Dichlorobenzidine-DNA binding catalyzed by peroxidative activation in Salmonella typhimurium

Peroxidative oxidation of dichlorobenzidine in vitro results in covalent binding to exogenous DNA. In a modified Ames assay, mutagenicity is observed in S. typhimurium strain TA98 following the incubation of dichlorobenzidine, bacteria, and hydrogen peroxide. In this paper, we demonstrate that [14C]dichlorobenzidine becomes covalently bound to S. typhimurium macromolecules, including DNA, when exogenous hydrogen peroxide is supplied. We compared the levels of binding in a pair of otherwise isogenic strains with wild-type (oxyR+) versus constitutive (oxyR1) expression of the hydrogen peroxide stress-induced regulon. Binding was approximately twofold higher in TA4124 (oxyR1) than in TA4123 (oxyR+). Bacterial hydroperoxidases may catalyze the activation of dichlorobenzidine to mutagenic and DNA binding species in this system.     

Courtship Behavior of Zaprionus indianus (Gupta) (Diptera: Drosophilidae) from Populations Colonizing South America

We describe for the first time the sexual behavior and the courtship song of males of the African fly Zaprionus indianus (Gupta), a recent invader of South America. The male courtship song is formed by monocyclic pulses and the courtship behavior is simple when compared to that of species of Drosophila. Two interpulse interval (IPI) distributions were observed: pre-mounting and mounting. No significant difference was observed between the pre-mounting IPIs of males that descended from three geographical populations from South America. We also observed the songs produced by females and the homosexual behavior exhibited by males. A sequence of bursts is produced by females as a refusal signal against males, while males emit a characteristic song that identifies sex genus, which differs from the courtship song. The short courtship and mating latencies recorded reveal vigorous males and receptive females, respectively.     

Modeling electroporation in a single cell. I. Effects Of field strength and rest potential.

This study develops a model for a single cell electroporated by an external electric field and uses it to investigate the effects of shock strength and rest potential on the transmembrane potential V(m) and pore density N around the cell. As compared to the induced potential predicted by resistive-capacitive theory, the model of electroporation predicts a smaller magnitude of V(m) throughout the cell. Both V(m) and N are symmetric about the equator with the same value at both poles of the cell. Larger shocks do not increase the maximum magnitude of V(m) because more pores form to shunt the excess stimulus current across the membrane. In addition, the value of the rest potential does not affect V(m) around the cell because the electroporation current is several orders of magnitude larger than the ionic current that supports the rest potential. Once the field is removed, the shock-induced V(m) discharges within 2 micros, but the pores persist in the membrane for several seconds. Complete resealing to preshock conditions requires approximately 20 s. These results agree qualitatively and quantitatively with the experimental data reported by Kinosita and coworkers for unfertilized sea urchin eggs exposed to large electric fields.     

Transgenic alteration of ethylene biosynthesis increases grain yield in maize under field drought‐stress conditions

A transgenic gene‐silencing approach was used to modulate the levels of ethylene biosynthesis in maize (Zea mays L.) and determine its effect on grain yield under drought stress in a comprehensive set of field trials. Commercially relevant transgenic events were created with down‐regulated ACC synthases (ACSs), enzymes that catalyse the rate‐limiting step in ethylene biosynthesis. These events had ethylene emission levels reduced approximately 50% compared with nontransgenic nulls. Multiple, independent transgenic hybrids and controls were tested in field trials at managed drought‐stress and rain‐fed locations throughout the US. Analysis of yield data indicated that transgenic events had significantly increased grain yield over the null comparators, with the best event having a 0.58 Mg/ha (9.3 bushel/acre) increase after a flowering period drought stress. A (genotype × transgene) × environment interaction existed among the events, highlighting the need to better understand the context in which the down‐regulation of ACSs functions in maize. Analysis of secondary traits showed that there was a consistent decrease in the anthesis‐silking interval and a concomitant increase in kernel number/ear in transgene‐positive events versus nulls. Selected events were also field tested under a low‐nitrogen treatment, and the best event was found to have a significant 0.44 Mg/ha (7.1 bushel/acre) yield increase. This set of extensive field evaluations demonstrated that down‐regulating the ethylene biosynthetic pathway can improve the grain yield of maize under abiotic stress conditions.     

A new technique for staining mast cells using ferroin

We describe here a new method for specific staining of mast cells using ferroin. Different hamster tissues were fixed in 4% formalin and processed for paraffin embedding. Sections were stained with hematoxylin followed by ferroin acidified with 2.5 N sulfuric acid to pH 4.0. Mast cells stained an intense orange color that contrasted markedly with bluish violet nuclei. High contrast was also observed when ferroin colored sections were counterstained with light green instead of hematoxylin. To evaluate the specificity of the stain, hamster cheek pouch sections were stained with toluidine blue, alcian blue-safranin O, and ferroin. Quantitative evaluation of mast cells stained with the three techniques showed no statistical difference. The simplicity and selectivity of this method is sufficient for image analysis of mast cells.     

Organization and evolution of the alcohol dehydrogenase gene in Drosophila

The alcohol dehydrogenase (Adh) gene was isolated from Drosophila simulans and D. mauritiana, and the DNA sequence of a 4.6-kb region, containing the structural gene and flanking sequence, was determined for each. These sequences were compared with the Adh region of D. melanogaster to characterize changes that occur in the Drosophila genome during evolution and to identify conserved sequences of functional importance. Drosophila simulans and D. mauritiana Adh are organized in a manner similar to that of D. melanogaster Adh, including the presence of two promoters for the single Adh gene. This study identified conserved flanking elements that, in conjunction with other studies, suggest regions that may be involved in the control of Adh expression. Inter- and intraspecies comparisons revealed differences in the kinds of sequence changes that have accumulated. Sequence divergence in and around the Adh gene was used to assess inter- and intraspecies evolutionary relationships. Finally, there appears to be an unrelated structural gene located directly 3' of the Adh transcribed region.      

Modeling electroporation in a single cell. II. Effects Of ionic concentrations.

This study expands a previously developed model of a single cell electroporated by an external electric field by explicitly accounting for the ionic composition of the electroporation current. The previous model with non-specific electroporation current predicts that both the transmembrane potential V(m) and the pore density N are symmetric about the equator, with the same values at either end of the cell. The new, ion-specific case predicts that V(m) is symmetric and almost identical to the profile from the non-specific case, but N has a profound asymmetry with the pore density at the hyperpolarized end of the cell twice the value at the depolarized end. These modeling results agree with the experimentally observed preferential uptake of marker molecules at the hyperpolarized end of the cell as reported in the literature. This study also investigates the changes in intracellular ionic concentrations induced around an electroporated single cell. For all ion species, the concentrations near the membrane vary significantly, which may explain the electrical disturbances observed experimentally after large electric shocks are delivered to excitable cells and tissues.     

A new technique for staining mast cells using ferroin

We describe here a new method for specific staining of mast cells using ferroin. Different hamster tissues were fixed in 4% formalin and processed for paraffin embedding. Sections were stained with hematoxylin followed by ferroin acidified with 2.5 N sulfuric acid to pH 4.0. Mast cells stained an intense orange color that contrasted markedly with bluish violet nuclei. High contrast was also observed when ferroin colored sections were counterstained with light green instead of hematoxylin. To evaluate the specificity of the stain, hamster cheek pouch sections were stained with toluidine blue, alcian blue-safranin O, and ferroin. Quantitative evaluation of mast cells stained with the three techniques showed no statistical difference. The simplicity and selectivity of this method is sufficient for image analysis of mast cells.     

Modelling of the enzymatic kinetically controlled synthesis of cephalexin: influence of diffusion limitation

In this study the influence of diffusion limitation on enzymatic kinetically controlled cephalexin synthesis from phenylglycine amide and 7-aminodeacetoxycephalosporinic acid (7-ADCA) was investigated systematically. It was found that if diffusion limitation occurred, both the synthesis/hydrolysis ratio (S/H ratio) and the yield decreased, resulting in lower product and higher by-product concentrations. The effect of pH, enzyme loading, and temperature was investigated, their influence on the course of the reaction was evaluated, and eventually diffusion limitation was minimised. It was found that at pH >or=7 the effect of diffusion limitation was eminent; the difference in S/H ratio and yield between free and immobilised enzyme was considerable. At lower pH, the influence of diffusion limitation was minimal. At low temperature, high yields and S/H ratios were found for all enzymes tested because the hydrolysis reactions were suppressed and the synthesis reaction was hardly influenced by temperature. The enzyme loading influenced the S/H ratio and yield, as expected for diffusion-limited particles. For Assemblase 3750 (the number refers to the degree of enzyme loading), it was proven that both cephalexin synthesis and hydrolysis were diffusion limited. For Assemblase 7500, which carries double the enzyme load of Assemblase 3750, these reactions were also proven to be diffusion limited, together with the binding-step of the substrate phenylglycine amide to the enzyme. For an actual process, the effects of diffusion limitation should preferably be minimised. This can be achieved at low temperature, low pH, and high substrate concentrations. An optimum in S/H ratio and yield was found at pH 7.5 and low temperature, where a relatively low reaction pH can be combined with a relatively high solubility of 7-ADCA. When comparing the different enzymes at these conditions, the free enzyme gave slightly better results than both immobilised biocatalysts, but the effect of diffusion limitation was minimal.