The Tropical Biominer Project: mining old sources for new drugs

The Tropical Biominer Project is a recent initiative from the Federal University of Minas Gerais (UFMG) and the Oswaldo Cruz foundation, with the participation of the Biominas Foundation (Belo Horizonte, Minas Gerais, Brazil) and the start-up Homologix. The main objective of the project is to build a new resource for the chemogenomics research, on chemical compounds, with a strong emphasis on natural molecules. Adopted technologies include the search of information from structured, semi-structured, and non-structured documents (the last two from the web) and datamining tools in order to gather information from different sources. The database is the support for developing applications to find new potential treatments for parasitic infections by using virtual screening tools. We present here the midpoint of the project: the conception and implementation of the Tropical Biominer Database. This is a Federated Database designed to store data from different resources. Connected to the database, a web crawler is able to gather information from distinct, patented web sites and store them after automatic classification using datamining tools. Finally, we demonstrate the interest of the approach, by formulating new hypotheses on specific targets of a natural compound, violacein, using inferences from a Virtual Screening procedure.     

3-hydroxyglutaric acid enhances glutamate uptake into astrocytes from cerebral cortex of young rats

A predominantly neurological presentation is common in patients with glutaric acidemia type I (GA-I). 3-hydroxyglutaric acid (3-OHGA), which accumulates in affected patients, has recently been demonstrated to play a central role in the neuropathogenesis of this disease. In the present study, we investigated the in vitro effects of 3-OHGA at concentrations ranging from 10 to 1000 microM on various parameters of the glutamatergic system, such as the basal and potassium-induced release of [3H]glutamate by synaptosomes, as well as on Na+-dependent [3H]glutamate uptake by synaptosomes and astrocytes and Na+-independent [3H]glutamate uptake by synaptic vesicles from cerebral cortex of 30-day-old Wistar rats. First, we observed that exposure of cultured astrocytes to 3-OHGA for 20 h did not reduce their viability. Furthermore, 3-OHGA significantly increased Na+-dependent [3H]glutamate uptake by astrocytes by up to 80% in a dose-dependent manner at doses as low as 30 microM. This effect was not dependent on the presence of the metabolite during the uptake assay, since it occurred even when 3-OHGA was withdrawn from the medium after cultured cells had been exposed to the acid for approximately 1 h. All other parameters investigated were not influenced by this organic acid, indicating a selective action of 3-OHGA on astrocyte transporters. Although the exact mechanisms involved in 3-OHGA-stimulatory effect on astrocyte glutamate uptake are unknown, the present findings contribute to the understanding of the pathophysiology of GA-I, suggesting that astrocytes may protect neurons against excitotoxic damage caused by 3-OHGA by increasing glutamate uptake and therefore reducing the concentration of this excitatory neurotransmitter in the synaptic cleft.     

Inhibition of creatine kinase activity from rat cerebral cortex by D-2-hydroxyglutaric acid in vitro

D-2-Hydroxyglutaric acid (DGA) is the biochemical hallmark of patients affected by the neurometabolic disorder known as D-2-hydroxyglutaric aciduria (DHGA). Although this disease is predominantly characterized by severe neurological findings, the underlying mechanisms of brain injury are virtually unknown. In the present study, we investigated the effect of DGA on total, cytosolic, and mitochondrial creatine kinase (CK) activities from cerebral cortex of 30-day-old Wistar rats. Total CK activity (tCK) was measured in whole cell homogenates, whereas cytosolic and mitochondrial activities were measured in the cytosolic and mitochondrial preparations from cerebral cortex. We verified that CK activities were significantly inhibited by DGA (11-34% inhibition) at concentrations as low as 0.25 mM, being the mitochondrial fraction the most affected activity. Kinetic studies revealed that the inhibitory effect of DGA was non-competitive in relation to phosphocreatine. We also observed that this inhibition was fully prevented by pre-incubation of the homogenates with reduced glutathione, suggesting that the inhibitory effect of DGA on tCK activity is possibly mediated by oxidation of essential thiol groups of the enzyme. Considering the importance of CK activity for brain metabolism homeostasis, our results suggest that inhibition of this enzyme by increased levels of DGA may be related to the neurodegeneration of patients affected by DHGA.     

Detection of high-level tetracycline resistance in clinical isolates of Helicobacter pylori using PCR-RFLP

Tetracycline is one of four antibiotics commonly used for the treatment of Helicobacter pylori infection, but its effectiveness is decreasing as the incidence of tetracycline resistance is increasing. In five Brazilian tetracycline-resistant (Tet(R)) H. pylori isolates, high-level tetracycline resistance is mediated by the triple-base-pair substitution AGA(926-928)-->TTC in both 16S rRNA genes, as was previously observed in two independent high-level Tet(R) H. pylori strains. A polymerase chain reaction-based restriction fragment length polymorphism (PCR-RFLP) assay was developed for the detection of the AGA(926-928)-->TTC substitution, and confirmed the presence of the aforementioned triple-base-pair substitution in all five Brazilian Tet(R) isolates. This PCR-RFLP-based approach distinguishes the high-level Tet(R) isolates from low-level Tet(R) and Tet(S) H. pylori strains and thus allows the direct detection of Tet(R) H. pylori isolates.     

Alterations in atrial natriuretic peptide and its receptors in streptozotocin-induced diabetic rat kidneys

In this study the effect of diabetes mellitus on atrial natriuretic peptide (ANP) receptors in streptozotocin- (STZ-) induced diabetic rat kidneys was studied. Moreover, plasma ANP concentration was evaluated in diabetic and control rats by using radioimmunoassay. In addition, the expression of ANP in the kidneys of control and diabetic rats was evaluated by immunohistochemistry. Body-weight loss and increased glucose levels were used as indices of diabetes mellitus in the STZ-induced rats. There was a significant loss in the body weight of the diabetic rats compared to controls. The efficacy of STZ administration was confirmed by rising blood glucose levels, which were significantly higher in diabetic rats compared to controls. Plasma ANP concentration was significantly greater in the diabetic rats in comparison with controls. Moreover, our immunohistochemical results show that the expression of ANP in diabetic rats was higher than that in age-matched controls. ANP was observed in the cells lining the proximal convoluted tubules in the cortex. The distribution and levels of ANP receptors in the kidneys of diabetic rats and age-matched controls were investigated using quantitative receptor autoradiography. Our results demonstrate significant decrease in ANP receptors in the kidneys of the diabetic rats compared to controls. The significant decrease was found in the juxtaglomerular medulla, inner medulla, and the papillae. The decrease in ANP receptors observed in the diabetic kidneys could have pathological consequences resulting in renal resistance to ANP in diabetes.     

Prevalence and Exoenzyme Secretion by Candida Albicans Isolates from Oral and Vaginal Mucosas of HIV-Infected Women

A cross-sectional study was performed to evaluate the prevalence and the aetiology of forms of mucosal fungal infections of HIV-negative and HIV-positive women. Candida albicans was the predominate specie isolated from both groups of patients, with remarkable proportion of isolation from symptomatic women. All 239 C. albicans isolates, regardless of their source, showed activity of proteinase and phospholipase. It was verified that isolates with particularly higher levels of exoenzymes production were significantly more common in HIV-positive patients. However, isolates obtained from the HIV-positive patients in use of HAART, with protease inhibitor, presented lower levels of these exoenzymes, similar to the levels observed in the isolates from HIV-negative patients.     

Diagnosis of Clinical Bovine Mastitis by Fine Needle Aspiration Followed by Staining and Scanning Electron Microscopy in a Prototheca zopfii Outbreak

Fruit anthracnose of ugurassa caused by Colletotrichum acutatum is hereby reported for the first time in Sri Lanka and it is proposed that C. acutatum is considered together with C. gloeosporioides, as a causal agent of this disease. C. acutatum was characterised by fusiform conidia and white to orange colonies with slight shades of light mouse grey aerial mycelia. C. gloeosporioides produced grey colonies with a dark mouse grey centre and conidia were cylindrical. The other differences between the ugurassa isolate of C. acutatum and C. gloeosporioides were slower growth at temperatures ranging from 15-30 degrees C and extremely high tolerance of two fungicides, carbendazim and thiophanate methyl.