Chromosomal polymorphism,syntenic relationships,and ploidy in the pathogenic fungus Paracoccidioides brasiliensis

Pulsed field gel electrophoresis (PFGE) and DNA hybridization were used to establish and compare the electrophoretic karyotypes of 12 clinical and environmental Paracoccidioides brasiliensis isolates from different geographic areas. Gene mapping allowed the identification of synteny groups and the use of isolated whole chromosomal bands to probe chromoblots indicated the existence of repetitive sequences, contributing to a better understanding of the structure and organization of the fungus genome. This represents the first comparative mapping study among different isolates. The results are indicative of the existence of genetic differences among natural isolates. DNA content of DAPI-stained nuclei of each isolate was estimated by confocal microscopy. Comparison of the genome sizes estimated by PFGE with those calculated by microfluorometry indicated the possible existence of haploid and diploid (or aneuploid) isolates of the fungus.     

Immunolocalization of proacrosin/acrosin in bovine sperm and sperm penetration through the zona pellucida

The aim of the present work was to immunolocalize acrosin in bull spermatozoa incubated for up to 6 h in capacitating culture medium (TALP-heparin), in order to study the kinetics of its release during the acrosome reaction and in vitro sperm penetration. Six replicates from semen of one bull were used. Acrosin was localized by the silver-enhanced immunogold technique using anti-bovine acrosin monoclonal antibody ACRO-C2E5. Spermatozoa thus showed the presence of acrosin only at the acrosomal region. Four different patterns were seen: (1) no labeling: (2) intense labeling on the rim of the portion of the acrosome; (3) diffuse label over the entire acrosomal region; and (4) intense label over the entire acrosomal region. Spermatozoa incubated in capacitating medium for 4 h showed that unlabeled (pattern 1) spermatozoa decreased from 72% to 28% difference that was found to be significant (p<0.05). Patterns 3 and 4 increased from about 10% to 20-29%, (p<0.05). With further incubation (4-6 h), pattern 1 increased while patterns 3 and 4 decreased differences were not significant (p0.05). The incidence of pattern 2 did not change through the whole incubation period. Sperm penetration through the zona pellucida of in vitro matured bovine oocytes (57%) or empty zonae pellucida (70.5%) increased (p<0.05) as a function of sperm incubation time in capacitating medium. The presence of acrosin, as determined by the silver-enhanced immunogold technique, was highly correlated with sperm penetration of in vitro mature bovine oocyte (r=0.98) and cryopreserved zonae pellucidae (r=0.93) (p<0.01).     

Characterization of constitutive heterochromatin of Callithrix geoffroyi (Callitrichidae, Primates) by restriction enzymes and fluorochrome bands

The neotropical primate genus Callithrix comprises two groups of species, jacchus and argentata, which inhabit distinct geographical regions and manifest different fur coloration and constitutive heterochromatin (CH) markers in their karyotypes. In this investigation the CH of a representative of the jacchus group, Callithrix geoffroyi, was analysed using fluorochromes and restriction enzymes in situ. To clarify the source of the constitutive heterochromatin of both groups, the data obtained in the jacchus group were compared with those published in the argentata group obtained by the same techniques. The C-bands of C. geoffroyi (four specimens, 2n = 46) were centromeric in all chromosomes, and distally located in pairs 6 and 22. The Alu I, Hae III, Hin fI, Rsa I, Dde I, Mbo I, and Msp I restriction endonucleases and CMA3 and DAPI fluorochromes produced different bands, which allowed the characterization of four distinct types of constitutive heterochromatin in the C. geoffroyi genome. Several of these types of heterochromatin were present in the ancestor of the two groups of species, jacchus and argentata, while others originated after their cladogenesis.     

Preparation of sucrose heptaesters unsubstituted at the C-1 hydroxy group of the fructose moiety via selective O-desilylation

Selective O-desilylation of 6,1',6'-tri-O-tert-butyldiphenylsilyl-2,3,4,3',4'-penta-O-benzo ylsucrose with hydrofluoric acid in acetonitrile led to the 1'-O-tert-butyldiphenylsilyl derivative (96% yield), which was further perbenzoylated and deprotected at OH-1' with tetrabutylammonium fluoride (86%). An analogous sequence with the corresponding O-acetylated sucrose derivative and tetrabutylammonium fluoride as desilylating agent resulted in a lower yield of the C-1' hydroxy derivative.     

Effect of nitric oxide synthase inhibition on hypercapnia-induced hypothermia and hyperventilation

Hypercapnia elicits hypothermia in a numberof vertebrates, but the mechanisms involved are not well understood. Inthe present study, we assessed the participation of the nitric oxide(NO) pathway in hypercapnia-induced hypothermia and hyperventilation bymeans of NO synthase inhibition by usingN-nitro-L-arginine(L-NNA). Measurements ofventilation, body temperature, and oxygen consumption were performed inawake unrestrained rats before and afterL-NNA injection(intraperitoneally) and L-NNA injection followed by hypercapnia (5%CO₂). Control animals received saline injections. L-NNA alteredthe breathing pattern during the control situation but not duringhypercapnia. A significant (P < 0.05) drop in body temperature was measured after bothL-NNA (40 mg/kg) and 5%inspired CO₂, with a drop inoxygen consumption in the first situation but not in the second.Hypercapnia had no effect onL-NNA-induced hypothermia. Theventilatory response to hypercapnia was not changed byL-NNA, even thoughL-NNA caused a drop in bodytemperature. The present data indicate that the two responses elicitedby hypercapnia, i.e., hyperventilation and hypothermia, do not share NOas a common mediator. However, theL-arginine-NO pathwayparticipates, although in an unrelated way, in respiratory function andthermoregulation.

     

Synthesis and biodistribution studies of carbohydrate derivatives radiolabeled with technetium-99m

Three carbohydrate derivatives, MAG₃-Gl, MAG₃-Ga, MAG₃-NG, were synthesized and radiolabeled in high yields. These substances were injected in health Swiss mice and their biodistribution were evaluated. Among them, ^99mTc-MAG₃-Ga displayed higher accumulation in hepatic tissue, due to the presence of specific receptors in the liver for this carbohydrate. Thus, the use of ^99mTc-MAG₃-Ga to assess hepatic function can be considered.     

Conventional sampling plan for the green leafhopper Empoasca kraemeri in common beans

Abstract:  A conventional sampling plan for the leafhopper Empoasca kraemeri Ross & Moore (Auchenorrhyncha: Cicadellidae) in common beans (Phaseolus vulgaris L.) is presented based on samples from 51 commercial fields. The research encompassed three phases. The first phase had the objective of determining the most suitable leaf to be used as a precise and representative sampling unit. In the second phase of the project, the available sampling methods for E. kraemeri were compared, including the approach identified in the first phase. In the third phase, the theoretical frequency distribution of the sampling data was assessed and the number of samples necessary for the sampling plan was established. The best leaves to sample adult leafhoppers were the fourth and the fifth from the apex, while the best leaf for sampling nymphs was the fifth from the apex. The best sampling technique for nymphs and adults was the beating of the apical leaves against a plastic tray. The sampling data obtained with this technique were fit to a negative binomial distribution with common aggregation parameter for adults ( K _common = 1.04) and nymphs ( K _common = 0.47). This sampling plan required 16.8 and 31.2 min to be carried out at an expense of US$ 0.31 and US$ 0.54 for sampling adults and nymphs in a field, respectively.     

Bovicin HC5 inhibits wasteful amino acid degradation by mixed ruminal bacteria in vitro

Streptococcus bovis HC5 produces a broad spectrum lantibiotic (bovicin HC5) that inhibits pure cultures of hyper ammonia-producing bacteria (HAB). Experiments were preformed to see if: (1) S. bovis HC5 cells could inhibit the deamination of amino acids by mixed ruminal bacteria taken directly from a cow, (2) semi-purified bovicin was as effective as S. bovis HC5 cells, and 3) semi-purified and the feed additive monensin were affecting the same types of ammonia-producing ruminal bacteria. Because purified and semi-purified bovicin HC5 was as effective as S. bovis HC5 cells, it appeared that bovicin HC5 was penetrating the cell membranes of HAB before it could be degraded by peptidases and proteinases. Mixed ruminal bacteria that were successively transferred and enriched nine times with trypticase did not become significantly more resistant to either bovicin HC5 (50 AU mL⁻¹) or monensin (5 μM), and amplified rDNA restriction analysis indicated that bovicin HC5 and monensin appeared to be selecting against the same types of bacteria.