Inhibition of the Ecto-ATPdiphosphohydrolase of Schistosoma mansoni by Thapsigargin

ATPdiphosphohydrolases (ATPDases) are ubiquitous enzymes capable ofhydrolyzing nucleoside di- and triphosphates. Although a number ofpossible physiological roles have been proposed for ATPDases, detailedstudies on structure-function relationships have generally been hamperedby the lack of specific inhibitors of these enzymes. We have previouslycharacterized a Ca²⁺-activated ATPDase on the external surface ofthe tegument of Schistosoma mansoni, the etiologic agent of humanschistosomiasis. In the present work, we have examined the effectsof thapsigargin, a sesquiterpene lactone known as a high affinityinhibitor of sarco-endoplasmic reticulum calcium transport (SERCA)ATPase, on ATPDase activity. Whereas other lactones tested had littleor no inhibitory action, thapsigargin inhibited ATP hydrolysis by theATPDase (K _i20 M). Interestingly, hydrolysis of ADP was notinhibited by thapsigargin. The lack of inhibition of ATPase activityby orthovanadate, a specific inhibitor of P-type ATPases, and theinhibition of the Mg²⁺-stimulated ATP hydrolysis by thapsigarginruled out the possibility that the observed inhibition of the ATPDaseby thapsigargin could be due to the presence of contaminating SERCAATPases in our preparation. Kinetic analysis indicated that a singleactive site in the ATPDase is responsible for hydrolysis of both ATPand ADP. Thapsigargin caused changes in both V _max and K _m for ATP,indicating a mixed type of inhibition. Inhibition by thapsigarginwas little or not affected by changes in free Ca²⁺ or Mg²⁺concentrations. These results suggest that interaction of thapsigarginwith the S. mansoni ATPDase prevents binding of ATP or its hydrolysisat the active site, while ADP can still undergo catalysis.     

Isospora lutrae n. sp. (Apicomplexa: Eimeriidae), a new coccidium from the European otter Lutra lutra (L.) (Carnivora: Mustelidae) from Spain

Parasitological examination of European otter originating from Extremadura, Spain revealed the presence of a new isosporan species. Oöcysts of Isospora lutrae n. sp. are spherical to subspherical, 31.2 (27.5–32) × 29.6 (28–31) m and have a smooth wall c. 1 m thick. Sporocysts are ellipsoidal, 18.2 (17–19) × 14.4 (14–16) m and lack Stieda and substieda bodies. A spherical sporocyst residuum is present, consisting of granules scattered among the sporozoites. Sporozoites are spindle-shaped, 12.4 × 2.5 m and have anterior and posterior refractile bodies. Based on its unique morphologic structure and host, I. lutrae is considered to be new.     

Propionic acid metabolism and poly-3-hydroxybutyrate-co-3-hydroxyvalerate (P3HB-co-3HV) production by Burkholderia sp

Mutants of Burkholderia sp. that are unable to grow on propionic acid (prp) but still accumulate P3HB-co-3HV from carbohydrate and propionic acid were studied. In shaken flask tests, yields of 3HV from propionic acid (Y(3HV/Prop)) increased from 0.10 g g(-1) in the wild type to c.a. 0.35 g g(-1) in mutants affected in alpha-oxidation pathway or to 0.80 g g(-1) in mutants not affected in that pathway. In bioreactor tests, mutant IPT 189 showed Y(3HV/Prop) = 1.20 g g(-1), a yield very close to the theoretical maximum of 1.35 g g(-1). Accumulation of 3HV units from unrelated carbon sources was undetectable in these mutants indicating that 3HV units are produced directly from propionic acid. Thus, the industrial use of those mutants to produce the copolymer from sucrose and propionic acid could significantly reduce the production costs. The results strongly suggest the existence of at least two pathways that are involved in the oxidation of propionic acid in Burkholderia sp. Their rates would be modulated by the availability of propionic acid.     

Skeletal muscle abnormalities associated with occupational exposure to mercury vapours

There is scarce information on the possible effects of chronic exposure to mercury on skeletal muscle. Dental personnel are frequently exposed to inhalation of metallic mercury vapours. The skeletal muscle of five technicians and one dentist (females, age 36-55) was studied. All of them presented symptoms of chronic mercury poisoning. Needle biopsy was taken from the quadriceps femoris muscle and samples were prepared for light microscope histochemistry and for transmission electron microscopy. Selective atrophy of type IIB muscle fibres was found in patients, and in one of them there was fibre grouping. Most of the muscles showed increased fibre area per capillary. Atrophy was confirmed by the ultrastructural study, demonstrating increase of intermyofibrillar spaces, loss of myofibrils or complete disappearance in some fibres, and sarcolemmal foldings. Splitting of the fibres was also found. Some capillaries were altered, showing endothelial infoldings into the lumen, thickened basement membrane and partial or total occlusion. The alterations found in muscle may be secondary to nerve damage, to ischemia caused by capillary lesion and/or to a direct effect of mercury on muscle fibre proteins.     

Elastin variations implicating in vascular smooth muscle cells phenotype in human tortuous arteries

The aim of the present work was to study the morphological implications between the elastin and the phenotypic expression of the vascular smooth muscle cells. For this purpose, sixty human tortuous arteries from different territories have been studied. We have measured the morphometric indexes Intimal Thickening Index and Elastolyse Index and they have been quantified with computer system analysis, image-colour corresponding to the orcein and Verho?ff reactions for detecting elastin and the alpha-actin in the smooth muscle cells. We compared both territorial arteries from the cranial and from abdominal origin. The elastin concentration was similar in both territories, but not its morphology according to its spatial distribution. We have observed a relationship between the elastin structural organisation from the media of arteries and of the internal elastic lamina in these territories and the variation of reactivity to the smooth muscle alpha-actin as a marker of the phenotypic state. Our results confirm the hypothesis that elastin, besides intervening in the architecture of the arterial wall, is a factor implicated in the phenotypic variability of the smooth muscle cells and in the development and evolution of the intimal thickenings in human atherosclerosis.     

MRNA encoding a putative RNA helicase of the DEAD-box gene family is up-regulated in trypomastigotes of Trypanosoma cruzi

Differential display of mRNAs from Trypanosoma cruzi epimastigote and metacyclic trypomastigote stages showed several mRNA species differing in their expression level. The cDNA corresponding to one of these mRNAs was used as a probe in Northern blots and identified a RNA product of 2.6 kb with an expression level eight or more times higher in trypomastigotes than in epimastigotes. This probe was also used to screen a genomic library of T. cruzi CL Brener clone prepared in lambda FIX. A clone of about 15 kb was selected that, after partial sequencing, revealed an open reading frame of 688 amino acids encoding a deduced protein with similarity to RNA helicases of the DEAD-box gene family. The presence of the eight conserved motifs characteristic of the DEAD protein family was observed in the T. cruzi sequence, indicating that it corresponds to a putative RNA helicase gene, which we named HelTc. Southern blot analysis indicated that HelTc is a single-copy gene. Pulsed-field gel electrophoresis separation of chromosomes of several isolates of T. cruzi showed that this gene was localized in one or two chromosomal bands.     

Cultural characteristics of some ectomycorrhizal fungi in pure culture

Macro and microscopic features for 21 species of ectomycorrhizal fungi are presented, with regard to their particular importance as possible taxonomic criteria. The culture media used are MMN and BAF. Cultural characteristics of some of our isolates, like color or morphology of the mycelia, were comparable to those observed by different authors for the same species, or species which are closely related taxonomically, as for example Boletus satanas and Lactarius deliciosus in BAF media, or Hebeloma edurum, Lycoperdon perlatum, Paxillus atrotomentosus, Rhizopogon roseolus and Suillus collinitus in MMN media. Macroscopically the color of mycelia, the amount of aerial mycelia and microscopically the exudates and the presence of inflated and flexuous cells were the most variable characteristics within species.     

Effects of Dihydrotestosterone on Brain mRNA Levels of Steroid 5α-Reductase Isozymes in Early Postnatal Life of Rat

We studied the expression of type 1 (5α-R1) and type 2 (5α-R2) 5α-reductase isozymes (5α-R) and their regulation by dihydrotestosterone (DHT) in the prefrontal cortex of male and female rats during postnatal sexual differentiation of the central nervous system (CNS), using one-step quantitative RT-PCR coupled with laser-induced fluorescence capillary electrophoresis. We found a higher expression of 5α-R2, which is considered a masculinizing enzyme, in the female versus male CNS, and observed sexual dimorphism in the regulation of both 5α-R isozymes by DHT. These results open up a new research line that could improve understanding of the role of 5α-R isozymes in the physiology of the CNS.     

Three new species of Isospora schneider, 1881 (Apicomplexa: Eimeriidae) from the double-collared seed eater, Sporophila caerulescens (Passeriformes: Emberizidae), from Eastern Brazil

Three isosporan species are described from the double-collared seedeater, Sporophila caerulescens from Eastern Brazil. Isospora sporophilae n. sp. oocysts spherical to subspherical; oocyst wall bi-layered, smooth, inner layer colorless to pale yellowish, 21.6 x 20.9 (19.20-23.20 x 18.40-22.60) microm, shape-index 1.03 +/- 0.02 (1-1.10), with no micropyle or oocyst residuum. Polar bodies splinter-like or comma-like. Sporocysts ovoidal, 15.2 x 10.6 (17.40-12.80 x 12.60-8.40) microm, shape-index 1.43 +/- 0.14 (1.17-1.81), with knob-like Stieda body and residuum. Large crystalloid body in the center of the sporocyst. Isospora flausinoi n. sp. oocysts spherical to subspherical, oocyst wall bi-layered, smooth, colorless, 17.30 x 16.53 (14-20 x 13.60-20) microm, shape-index 1.05 +/- 0.04 (1-1.21). Micropyle and oocyst residuum absent; presence of a large polar body. Sporocystpiriform, 14.88 x 10.70 (11.80-18 x 8-12.40) microm, shape-index 1.40 +/- 0.18 (1.07-1.77), with smooth, thin, single-layered wall. Sporocyst with rounded Stieda body with no substieda body, and residuum composed of granular material. Isospora teixeirafilhoi n. sp. oocysts spherical to subspherical, oocyst wall bi-layered, smooth, colorless, 17.41 x 16.81 (15.60 - 19.40 x 14.20-18.80) microm. Shape-index 1.04 +/- 0.08 (1-1.12). Micropyle and oocyst residuum absent; presence of a small double-lobuled polar body. Sporocyst ovoid, 11.74 x 8.12 (9-14.20 x 6.20-9.40) microm. Shape-index 1.46 +/- 0.23 (1.06-1.88). Sporocyst with knob-like Stieda body, no sub-Stieda body and residuum composed of granular material.