Biodegradation of Bis(2-Chloroethyl) Ether by Xanthobacter sp. Strain ENV481

Degradation of bis(2-chloroethyl) ether (BCEE) was observed to occur in two bacterial strains. Strain ENV481, a Xanthobacter sp. strain, was isolated by enrichment culturing of samples from a Superfund site located in the northeastern United States. The strain was able to grow on BCEE or 2-chloroethylethyl ether as the sole source of carbon and energy. BCEE degradation in strain ENV481 was facilitated by sequential dehalogenation reactions resulting in the formation of 2-(2-chloroethoxy)ethanol and diethylene glycol (DEG), respectively. 2-Hydroxyethoxyacetic acid was detected as a product of DEG catabolism by the strain. Degradation of BCEE by strain ENV481 was independent of oxygen, and the strain was not able to grow on a mixture of benzene, ethylbenzene, toluene, and xylenes, other prevalent contaminants at the site. Another bacterial isolate, Pseudonocardia sp. strain ENV478 (S. Vainberg et al., Appl. Environ. Microbiol. 72:5218-5224, 2006), degraded BCEE after growth on tetrahydrofuran or propane but was not able to grow on BCEE as a sole carbon source. BCEE degradation by strain ENV478 appeared to be facilitated by a monooxygenase-mediated O-dealkylation mechanism, and it resulted in the accumulation of 2-chloroacetic acid that was not readily degraded by the strain.     

Role of spike-frequency adaptation in shaping neuronal response to dynamic stimuli

Spike-frequency adaptation is the reduction of a neuron’s firing rate to a stimulus of constant intensity. In the locust, the Lobula Giant Movement Detector (LGMD) is a visual interneuron that exhibits rapid adaptation to both current injection and visual stimuli. Here, a reduced compartmental model of the LGMD is employed to explore adaptation’s role in selectivity for stimuli whose intensity changes with time. We show that supralinearly increasing current injection stimuli are best at driving a high spike count in the response, while linearly increasing current injection stimuli (i.e., ramps) are best at attaining large firing rate changes in an adapting neuron. This result is extended with in vivo experiments showing that the LGMD’s response to translating stimuli having a supralinear velocity profile is larger than the response to constant or linearly increasing velocity translation. Furthermore, we show that the LGMD’s preference for approaching versus receding stimuli can partly be accounted for by adaptation. Finally, we show that the LGMD’s adaptation mechanism appears well tuned to minimize sensitivity for the level of basal input. This article is part of a special issue on Neuronal Dynamics of Sensory Coding.     

Structure and expression of ubiquitin genes of Drosophila melanogaster.

We isolated and characterized two related ubiquitin genes from Drosophila melanogaster, polyubiquitin and UB3-D. The polyubiquitin gene contained 18 repeats of the 228-base-pair monomeric ubiquitin-encoding unit arranged in tandem. This gene was localized to a minor heat shock puff site, 63F, and it encoded a constitutively expressed 4.4-kilobase polyubiquitin-encoding mRNA, whose level was induced threefold by heat shock. To investigate the pattern of expression of the polyubiquitin gene in developing animals, a polyubiquitin-lacZ fusion gene was introduced into the Drosophila genome by germ line transformation. The fusion gene was expressed at high levels in a tissue-general manner at all life stages assayed. The ubiquitin-encoding gene, UB3-D, consisted of one ubiquitin-encoding unit directly fused, in frame, to a nonhomologous tail sequence. The amino acid sequence of the tail portion of the protein had 65% positional identity with that of yeast UBI3 protein, including a region that contained a potential nucleic acid-binding motif. The Drosophila UB3-D gene hybridized to a 0.9-kilobase mRNA that was constitutively expressed, and in contrast to the polyubiquitin gene, it was not inducible by heat shock.     

Identification and activity of a series of azole-based compounds with lactate dehydrogenase-directed anti-malarial activity

Plasmodium falciparum, the causative agent of malaria, relies extensively on glycolysis coupled with homolactic fermentation during its blood-borne stages for energy production. Selective inhibitors of the parasite lactate dehydrogenase (LDH), central to NAD(+) regeneration, therefore potentially provide a route to new antimalarial drugs directed against a novel molecular target. A series of heterocyclic, azole-based compounds are described that preferentially inhibit P. falciparum LDH at sub-micromolar concentrations, typically at concentrations about 100-fold lower than required for human lactate dehydrogenase inhibition. Crystal structures show these competitive inhibitors form a network of interactions with amino acids within the active site of the enzyme, stacking alongside the nicotinamide ring of the NAD(+) cofactor. These compounds display modest activity against parasitized erythrocytes, including parasite strains with known resistance to existing anti-malarials and against Plasmodium berghei in BALB/c mice. Initial toxicity data suggest the azole derivatives have generally low cytotoxicity, and preliminary pharmoco-kinetic data show favorable bioavailability and circulation times. These encouraging results suggest that further enhancement of these structures may yield candidates suitable for consideration as new therapeutics for the treatment of malaria. In combination these studies also provide strong support for the validity of targeting the Plasmodium glycolytic pathway and, in particular, LDH in the search for novel anti-malarials.     

Immunosenescence in wild animals: meta‐analysis and outlook

Immunosenescence, the decline in immune defense with age, is an important mortality source in elderly humans but little is known of immunosenescence in wild animals. We systematically reviewed and meta‐analysed evidence for age‐related changes in immunity in captive and free‐living populations of wild species (321 effect sizes in 62 studies across 44 species of mammals, birds and reptiles). As in humans, senescence was more evident in adaptive (acquired) than innate immune functions. Declines were evident for cell function (antibody response), the relative abundance of naïve immune cells and an in vivo measure of overall immune responsiveness (local response to phytohaemagglutinin injection). Inflammatory markers increased with age, similar to chronic inflammation associated with human immunosenescence. Comparisons across taxa and captive vs free‐living animals were difficult due to lack of overlap in parameters and species measured. Most studies are cross‐sectional, which yields biased estimates of age‐effects when immune function co‐varies with survival. We therefore suggest longitudinal sampling approaches, and highlight techniques from human cohort studies that can be incorporated into ecological research. We also identify avenues to address predictions from evolutionary theory and the contribution of immunosenescence to age‐related increases in disease susceptibility and mortality.     

Synthesis of Bacterial Flagella: Chromosomal Synchrony and Flagella Synthesis

Synchronous cultures of Bacillus subtilis 168 M were obtained from light-density spores germinated at 46 C and grown at 37 C. This procedure synchronizes both cell division and chromosome replication. The chromosome synchrony was demonstrated by using transformation to measure changes in marker frequency during the cell cycle. The synthesis of two enzymes and of bacterial flagellar protein was also followed. All of the proteins were found to be synthesized continuously with an abrupt doubling in the rate of synthesis at a specific time in the cell cycle. The time at which the doubling occurred for each enzyme corresponded to the time at which the structural gene for the enzyme was replicated. The doubling of the rate of flagella synthesis corresponded to the time of replication of the hisA1 gene. We conclude that the genetic locus for the factors involved in the rate-limiting steps in flagella synthesis are located on the genetic map near the hisA1 locus.     

1-Methoxy-, 1-deoxy-11-hydroxy- and 11-Hydroxy-1-methoxy-Δ-tetrahydrocannabinols: new selective ligands for the CB2 receptor

Three series of new cannabinoids were prepared and their affinities for the CB₁ and CB₂ cannabinoid recptors were determined. These are the 1-methoxy-3-(1′,1′-dimethylalkyl)-, 1-deoxy-11-hydroxy-3-(1′,1′-dimethylalkyl)- and 11-hydroxy-1-methoxy-3-(1′,1′-dimethylalkyl)-Δ⁸-tetrahydrocannabinols, which contain alkyl chains from dimethylethyl to dimethylheptyl appended to C-3 of the cannabinoid. All of these compounds have greater affinity for the CB₂ receptor than for the CB₁ receptor, however only 1-methoxy-3-(1′,1′-dimethylhexyl)-Δ⁸-THC (JWH-229, 6e) has effectively no affinity for the CB₁ receptor (K_i=3134±110 nM) and high affinity for CB₂ (K_i=18±2 nM).     

The overlooked role of a biotin precursor for marine bacteria - desthiobiotin as an escape route for biotin auxotrophy

Biotin (vitamin B₇) is involved in a wide range of essential biochemical reactions and a crucial micronutrient that is vital for many pro- and eukaryotic organisms. The few biotin measurements in the world’s oceans show that availability is subject to strong fluctuations. Numerous marine microorganisms exhibit biotin auxotrophy and therefore rely on supply by other organisms. Desthiobiotin is the primary precursor of biotin and has recently been detected at concentrations similar to biotin in seawater. The last enzymatic reaction in the biotin biosynthetic pathway converts desthiobiotin to biotin via the biotin synthase (BioB). The role of desthiobiotin as a precursor of biotin synthesis in microbial systems, however, is largely unknown. Here we demonstrate experimentally that bacteria can overcome biotin auxotrophy if they retain the bioB gene and desthiobiotin is available. A genomic search of 1068 bacteria predicts that the biotin biosynthetic potential varies greatly among different phylogenetic groups and that 20% encode solely bioB and thus can potentially overcome biotin auxotrophy. Many Actino- and Alphaproteobacteria cannot synthesize biotin de novo, but some possess solely bioB, whereas the vast majority of Gammaproteobacteria and Flavobacteriia exhibit the last four crucial biotin synthesis genes. We detected high intra- and extracellular concentrations of the precursor relative to biotin in the prototrophic bacterium, Vibrio campbellii, with extracellular desthiobiotin reaching up to 1.09 ± 0.15*10⁶ molecules per cell during exponential growth. Our results provide evidence for the ecological role of desthiobiotin as an escape route to overcome biotin auxotrophy for bacteria in the ocean and presumably in other ecosystems.Subject terms: Water microbiology, Ecosystem ecology, Marine microbiology