The detection of fish pathogenVibrio anguillarum in water and fish using a species-specific DNA probe combined with membrane filtration

The marine bacteriumVibrio anguillarum causes disease in fish worldwide and is particularly devastating in aquaculture. Little is known about the ecology ofV. anguillarum in the environment and how this may relate to the pathogenicity of this organism. Combining membrane filtration and a species-specific DNA probe, culturableV. anguillarum cells were detected in water from three habitats and in chinook salmon (Onchorynchus tshawytscha) tissue samples. Results show that different marine habitats have a marked effect on cell numbers and that water temperature may play a role in the culturability and distribution ofV. anguillarum. Vibrio anguillarum was detected from the gills of salmon within 24 h of transfer of fingerlings from freshwater to seawater, with cell numbers reaching a concentration of 1.9 × 10² cells g^–1 tissue 28 days post transfer.Vibrio anguillarum cell numbers were low in the colon throughout the study, andV. anguillarum was not detected in healthy kidney samples. The methodology reported in this paper allows the accurate quantification of culturableV. anguillarum cells and has allowed a preliminary study of the ecology of this species.     

Development of a DNA probe using differential hybridization to detect the fish pathogenVibrio anguillarum

The fish pathogenVibrio anguillarum causes significant economic losses in commercially cultured fish species worldwide. At present, identification ofV. anguillarum requires conventional isolation and culturing techniques. Using differential hybridization, a 310 base pairV. anguillarum-specific DNA fragment was isolated for use as a probe. In specificity studies against 19 different bacterial species, including twoVibrio sp. and fish pathogens, and 223 marine bacterial isolates, the probe hybridized exclusively toV. anguillarum strains. The probe also strongly hybridizes to 7 of 9 serotypes tested, with serotype 09 giving a weak probe reaction and serotype O7 negative. The probe allows rapid and accurate detection of both pathogenic and environmental strains ofV. anguillarum.     

Activity of phosphoenolpyruvate carboxylase of an anthracycline-producing streptomycete

During fermantation studies on the production of anthracycline antibiotics by Streptomyces C5, it was observed that among the intermediate metabolism enzymes tested, only phosphoenolpyruvate carboxylase (PEPCase; EC 4.1.1.31) increased significantly in specific activity during stationary phase. The specific activity of the Streptomyces C5 PEPCase increased ca. 3-fold during antibiotic production phase from the logarithmic phase levels. To characterize the regulation of the enzyme further, the Streptomyces C5 PEPCase was purified 150-fold from crude extracts. Acetyl-CoA and Mg2+ were shown to be required for PEPCase activity. The activity of the partially purified PEPCase was stimulated slightly by fructose 1,6-bisphosphate and AMP, and was inhibited severely by oxaloacetate, aspartate, malate, succinate, ATP, citrate, and CoASH.     

Habenular modulation of raphe indoleamine metabolism

Anatomical and neurophysiological findings have demonstrated neuronal connections between the diencephalic habenular nuclei and brain stem serotonergic raphe nuclei. Therefore we examined some neurochemical consequences of habenular lesions. Sixteen hours and one week after bilateral lesions serotonin metabolism (as reflected by concentrations of its metabolite, 5-hydroxyindoleacetic acid) was significantly increased in the dorsal but not the median raphe nuclei. Unilateral lesions produced a proportionally smaller augmentation. Motron locomotor activity was enhanced during the light and dark illumination phases in lesioned animals but only attained statistical significance during the day.     

Inhibition by S-adenosyl-L-homocysteine of dopamine dependent adenylate cyclase in rat retina

S-adenosyl-L-homocysteine (S-AH), a potent inhibitor of biological transmethylation, decreased the response of rat retina adenylate cyclase to dopamine and to 2-amino-6, 7-dihydroxytetrahydronaphtalene (ADTN). This effect appeared for 10^?7M of S-adenosyl-L-homocysteine and was linear for concentration ranging to 10^?4M. S-adenosyl-L-homocysteine did not decrease the cyclic AMP accumulation with sodium fluoride, a non specific adenylate cyclase activator. On the other hand, the incorporation of methyl group was reduced in rat retina homogenates by S-adenosyl-L-homocysteine. These findings suggest that the activity of the dopamine dependent adenylate cyclase is linked to a methylation process.     

Nonintegrative transformation in the filamentous fungus Podospora anserina: stabilization of a linear vector by the chromosomal ends of Tetrahymena thermophila.

The effect of the chromosomal ends of Tetrahymena thermophila on the stability of linear transforming molecules in the filamentous fungus Podospora anserina was tested. A derivative of an integrative vector for this fungus has been constructed, so that after linearization, the ends of the plasmid are the telomeric sequences of T. thermophila. After transformation, this linear molecule was maintained as an extrachromosomal plasmid with no integrated copies in about 50% of the transformants. Under selective conditions, there was approximately one linear molecule per 5 to 10 nuclei, and these extrachromosomal molecules were rapidly lost under nonselective conditions. The circular plasmid carrying an inverted repeat of T. thermophila telomeres could be linearized and processed in vivo.     

Maleylated-BSA suppresses IFN-gamma-mediated Ia expression in murine peritoneal macrophages

Maleylated bovine serum albumin (maleyl-BSA) and other polyanionic polymers that are recognized by cell surface receptors on macrophages have been shown to induce chemotaxis, protease secretion, and tumoricidal function in this cell type. In this paper the effect of maleyl-BSA on Ia antigen expression has been evaluated. In a fashion similar to LPS, maleyl-BSA suppressed IFN-gamma-induced expression of Ia in a time- and dose-dependent manner. Also like LPS, maleyl-BSA stimulated the production and secretion of substantial amounts of PGE2 over a 24-hr period. This did not, however, appear to be the primary mechanism by which expression of Ia was suppressed, because co-treatment of the cells with indomethacin, which totally inhibited the production of PGE2, only minimally affected the suppressive activity. Surprisingly, the suppressive activity of both maleyl-BSA and LPS could be largely abrogated by co-treatment of the cells with cyclohexamide during the time period when Ia expression was sensitive to suppression. This effect was selective in that PGE2- or dibutyryl cyclic AMP-induced suppression of Ia expression was not affected by cyclohexamide treatment. The data support the concept that there are multiple molecular mechanisms involved in the negative regulation of IFN-gamma-induced Ia expression in macrophages. Such mechanisms may include, in addition to the synthesis of PGE2 and consequent elevation in intracellular levels of cyclic AMP, one or more proteins made early after treatment with either maleyl-BSA or LPS. Thus the function of some of these early gene products may be to regulate expression of functional genes such as that encoding Ia antigen.