Parasite and egg burden, hepatic collagen and histologic pattern of liver granulomas in selection III high and low antibody responder mice infected with Schistosoma mansoni

Selection III mice have particular immunological characteristics: they are high (H III) or low (L III) antibody producer animals, yet both lines display similar T cell responses and macrophage activities. We submittedthese mice to infection with Schistosoma mansoni to assess in vivo parasite and egg burden, hepatic collagen and cellular composition of granulomas in both lines. Titration of anti-Schistosoma IgG by ELISA showed remarkably higher values inH III line, at both studied periods (8th and 12th weeks post-infection). Nevertheless, the number of adult worms recovered from the portal system was similar inboth lines, being not associated with anti-Schistosoma antibody levels. There isan increase in hepatic collagen from the 8th to the 12th weeks post-infection, which is paralleled by an increase in the number of eggs in the liver. This association apparently occurs at the same radio in H III and L III animals. The most important difference found between the two lines was the outstanding contrast interms of volume and eosinophil counts in the granulomas, with lesions from H IIImice clearly being larger and containing more of these cells than LIII lesions.

Genetic Variability and Microdistribution of Triatoma infestans Genotypes and Trypanosoma cruzi Clones in Arequipa Region (Peru)

The genetic variability of Triatoma infestans and Trypanosoma cruzi populations was studied by isoenzyme analysis in two distinct areas of Arequipa province (Peru); one, Santa Rita de Siguas, being an endemic area for Chagas' disease, the second, Arequipa, recently infected. Analysis of T. infestans genetic variability indicates, (i) temporal stability of genotypes found in Santa Rita de Siguas, (ii) high genetic differences between Arequipa and Santa Rita de Siguas populations suggesting minor contact between them, (iii) multiple origin of the T. infestans population in Arequipa, and (iv) poor dispersal capacity of T. infestans: the panmictic unit could be reduce to a house. Parasite isoenzyme analysis was performed in 29 Peruvian stocks of T. cruzi, mainly isolated from bugs taken in a single locality, Santa Rita de Siguas. The results show, (i) a high genetic polymorphism, (ii) nine different multilocus genotypes were detected and clustered in two different clades, (iii) most of the parasite isolates pertained to one of the clade and were genetically similar to those analyzed 12 years before. This sample allowed the study of the mating system of T. cruzi in strict sympatic conditions and gave more strength to the hypothesis of the clonal structure of T. cruzi populations

Haemolymph and Fat Body Metallo-protease Associated with Enterobacter cloacae Infection in the Bloodsucking Insect, Rhodnius prolixus

Analysis of zymograms with SDS-polyacrilamide gel electrophoresis containing gelatin as substrate, and performed on samples of haemolymph or fat body taken from Rhodnius prolixus inoculated or not with Enterobacter cloacae, demonstrated distinct patterns of protease activities: (i) in the haemolymph two proteases were induced in insects inoculated with bacteria; (ii) two proteases were detected in the fat bodies derived from non-inoculated controls or insect inoculated with sterile culture medium; (iii) haemolymph and fat body had both the same apparent molecular weights proteases (46 and 56 kDa); and (iv) these enzymes were characterized as metallo-proteases. The association of these enzymes in Rhodnius infected with bacteria was discussed.

Seroprevalence of Toxoplasma gondii infection in goats by the indirect haemagglutination, immunofluorescence and immunoenzymatic tests in the region of Uberlândia, Brazil

A comparative study of the indirect haemagglutination (IHA), immunofluorescence (IFAT) and immunoenzymatic (ELISA) tests was carried out to determine the prevalence of Toxoplasma gondii antibodies in goats. One hundred seventy-four serum samples were obtained from four goat herds from the region of Uberlândia, State of Minas Gerais. The distribution of the animals, according to their origin, was as follow: 71 from herd I; 39 from herd II; 37 from herd III; and 27 from herd IV. Serum samples were analyzed by IHA, IFAT and ELISA, considering the reactivity of the serum samples at dilution ≥ 1:64 as cut off titer for the three tests. A global seroprevalence of 18.4% was observed, with significantly higher positivity rate in the herd II (66.7%) and older animals (> 36 months). A high and significant positive correlation was found between the titers obtained by the IHA versus IFAT, IHA versus ELISA, and ELISA versus IFAT. Therefore, it can be concluded that the three analyzed tests have shown to be highly concordant and appropriate for epidemiological surveys of Toxoplasma infection in goats. Although the seroprevalence of T. gondii infection in goats is relatively low in this region as compared to other regions of the country, adequate management might be useful and essential to control the infection in the goat herds.

Life cycle, feeding and defecation patterns of Rhodnius ecuadoriensis (Lent & León 1958) (Hemiptera: Reduviidae: Triatominae) under laboratory conditions

Rhodnius ecuadoriensis is the second most important vector of Chagas Disease (CD) in Ecuador. The objective of this study was to describe (and compare) the life cycle, the feeding and defecation patterns under laboratory conditions of two populations of this specie [from the provinces of Manabí (Coastal region) and Loja (Andean region)]. Egg-to-adult (n = 57) development took an average of 189.9 ± 20 (Manabí) and 181.3 ± 6.4 days (Loja). Mortality rates were high among Lojan nymphs. Pre-feeding time (from contact with host to feeding initiation) ranged from 4 min 42 s [nymph I (NI)] to 8 min 30 s (male); feeding time ranged from 14 min 45 s (NI)-28 min 25 s (male) (Manabí) and from 15 min 25 s (NI)-28 min 57 s (nymph V) (Loja). The amount of blood ingested increased significantly with instar and was larger for Manabí specimens (p < 0.001). Defecation while feeding was observed in Manabí specimens from stage nymph III and in Lojan bugs from stage nymph IV. There was a gradual, age-related increase in the frequency of this behaviour in both populations. Our results suggest that R. ecuadoriensis has the bionomic traits of an efficient vector of Trypanosoma cruzi. Together with previous data on the capacity of this species to infest rural households, these results indicate that control of synanthropic R. ecuadoriensis populations in the coastal and Andean regions may have a significant impact for CD control in Ecuador and Northern Peru.

The utility of anti-trypomastigote lytic antibodies for determining cure of Trypanosoma cruzi infections in treated patients: an overview and perspectives

In previous work, we proposed alternative protocols for following patients with treated Chagas disease and these are reviewed herein. Evidence was provided to support the following: (i) functional anti-trypomastigote antibodies are indicative of ongoing chronic Trypanosoma cruzi infections; (ii) specific antibodies detected by conventional serology (CS) with epimastigote extracts, fixed trypomastigotes or other parasite antigens may circulate years after parasite elimination; (iii) functional antibodies are evidenced by complement-mediated lysis of freshly isolated trypomastigotes, a test which is 100% specific, highly sensitive, and the first to revert after T. cruzi elimination and (iv) the parasite target for the lytic antibodies is a glycoprotein of high molecular weight (gp160) anchored at the parasite surface. The complement regulatory protein has been cloned, sequenced and produced as a recombinant protein by other groups and is useful for identifying functional anti-T. cruzi antibodies in ELISA tests, thus dispensing with the need for live trypomastigotes to manage treated patients. If used instead of CS to define cures for Chagas patients, ELISA will avoid unnecessary delays in finding anti-T. cruzi drugs. Other highly sensitive techniques for parasite DNA detection, such as PCR, need to be standardized and included in future protocols for the management of patients with drug-treated Chagas disease.

Expression of bacterial virulence factors and cytokines during in vitro macrophage infection by enteroinvasive Escherichia coli and Shigella flexneri: a comparative study

Enteroinvasive Escherichia coli (EIEC) and Shigellaspp cause bacillary dysentery in humans by invading and multiplying within epithelial cells of the colonic mucosa. Although EIEC and Shigellashare many genetic and biochemical similarities, the illness caused by Shigellais more severe. Thus, genomic and structure-function molecular studies on the biological interactions of these invasive enterobacteria with eukaryotic cells have focused on Shigella rather than EIEC. Here we comparatively studied the interactions of EIEC and of Shigella flexneriwith cultured J774 macrophage-like cells. We evaluated several phenotypes: (i) bacterial escape from macrophages after phagocytosis, (ii) macrophage death induced by EIEC and S. flexneri, (iii) macrophage cytokine expression in response to infection and (iv) expression of plasmidial (pINV) virulence genes. The results showed thatS. flexneri caused macrophage killing earlier and more intensely than EIEC. Both pathogens induced significant macrophage production of TNF, IL-1 and IL-10 after 7 h of infection. Transcription levels of the gene invasion plasmid antigen-C were lower in EIEC than in S. flexneri throughout the course of the infection; this could explain the diminished virulence of EIEC compared to S. flexneri.

Pharmacodynamics of mefloquine and praziquantel combination therapy in mice harbouring juvenile and adult Schistosoma mansoni

Praziquantel (PZQ) is currently the only drug widely used for the treatment of schistosomiasis, but the antimalarial drug mefloquine (Mef) possesses interesting antischistosomal properties. Combination therapy with these two drugs has been suggested as a strategy for transmission control, as PZQ is active against adult worms and Mef is active against schistosomula. To examine the efficacy of combination therapy, Schistosoma mansoni-reinfected mice were separated into seven groups: untreated (I), treated with PZQ in doses of 200 mg/kg (II) or 1,000 mg/kg (III), treated with Mef in doses of 200 mg/kg (IV) or 400 mg/kg (V); each dose was divided equally and given on two consecutive days. Group VI was treated with doses of PZQ + Mef as in groups II and IV, respectively, while group VII was treated with PZQ + Mef as in groups III and V, respectively. PZQ + Mef at the reduced doses of 200 mg/kg each enhanced the therapeutic efficacy over the reduced PZQ dose alone as shown by a very high reduction in the total numbers of mature worms (95% vs. 49%), immature worms (96% vs. 29%) and the complete eradication of immature females, mature females and immature eggs. The reduction in worm burden was associated with the healing of hepatic granulomatous lesions and the normalisation of all liver enzymes. Therefore, the use of Mef with PZQ is more effective than PZQ alone and should be considered for clinical trials in humans as a potential treatment regimen to prevent treatment failures in areas with high rates of schistosomiasis.

Chagas disease: control, elimination and eradication. Is it possible?

From an epidemiological point of view, Chagas disease and its reservoirs and vectors can present the following characteristics: (i) enzooty, maintained by wild animals and vectors, with broad occurrence from southern United States of America (USA) to southern Argentina and Chile (42ºN 49ºS), (ii) anthropozoonosis, when man invades the wild ecotope and becomes infected with Trypanosoma cruzi from wild animals or vectors or when the vectors and wild animals, especially marsupials, invade the human domicile and infect man, (iii) zoonosis-amphixenosis and exchanged infection between animals and humans by domestic vectors in endemic areas and (iv) zooanthroponosis, infection that is transmitted from man to animals, by means of domestic vectors, which is the rarest situation in areas endemic for Chagas disease. The characteristics of Chagas disease as an enzooty of wild animals and as an anthropozoonosis are seen most frequently in the Brazilian Amazon and in the Pan-Amazon region as a whole, where there are 33 species of six genera of wild animals: Marsupialia, Chiroptera, Rodentia, Edentata (Xenarthra), Carnivora and Primata and 27 species of triatomines, most of which infected with T. cruzi . These conditions place the resident populations of this area or its visitors - tourists, hunters, fishermen and especially the people whose livelihood involves plant extraction - at risk of being affected by Chagas disease. On the other hand, there has been an exponential increase in the acute cases of Chagas disease in that region through oral transmission of T. cruzi , causing outbreaks of the disease. In four seroepidemiological surveys that were carried out in areas of the microregion of the Negro River, state of Amazonas, in 1991, 1993, 1997 and 2010, we found large numbers of people who were serologically positive for T. cruzi infection. The majority of them and/or their relatives worked in piassava extraction and had come into contact with and were stung by wild triatomines in that area. Finally, a characteristic that is greatly in evidence currently is the migration of people with Chagas disease from endemic areas of Latin America to non-endemic countries. This has created a new dilemma for these countries: the risk of transmission through blood transfusion and the onus of controlling donors and treating migrants with the disease. As an enzooty of wild animals and vectors, and as an anthropozoonosis, Chagas disease cannot be eradicated, but it must be controlled by transmission elimination to man.

Variability of root traits in common bean genotypes at different levels of phosphorus supply and ontogenetic stages

Selection of common bean (Phaseolus vulgaris L.) cultivars with enhanced root growth would be a strategy for increasing P uptake and grain yield in tropical soils, but the strong plasticity of root traits may compromise their inclusion in breeding programs. The aim of this study was to evaluate the magnitude of the genotypic variability of root traits in common bean plants at two ontogenetic stages and two soil P levels. Twenty-four common bean genotypes, comprising the four growth habits that exist in the species and two wild genotypes, were grown in 4 kg pots at two levels of applied P (20 and 80 mg kg-1) and harvested at the stages of pod setting and early pod filling. Root area and root length were measured by digital image analysis. Significant genotype × P level and genotype × harvest interactions in analysis of variance indicate that the genotypic variation of root traits depended on soil nutrient availability and the stage at which evaluation was made. Genotypes differed for taproot mass, basal and lateral root mass, root area and root length at both P levels and growth stages; differences in specific root area and length were small. Genotypes with growth habits II (upright indeterminate) and III (prostrate indeterminate) showed better adaptation to limited P supply than genotypes of groups I (determinate) and IV (indeterminate climbing). Between the two harvests, genotypes of groups II and III increased the mass of basal and lateral roots by 40 and 50 %, respectively, whereas genotypes of groups I and IV by only 7 and 19 %. Values of the genotypic coefficient of determination, which estimates the proportion of phenotypic variance resulting from genetic effects, were higher at early pod filling than at pod setting. Correlations between shoot mass and root mass, which could indicate indirect selection of root systems via aboveground biomass, were higher at early pod filling than at pod setting. The results indicate that selection for root traits in common bean genotypes should preferentially be performed at the early pod-filling stage.

Antioxidant compoundsand total antioxidant activity in fruits of acerola from cv. Flor Branca, Florida Sweet and BRS 366

Information on antioxidant properties at different ontological stages may help producers and food technologists to identify which cultivar and/or maturity stage are most adequate for their need, therefore this work aimed to study the changes in the antioxidant metabolism during acerola development. Fruit from cv. Flor Branca, BRS366 and Florida Sweet were harvested at different stages: immature green colored (I), physiologically mature with green color and maximum size (II), breaker (III) and full red ripe (IV). After harvest, fruits were selected, divided into four replications with 500 g each and evaluated regarding their titratable acidity, pH, soluble solids, total soluble sugar, vitamin C, polyphenol, anthocyanin, yellow flavonoid, total antioxidant activity and antioxidant enzyme activity. Anthocyanin and flavonoid were determined through LC-DAD-ESI/MS and all analysis followed a completely randomized factorial 3 x 4 design. Fruits of 'Florida Sweet' presented significantly higher soluble solids (9.46ºBrix). Vitamin C content decreased during ripening, but ripe 'BRS 366' fruits showed the greatest values (1363 mg.100 g-1) and highest TAA with 42.36 µM TEAC.g-1FW. Cyanidin 3-rhamnoside (520.76 mg.100 g-1 DM) and quercetin 3-rhamnoside (33.72 mg.100 g-1 DM) were the most abundant anthocyanin and yellow flavonoids found mainly in 'Flor Branca' fruit of acerola, whose antioxidant enzymes activities were also higher. Ripe 'Florida Sweet' fruit presents a great potential for fresh consumption, meanwhile physiologically mature 'BRS 366' fruit seems the best option for the bioactive compounds processing industry. As 'Flor Branca' fruit of acerola kept the highest activity levels, it could be an indicative of greater potential for postharvest conservation.

Determinação indireta de N-acetil-L-cisteína por injeção em fluxo empregando Ce(IV) e ferroína

An indirect flow injection spectrophotometric procedure is proposed for the determination of N-acetyl-L-cysteine in pharmaceutical formulations. In this system, ferroin ([Fe(II)-(fen)2]2+) in excess, with a strong absorption at 500 nm, is oxidized by cerium(IV) yielding cerium(III) and [Fe(III)-(fen)2]3+ (colorless), thus producing a baseline. When N-acetyl-L-cysteine solution is introduced into the flow injection system, it reacts with cerium(IV) increasing the analytical signal in proportion to the drug concentration. Under optimal experimental conditions, the linearity of the analytical curve for N-acetyl-L-cysteine ranged from 6.5x10-6 to 1.3x10-4 mol L-1. The detection limit was 5.0x10-6 mol L-1and recoveries between 98.0 and 106% were obtained. The sampling frequency was 60 determinations per hour and the RSD was smaller than 1.4% for 2.2x10-5 mol L-1 N-acetyl-L-cysteine.

Myoglobins: the link between discoloration and lipid oxidation in muscle and meat

Aerobic metabolism changes rapidly to glycolysis post-mortem resulting in a pH-decrease during the transformation of muscle in to meat affecting ligand binding and redox potential of the heme iron in myoglobin, the meat pigment. The "inorganic chemistry" of meat involves (i) redox-cycling between iron(II), iron(III), and iron(IV)/protein radicals; (ii) ligand exchange processes; and (iii) spin-equilibra with a change in coordination number for the heme iron. In addition to the function of myoglobin for oxygen storage, new physiological roles of myoglobin are currently being discovered, which notably find close parallels in the processes in fresh meat and nitrite-cured meat products. Myoglobin may be characterized as a bioreactor for small molecules like O2, NO, CO, CO2, H2O, and HNO with importance in bio-regulation and in protection against oxidative stress in vivo otherwise affecting lipids in membranes. Many of these processes may be recognised as colour changes in fresh meat and cured meat products under different atmospheric conditions, and could also be instructive for teaching purposes.

Pré-concentração de Cr(III) e análise de especiação de cromo empregando montmorilonita saturada com íons potássio

The clay mineral montmorillonite-K10 (MT), treated under acidic medium and saturated with potassium ions (MTK), was employed in sorption and desorption studies aiming the preconcentration of Cr(III) and the speciation analysis of chromium. The sorption process of Cr(III) was close to 100%, suggesting that MTK was a good material for Cr(III) preconcentration, although, the maximum recovery in HNO3 solution was near 89%. On the other hand, Cr(VI) practically was not retained in MTK, suggesting this material as an appropriate mineral phase to be used in speciation analysis of chromium in aqueous medium.

Preparative separation of polyphenols from artichoke by polyamide column chromatography and high-speed counter-current chromatography

An efficient method for the rapid separation and purification of polyphenols from artichoke by polyamide column chromatography in combination with high-speed counter-current chromatography (HSCCC) was successfully built. The crude ethanol extracts from dry artichoke were first pre-separated by polyamide column chromatography and divided in two parts as sample 1 and sample 2. Then, the samples were further separated by HSCCC and yielded 7.8 mg of chlorogenic acid (compound I), 24.5 mg of luteolin-7-O-β-D-rutinoside (compound II), 18.4 mg of luteolin-7-O-β-D-glucoside (compound III), and 33.4 mg of cynarin (compound IV) with purity levels of 92.0%, 98.2%, 98.5%, and 98.0%, respectively, as determined by high-performance liquid chromatography (HPLC) method. The chemical structures of these compounds were identified by electrospray ionization-mass spectrometry (ESI-MS) and nuclear magnetic resonance (NMR).