degrees C, respectively. The characteristic mass, based on integrated absorbance, was 56 pg, and the detection limits, calculated considering the variability of 20 consecutive measurements of platform inserted without sample was 32 pg. The accuracy of the procedure was checked with the analysis of two reference materials (IPT 62 and 63). The determined concentrations were in agreement with the recommended values (95% confidence level). Five sand samples were analyzed, and a good agreement (95% confidence level) was observed using the proposed method and conventional flame atomic absorption spectrometry. The relative standard deviations were lower than 25% (n = 5). The tube and boat platform lifetimes were around 1000 and 250 heating cycles, respectively.

Saturated Ceramides from the Sponge Dysidea robusta

Chemical investigation of the crude extract of a marine sponge Dysidea robusta led to the isolation of an inseparable mixture of saturated ceramides. These were identified from spectroscopic data as well as by hydrolysis followed by LC-MS analysis of the sphingosine moieties.