PfeR, an enterobactin-responsive activator of ferric enterobactin receptor gene expression in Pseudomonas aeruginosa.

PfeR (Regulator) and PfeS (Sensor), members of the superfamily of so-called two-component regulatory protein pairs, are required for the enterobactin-inducible production of the ferric enterobactin receptor (PfeA) in Pseudomonas aeruginosa. A pfeR knockout mutant failed to demonstrate enterobactin-inducible expression of a pfeA-lacZ fusion, indicating that PfeR acts at the level of pfeA gene expression. Consistent with this, PfeR overexpressed in P. aeruginosa bound, in bandshift assays, the promoter region of pfeA. Such binding was enhanced when PfeR-containing extracts were prepared from cells cultured in the presence of enterobactin, consistent with a model of PfeR as an enterobactin-responsive activator of pfeA expression. A region showing homology to the consensus binding sequence for the global iron repressor Fur was identified upstream of pfeR, suggesting that the pfeRS operon is iron regulated. As expected, expression of a pfeR-lacZ fusion in P. aeruginosa was increased under conditions of iron limitation. Enterobactin failed, however, to provide any enhancement of pfeR-lacZ expression under iron-limiting conditions, indicating that PfeR does not positively regulate pfeRS expression. A pfeA knockout mutant demonstrated enterobactin-inducible expression of a pfeA-lacZ fusion, indicating that the receptor is not required for the enterobactin inducibility of pfeA gene expression. Such mutants show growth, albeit reduced, in enterobactin-supplemented iron-limiting minimal medium, indicating that a second route of uptake across the outer membrane exists for ferric enterobactin in P. aeruginosa and may be important for the initial induction of pfeA in response to enterobactin.     

Cyclothiazide Unmasks an AMPA-Evoked Release of Arachidonic Acid from Cultured Striatal Neurones

Abstract: The joint, but not independent, activation of α-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) and metabotropic glutamate receptors induces liberation of arachidonic acid from cultured mouse striatal neurones. We examined whether blocking AMPA receptor desensitisation with cyclothiazide would modify this response. Cyclothiazide strongly potentiated the combined AMPA/(1 S ,3 R )-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD)-evoked release of arachidonic acid (EC₅₀ of ∼7 µ M ) but did not modulate the basal, ACPD, or NMDA response. The enhanced liberation of arachidonic acid, observed in the presence of cyclothiazide, was due to the appearance of a genuine AMPA response that was independent of an associative activation of metabotropic receptors. The potentiated and nonpotentiated AMPA responses were inhibited by both competitive [2,3-dihydroxy-6-nitro-7-sulphamoylbenzo( f )quinoxaline] and 2,3-benzodiazepine noncompetitive (GYKI 53655 and GYKI 52466) receptor antagonists. Cyclothiazide was equally effective at potentiating the AMPA response in either the presence or absence of glucose, suggesting that the increased glutamate-evoked arachidonic acid release observed in these cells under conditions of glucose deprivation is not due to reduced AMPA receptor desensitisation. The enhanced liberation of arachidonic acid measured in the presence of cyclothiazide appeared to result from a large (fourfold) elevation of the AMPA-induced increase in intracellular calcium level. Therefore, an AMPA-evoked mobilisation of arachidonic acid could potentially contribute to non-NMDA receptor-mediated neurotoxicity, which has been observed in neuronal cells in the presence of cyclothiazide.     

Biogeography of milkweed butterflies (Nymphalidae: Danainae) and mimetic patterns on tropical Paciflc archipelagos

Distributions of danaine butterfly species and associated mimetic patterns were compared among fifteen archipelagos of the tropical Pacific Ocean, and within five major archipelagos (the Bismarcks, Fiji, East and West Solomon Islands, and Vanuatu). Using both simple and stepwise linear regression analysis, variation in the total number of danaine species and number of mimetic patterns was assessed with respect to island size, isolation and elevation. Relative to interarchipelago distributions, the distribution of danaine species and number of mimetic patterns on islands within archipelagos exhibited less dependence upon interisland distance and island area. Geographical features influencing the number of mimetic patterns were similar to those of danaines as a whole. Analysis of residuals from stepwise linear regression suggested that factors influencing danaine distributions were different from those for non-danaine butterflies. This result is consistent with the hypothesis of enhancement of danaine species establishment through Müllerian mimicry, although other factors such as host plant availability and similar habitat use may also be important.     

Isozyme analysis of Galaxias species (Teleostei: Galaxiidae) from the Taieri River, South Island, New Zealand: a species complex revealed

We examined genetic differentiation among 23 samples of non-migratory river galaxias from 17 streams in the Taieri River system, South Island, New Zealand. Four major genetic types were found, two of which occur in narrow sympatry in one location. These were compared with topotypical material representing Galaxias anomalus from the Clutha system (Otago) and G. vulgaris from the Waimakariri system (Canterbury) in order to establish identity. Morphological examination of these four major genetic types revealed consistent concomitant differences. The results suggest that there are at least three species of river galaxias in the Taieri system: G. anomalus, G. vulgaris and at least one previously undescribed species. We propose that the genetic structuring and subsequent speciation of this group has been promoted by the absence of the marine juvenile phase that is found in five other members of the genus native to New Zealand. This structuring may be exacerbated by population fragmentation over the last century owing to the negative influence of introduced trout. The phylogenetic diversity within the river system mirrors the diverse flora and invertebrate fauna of the region, and has conservation implications that parallel those resulting from our improved knowledge of the New Zealand herpetofauna through the application of genetic analysis.     

Determination of half-reaction equilibrium in a ping-pong enzyme mechanism

Substituted enzyme (or ping-pong) mechanisms usually involve enzymes that exist in two forms that alternate during the catalytic reaction. A method is described here for determining the position of the equilibrium of a half reaction in a ping-pong enzyme mechanism that is based on the kinetics of the burst reaction which occurs upon addition of reactants that recycle the enzyme from one form to another. The theoretical basis for the analysis is developed, and the method is applied to the half reaction of the aldimine form of aspartate transaminase with difluoro-oxaloacetate. Special issue dedicated to Herman Bachelard     

Changes in extracellular levels of dopamine metabolites in somatosensory cortex after peripheral denervation

This study examined the effects of a nerve transection on monoamine release from primary somatosensory cortex. The technique of microdialysis was employed to sample extracellular levels of norepinephrine (NE), 3,4-dihydroxyphenylacetic acid (DOPAC), 5-hydroxyindole-3-acetic acid (5-HIAA) and homovanillic acid (HVA) in the barrel field of freely moving rats following the surgical transection of the contralateral infraorbital nerve. Microdialysates obtained 3, 4, and 5 days after deafferentation were analyzed using high-performance liquid chromatography with electrochemical detection. We found a significant increase in the release of the dopamine metabolites, DOPAC and HVA from the deafferented cortex. Three days after deafferentation the release of DOPAC was three-fold higher in the deafferented than in the control animals, and remained about 100% higher in the next two days in this group of animals. The release of HVA showed a gradual increase following the deafferentation procedure, since a 92% larger value on day 3 increased to a 338% difference on day 5. On the other hand, the release rate of NE and the levels of the serotonin metabolite 5-HIAA were not significantly affected by the deafferentation procedure. These results are discussed in the context of the possible participation of dopamine in the reorganization of the deafferented somatosensory cortex.     

Recent developments in vitamin E nutrition of turkeys

Research and field observations have shown that vitamin E status of poults is often inadequate during the 5 to 21-day posthatching period. Although overt consequences of this inadequacy may not be evident and clear-cut, subtle adverse effects on health and metabolic efficiency are probable. These latter effects can result in uneconomical performance of turkeys. It therefore seems advisable to supplement the first diet (starter or prestarter) of poults with 100 to 150 IU of vitamin E. The cost of doing so will contribute little to the total cost of production and may facilitate more efficient turkey production. In the meantime, other alternatives that may be effective and economical need to be evaluated.     

Hydroperoxide inactivation of enzymes within spores of Bacillus megaterium ATCC19213

Abstract Hydroperoxide inactivation of the protoplast enzymes enolase, aldolase and glucose-6-phosphate dehydrogenase in intact spores of Bacillus megaterium ATCC19213 was assessed by first treating the cells with lethal levels of H₂O₂, then germinating them in the presence of chloramphenicol prior to permeabilization and enzyme assays. Glucose-6-phosphate dehydrogenase proved to be more sensitive to H₂O₂than enolase or aldolase, in agreement with findings for isolated enzymes. Average D values (time for 90% inactivation) for spores treated with 0.50% H₂O₂ were 173 min for enolase, 67 min for aldolase and 32 min for glucose-6-phosphate dehydrogenase, compared with a D value of 34 min for spore killing. H₂O₂ killing of spores was found to be conditional in that recoveries of survivors were greater on complex medium than on minimal medium. Overall, it appeared that oxidative inactivation of enzymes may be important for hydroperoxide killing of spores.     

Characterization of the eaeA gene from rabbit enteropathogenic Escherichia coli strain RDEC-1 and comparison to other eaeA genes from bacteria that cause attaching-effacing lesions

Abstract A number of enteric pathogens, including enteropathogenic (EPEC) and enterohemorrhagic (EHEC) Escherichia coli , Hafnia alvei , a strain of Citrobacter freundii , and rabbit EPEC strain RDEC-1 cause attaching-effacing (AE) lesions in the gut mucosa. These bacteria have a pathogenicity cassette (locus of enterocyte effacement or LEE) containing the eaeA gene. This gene encodes intimin, an outer membrane protein required for production of AE lesions. RDEC-1, a non-invasive enteropathogen in young rabbits, produces AE lesions morphologically indistinguishable from lesions caused by human AE bacterial strains. The RDEC-1 example of E. coli diarrhea in rabbits is an important model for studying the pathogenesis of AE bacteria in a natural infection and for analyzing specific roles of the components of LEE. In order to better understand the role of intimin in the development of AE lesions, a portion of DNA within RDEC-1 LEE, containing the eaeA gene and an upstream open reading frame (ORF), was sequenced. The RDEC-1 eaeA gene shared 87%, 92%, and 93% DNA sequence identity and > 80% amino acid sequence identity with the eaeA genes of C. freundii biotype 4280, EHEC O157:H7, and EPEC O127:H6, respectively. The carboxy-terminal 280 amino acid residues of intimin has 80%, 56%, and 54% identity with C. freundii , EHEC O157:H7, and EPEC O127:H6 intimins, respectively. The predicted protein encoded by the upstream ORF (156 amino acids) shares 95%, 97%, and 99% amino acid identity with predicted proteins from C. freundii , EHEC O157:H7, and EPEC O127:H6, respectively. The high degree of sequence homology of the ORF and the eaeA gene of RDEC-1 with those of other AE bacteria suggests an evolutionary relationship of LEE and supports and facilitates the use of the RDEC-1 model for studying the role of LEE in pathogenesis.