Effect of hypoxia on the activity and binding of glycolytic and associated enzymes in sea scorpion tissues

The effect of hypoxia on the levels of glycogen, glucose and lactate as well as the activities and binding of glycolytic and associated enzymes to subcellular structures was studied in brain, liver and white muscle of the teleost fish, Scorpaena porcus. Hypoxia exposure decreased glucose levels in liver from 2.53 to 1.70 µmol/g wet weight and in muscle led to its increase from 3.64 to 25.1 µmol/g wet weight. Maximal activities of several enzymes in brain were increased by hypoxia: hexokinase by 23%, phosphoglucoisomerase by 47% and phosphofructokinase (PFK) by 56%. However, activities of other enzymes in brain as well as enzymes in liver and white muscle were largely unchanged or decreased during experimental hypoxia. Glycolytic enzymes in all three tissues were partitioned between soluble and particulate-bound forms. In several cases, the percentage of bound enzymes was reduced during hypoxia; bound aldolase in brain was reduced from 36.4 to 30.3% whereas glucose-6-phosphate dehydrogenase fell from 55.7 to 28.7% bound. In muscle PFK was reduced from 57.4 to 41.7% bound. Oppositely, the proportion of bound aldolase and triosephosphate isomerase increased in hypoxic muscle. Phosphoglucomutase did not appear to occur in a bound form in liver and bound phosphoglucomutase disappeared in muscle during hypoxia exposure. Anoxia exposure also led to the disappearance of bound fructose-1,6-bisphosphatase in liver, whereas a bound fraction of this enzyme appeared in white muscle of anoxic animals. The possible function of reversible binding of glycolytic enzymes to subcellular structures as a regulatory mechanism of carbohydrate metabolism is discussed.

The association of genetic markers and malaria infection in the Brazilian Western Amazonian region

Almost all individuals (182) belonging to an Amazonian riverine population (Portuchuelo, RO, Brazil) were investigated for ascertaining data on epidemiological aspects of malaria. Thirteen genetic blood polymorphisms were investigated (ABO, MNSs, Rh, Kell, and Duffy systems, haptoglobins, hemoglobins, and the enzymes glucose-6-phosphate dehydrogenase, glyoxalase, phosphoglucomutase, carbonic anhydrase, red cell acid phosphatase, and esterase D). The results indicated that the Duffy system is associated with susceptibility to malaria, as observed in other endemic areas. Moreover, suggestions also arose indicating that the EsD and Rh loci may be significantly associated with resistance to malaria. If statistical type II errors and sample stratification could be ruled out, hypotheses on the existence of a causal mechanism or an unknown closely linked locus involved in susceptibility to malaria infection may explain the present findings.

Sativa seeds against Schistosoma mansoni different stages

The schistosomicidal properties of Nigella sativaseeds were tested in vitro against Schistosoma mansoni miracidia, cercariae, and adult worms. Results indicate its strong biocidal effects against all stages of the parasite and also showed an inhibitory effect on egg-laying of adult female worms. In the present work we also studied the effects of crushed seeds on some antioxidant enzymes; which have a role in protection of adult worms against host oxidant killing; as well as some enzymes of glucose metabolism; which have a crucial role in the survival of adult worms inside their hosts. The data revealed that the used drug induce an oxidative stress against adult worms which indicated by a decrease in the activities of both antioxidant enzymes, superoxide dismutase, glutathione peroxidase, and glutathione reductase and enzymes of glucose metabolism, hexokinase and glucose-6-phosphate dehydrogenase. Disturbing of such enzymes of adult worms using N. sativa seeds could in turn render the parasite vulnerable to damage by the host and may play a role in the antischistosomal potency of the used drug.

Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species

In this study, PCR assays targeting different Leishmania heat-shock protein 70 gene (hsp70) regions, producing fragments ranging in size from 230-390 bp were developed and evaluated to determine their potential as a tool for the specific molecular diagnosis of cutaneous leishmaniasis (CL). A total of 70 Leishmania strains were analysed, including seven reference strains (RS) and 63 previously typed strains. Analysis of the RS indicated a specific region of 234 bp in the hsp70 gene as a valid target that was highly sensitive for detection of Leishmania species DNA with capacity of distinguishing all analyzed species, after polymerase chain reaction-restriction fragment length polymorfism (PCR-RFLP). This PCR assay was compared with other PCR targets used for the molecular diagnosis of leishmaniasis: hsp70 (1400-bp region), internal transcribed spacer (ITS)1 and glucose-6-phosphate dehydrogenase (G6pd). A good agreement among the methods was observed concerning the Leishmania species identification. Moreover, to evaluate the potential for molecular diagnosis, we compared the PCR targets hsp70-234 bp, ITS1, G6pd and mkDNA using a panel of 99 DNA samples from tissue fragments collected from patients with confirmed CL. Both PCR-hsp70-234 bp and PCR-ITS1 detected Leishmania DNA in more than 70% of the samples. However, using hsp70-234 bp PCR-RFLP, identification of all of the Leishmania species associated with CL in Brazil can be achieved employing a simpler and cheaper electrophoresis protocol.

G6PD deficiency in Latin America: systematic review on prevalence and variants

Plasmodium vivax radical cure requires the use of primaquine (PQ), a drug that induces haemolysis in glucose-6-phosphate dehydrogenase deficient (G6PDd) individuals, which further hampers malaria control efforts. The aim of this work was to study the G6PDd prevalence and variants in Latin America (LA) and the Caribbean region. A systematic search of the published literature was undertaken in August 2013. Bibliographies of manuscripts were also searched and additional references were identified. Low prevalence rates of G6PDd were documented in Argentina, Bolivia, Mexico, Peru and Uruguay, but studies from Curaçao, Ecuador, Jamaica, Saint Lucia, Suriname and Trinidad, as well as some surveys carried out in areas of Brazil, Colombia and Cuba, have shown a high prevalence (> 10%) of G6PDd. The G6PD A-202A mutation was the variant most broadly distributed across LA and was identified in 81.1% of the deficient individuals surveyed. G6PDd is a frequent phenomenon in LA, although certain Amerindian populations may not be affected, suggesting that PQ could be safely used in these specific populations. Population-wide use of PQ as part of malaria elimination strategies in LA cannot be supported unless a rapid, accurate and field-deployable G6PDd diagnostic test is made available.

Bioassay for detection of transgenic soybean seeds tolerant to glyphosate

Glyphosate is a systemic, nonselective, postemergence herbicide that inhibits growth of both weeds and crop plants. Once inside the plant, glyphosate interferes with biosynthesis of aromatic amino acids phenylalanine, tyrosine, and tryptophan, by inhibiting the activity of 5enolpyruvylshikimate-3-phosphate synthase (EPSPS), a key enzyme of the shikimate pathway. The objective of this work was to develop a simple, effective and inexpensible method for identification of transgenic soybean tolerant to glyphosate. This technique consisted in germinating soybean seeds in filter paper moistened with 100 to 200 muM of glyphosate. Transgenic soybean seeds tolerant to glyphosate germinated normally in this solution and, between 7 and 10 days, started to develop a primary root system. However non-transgenic seeds stopped primary root growth and emission of secondary roots.

Effect of exposure to sublethal concentrations of sodium cyanide on the carbohydrate metabolism of the Indian Major Carp Labeo rohita (Hamilton, 1822)

Experiments were designed to study in-vivo effects of sodium cyanide on biochemical endpoints in the freshwater fish Labeo rohita. Fish were exposed to two sublethal concentrations (0.106 and 0.064mg/L) for a period of 15 days. Levels of glycogen, pyruvate, lactate and the enzymatic activities of lactate dehydrogenase (LDH), succinate dehydrogenase (SDH), glucose-6-phosphate dehydrogenase (G6PDH), phosphorylase, alkaline phosphatase (ALP), acid phosphatase (AcP) were assessed in different tissues (liver, muscle and gills). Result indicated a steady decrease in glycogen, pyruvate, SDH, ALP and AcP activity with a concomitant increase in the lactate, phosphorylase, LDH and G6PD activity in all selected tissues. The alterations in all the above biochemical parameters were significantly (p<0.05) time and dose dependent. In all the above parameters, liver pointing out the intensity of cyanide intoxication compare to muscle and gills. Study revealed change in the metabolic energy by means of altered metabolic profile of the fish. Further, these observations indicated that even sublethal concentrations of sodium cyanide might not be fully devoid of deleterious influence on metabolism in L. rohita.

Glyphosate as a tool to produce shikimic acid in plants

Oseltamivir phosphate is a potent viral inhibitor produced from shikimic acid extracted from seeds of Ilicium verum, the most important natural source. With the site of action 5-enolpyruvylshikimate-3-phosphate synthase (EPSP), glyphosate is the only compound capable of inhibiting its activity with the consequent accumulation of shikimic acid in plants. Corn and soybean plants were sprayed with reduced rates of glyphosate (0.0 to 230.4 g a.i. ha¹) and shikimic acid content in the dry mass was determined by HPLC 3, 7 and 10 days after application. Results showed shikimic acid accumulation in dry mass with increases of up to 969% in corn and 33,000% on soybeans, with peak concentrations 3 days after treatment (DAT). Industrial feasibility for shikimic acid production, combined with favorable climatic conditions for growing corn and soybean in virtually all over Brazil, favor the use of reduced rates of glyphosate in shikimic acid biosynthesis, with potential for use as an inducer in exploration of alternative sources for production of oseltamivir phosphate with low environmental impact.

Changes in the TBARs content and superoxide dismutase, catalase and glutathione peroxidase activities in the lymphoid organs and skeletal muscles of adrenodemedullated rats

Thiobarbituric acid reactant substances (TBARs) content, and the activities of glucose-6-phosphate dehydrogenase (G6PDh), citrate synthase (CS), Cu/Zn- and Mn-superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPX) were measured in the lymphoid organs (thymus, spleen, and mesenteric lymph nodes (MLN)) and skeletal muscles (gastrocnemius and soleus) of adrenodemedullated (ADM) rats. The results were compared with those obtained for sham-operated rats. TBARs content was reduced by adrenodemedullation in the lymphoid organs (MLN (28%), thymus (40%) and spleen (42%)) and gastrocnemius muscle (67%). G6PDh activity was enhanced in the MLN (69%) and reduced in the spleen (28%) and soleus muscle (75%). CS activity was reduced in all tissues (MLN (75%), spleen (71%), gastrocnemius (61%) and soleus (43%)), except in the thymus which displayed an increment of 56%. Cu/Zn-SOD activity was increased in the MLN (126%), thymus (223%), spleen (80%) and gastrocnemius muscle (360%) and was reduced in the soleus muscle (31%). Mn-SOD activity was decreased in the MLN (67%) and spleen (26%) and increased in the thymus (142%), whereas catalase activity was reduced in the MLN (76%), thymus (54%) and soleus muscle (47%). It is particularly noteworthy that in ADM rats the activity of glutathione peroxidase was not detectable by the method used. These data are consistent with the possibility that epinephrine might play a role in the oxidative stress of the lymphoid organs. Whether this fact represents an important mechanism for the establishment of impaired immune function during stress remains to be elucidated.

The carotid body of the spontaneous insulin-dependent diabetic rat

The carotid bodies from adult spontaneous insulin-dependent diabetic rats (strain BB/S) were perfusion-fixed at normal arterial blood pressure with 3% phosphate-buffered glutaraldehyde and compared with the organs from control rats (strain BB/Sc) prepared in the same way. Serial 5-µm sections were cut, stained, and using an interactive image analysis system, were analysed to determine the volumes of the carotid body and its vascular and extravascular compartments. There was no evidence of systemic arterial disease in the carotid stem arteries in either group of animals, and the microvasculature of the organs appeared normal by light microscopy. The volume of the carotid body was unchanged 3 months after the onset of diabetes but was increased at 6 months. The total vascular volume of the organ was unchanged, but the volume of the small vessels (5-12 µm) was increased. In the control group the small vessels comprised 5% of the total volume of the carotid body, or about 44% of the vascular compartment. The percentage of small vessels increased at 3 months in the diabetic group, but had returned to normal at 6 months. The extravascular volume followed the same pattern as the total carotid body volume and so did not change appreciably when expressed as a percentage of the total volume of the organ. The increase in size of the carotid body in diabetic rats is due, therefore, to an augmented extravascular volume. In one diabetic specimen the carotid sinus nerve showed signs of diabetic neuropathy, axonal swelling and intramyelinic oedema. The clinical implications of these results are discussed.

Application of protein-phenolic based coating on tomatoes (Lycopersicum esculentum)

The aim of this study was to investigate the use of protein-phenolic based coating made from fermented rice bran on cherry tomatoes (Lycopersicum esculentum). Tests were performed with glycerol 3% (v/v), glycerol with protein-phenolic rice bran extract (5%), glycerol with protein-phenolic extract after 96 hours of fermentation (5%), and a control (without coating). The coated cherry tomatoes were kept at room temperature for 28 days. Mass loss, pH and acidity, total soluble solids, and carotenoids were determined every 96 hours. The coating made from the biomass extract reduced the carotenoid and acidity levels in the fruits studied by 17 and 21.1%, respectively, compared to the control. The coating proved an efficient barrier to water vapor with mass loss of 57% less than the control suggesting that it can be used as an alternative for vegetable tissue conservation.

The use of triphenyltetrazolium chloride in the study of dehydrogenase activity of Brucellae

Experiments for the investigation of dehydrogenase activity of washed cells of a strains of Br. abortus and another of Br. suis in presence of different single added substrates are reported. The activity was measured as the amount of formazan produced by the reduction of 2, 3, 5-triphenyltetrazolum chloride acting as a hydrogen ions acceptor, at pH 7.0. In a general manner the dehydrogenase activity of Br. suis was much more intense than that of Br. abortus (fig. 5). In the conditions of the experiments Br. abortus oxidized L-arabinose, D-galactose, D-glucose, glycerol, D-xylose, DL-alanine, D-fructose, and D-sorbitol. Brucella suis oxidized D-xylose, L-arabinose, D-glucose, D-galactose, DL-alanine, sodium acetate, maltose, glycine, D-fructose, and D-sorbitol. Glycerol was oxidized by Br. abortus but its oxidation by Br. suir was very slight. Sodium acetate and maltose were intensely oxidized by Br. suir but not by Br. abortus. The sites of more intense enzymatic acitivity were seen as small red colored round granules located in one pole of the cells.

Initial performance of pineapple and utilization of rock phosphate applied in combination with organic compounds to leaf axils

Rock phosphates have low solubility in water, but good solubility in acid. The use of organic compounds together with these phosphorus sources applied to the basal leaf axils of pineapple can increase the solubility of this phosfate source and increase the P availability to the crop. A greenhouse experiment was conducted using Araxá rock phosphate (10 g) in combination or not with solutions containing increasing concentrations of humic acids (0 to 40 mmol L-1 of carbon), with or without citric acid (0.005 mmol L-1), applied to basal leaf axils of pineapple cv. Pérola. Growth and nutritional characteristics of aerial plant parts were assessed. Growth rates of aerial parts and N, P, K, Ca and Mg contents increased curvilinearly with increasing concentration of carbon in the form of humic acids. Maximum values were found for the concentration of 9.3 mmol L-1 of carbon combined with 0.005 mmol L-1 of citric acid and natural phosphate.

Performance of pineapple slips inoculated with diazotrophic phosphate-solubilizing bacteria and rock phosphate

Besides fixing N2, some diazotrophic bacteria or diazotrophs, also synthesize organic acids and are able to solubilize rock phosphates, increasing the availability of P for plants. The application of these bacteria to pineapple leaf axils in combination with rock phosphate could increase N and P availability for the crop, due to the bacterial activity of biological nitrogen fixation and phosphate solubilization. The objectives of this study were: (i) to select and characterize diazotrophs able to solubilize phosphates in vitro and (ii) evaluate the initial performance of the pineapple cultivars Imperial and Pérola in response to inoculation with selected bacteria in combination with rock phosphate. The experiments were conducted at Universidade Estadual do Norte Fluminense Darcy Ribeiro, in 2009. In the treatments with bacteria the leaf contents of N, P and K were higher than those of the controls, followed by an increase in plant growth. These results indicate that the combined application of diazotrophic phosphate-solubilizing bacteria Burkholderia together with Araxá rock phosphate can be used to improve the initial performance of pineapple slips.

Factors affecting phosphate fixation in soils of the state of São Paulo

A laboratory experiment was carried out to study the effects of chemical and physical characteristics of the soil on the phosphate fixing capacity. One hundred samples collected from various localities were at first characterized chemically and their particlesize distribution determined. They were then tested as to their phosphate fixing capacities. The results obtained were statistically analysed by means of both simple linear and multiple correlation. The following conclusions could be drawn: 1. simple linear regression analysis indicated that % C, exchangeable Al+3, CEC, % clay, pH and % sand were the soil characteristics which significantly affected phosphate fixing capacity of São Paulo State soils; 2. multiple linear regression analysis indicated that % C, exchangeable Mg(+2)9 exchangeable- Al+3 and % clay were the soil characteristics which significantly affected the phosphate fixing capacity of São Paulo State soils; 3. the phosphate phonomena fixing as they occur in the soils of the São Paulo State can be best described by the following equation: Y = -2,266 - 3,484 + 3,514 + 5,559 + 1,005 %C Mg+2 Al+3 % clay exchangeable exchangeable 4. phosphate fixation in the soil is affected by the combined effects of both soil chemical and physical characteristics.