Exposure of fish to high‐intensity sonar does not induce acute pathology

This study investigated immediate effects of intense sound exposure associated with low‐frequency (170–320 Hz) or with mid‐frequency (2·8–3·8 kHz) sonars on caged rainbow trout Oncorhynchus mykiss, channel catfish Ictalurus punctatus and hybrid sunfish Lepomis sp. in Seneca Lake, New York, U.S.A. This study focused on potential effects on inner ear tissues using scanning electron microscopy and on non‐auditory tissues using gross and histopathology. Fishes were exposed to low‐frequency sounds for 324 or 628 s with a received peak signal level of 193 dB re 1 µPa (root mean square, rms) or to mid‐frequency sounds for 15 s with a received peak signal level of 210 dB re 1 µPa (rms). Although a variety of clinical observations from various tissues and organ systems were described, no exposure‐related pathologies were observed. This study represents the first investigation of the effects of high‐intensity sonar on fish tissues in vivo. Data from this study indicate that exposure to low and midfrequency sonars, as described in this report, might not have acute effects on fish tissues.     

Effects of genotype, explant source, and plant growth regulators on indirect shoot organogenesis in Dieffenbachia cultivars

The capacity for indirect shoot organogenesis of leaf and root explants of four Dieffenbachia cultivars were examined on a modified Murashige and Skoog (MS; Physiol Plant 15:473–495, 1962) medium supplemented with different plant growth regulators in 112 combinations. Callus formation was only observed from leaf explants on MS supplemented with 1–10 μM thidiazuron (TDZ) and 0.5–1.0 μM 2,4-dichlorophenoxyacetic acid (2,4-D) regardless of cultivars. The combination of 5 μM TDZ and 1 μM 2,4-D resulted in the greatest callus formation frequency among the four cultivars tested. Significant differences in callus and shoot formation from leaf explants were also observed among cultivars. Cultivars Camouflage, Camille, Octopus, and Star Bright produced green nodular, brown nodular, yellow friable, and green compact calli with corresponding maximum callus formation frequencies of 96%, 62%, 54%, and 52%, respectively. A maximum of 6.7 shoots/callus was observed in cv. Camouflage, followed by cvs. Camille and Star Bright at 3.7 and 3.5, respectively. Calli of cv. Octopus displayed no capacity for shoot organogenesis. Regardless of cultivar, callus formation was not observed on root explants. Regenerated shoots were successfully acclimatized in a shaded greenhouse condition with 100% survival.     

Enzymatic recycling-based amperometric immunosensor for the ultrasensitive detection of okadaic acid in shellfish

Electrochemical immunosensors based on a competitive indirect enzyme-linked immunosorbent assay (ciELISA) and an enzymatic recycling system were developed for the detection of okadaic acid (OA). OA-ovalbumin (OA-OVA) conjugate was immobilised on screen-printed electrodes (SPEs) and competition of a newly generated monoclonal antibody (MAb) for free and immobilised OA was subsequently performed. Secondary antibodies labelled with alkaline phosphatase (ALP) or horseradish peroxidase (HRP) were used for signal generation. Experimental parameters were firstly optimised by colorimetric ELISA on microtiter wells and on SPEs. The ELISA system was then tested by amperometry at +300 mV vs. Ag/AgCl (detection of p-aminophenol produced by the reaction of p-aminophenyl phosphate with ALP) or -200 mV vs. Ag/AgCl (detection of 5-methyl-phenazinium methyl sulfate, redox mediator in the HRP bioelectrocatalysis). The limits of detection (LODs) with standard solutions were 1.07 and 1.98 microgL(-1) when using ALP and HRP labels, respectively. An electrochemical signal amplification system based on diaphorase (DI) recycling was integrated into the ALP-based immunosensor, decreasing the LOD to 0.03 microgL(-1) and enlarging the working range by two orders of magnitude. Preliminary results with mussel and oyster extracts were obtained and compared with the colorimetric immunoassay, the colorimetric protein phosphatase inhibition assay (PPIA) and LC-MS/MS.     

Genetics and evolution of weedy Helianthus annuus populations: adaptation of an agricultural weed

Agricultural weeds are a major cost to economies throughout the world, and have evolved from numerous plant species in many different plant families. Despite their ubiquity, we do not yet know how easily or often weeds evolve from their wild ancestors or the kinds of genes underlying their evolution. Here we report on the evolution of weedy populations of the common sunflower Helianthus annuus. We analysed 106 microsatellites in 48 individuals from each of six wild and four weed populations of the species. The statistical tests lnRV and lnRH were used to test for significant reductions in genetic variability at each locus in weedy populations compared to nearby wild populations. Between 1% and 6% of genes were significant outliers with reduced variation in weedy populations, implying that a small but not insignificant fraction of the genome may be under selection and involved in adaptation of weedy sunflowers. However, there did not appear to be a substantial reduction in variation across the genome, suggesting that effective population sizes have remained very large during the recent evolution of these weedy populations. Additional analyses showed that weedy populations are more closely related to nearby wild populations than to each other, implying that weediness likely evolved multiple times within the species, although a single origin followed by gene flow with local populations cannot be ruled out. Together, our results point to the relative ease with which weedy forms of this species can evolve and persist despite the potentially high levels of geneflow with nearby wild populations.     

Synthesis and evaluation of xanomeline analogs--probing the wash-resistant phenomenon at the M1 muscarinic acetylcholine receptor

A series of xanomeline analogs were synthesized and evaluated for binding at the M(1) muscarinic acetylcholine receptor (M(1) receptor). Specifically, compounds that substitute the O-hexyl chain of xanomeline with polar, ionizable, or conformationally restricted moieties were assessed for their ability to bind to the M(1) receptor in a wash-resistant manner (persistent binding). From our screen, several novel ligands that persistently bind to the M(1) receptor with greater affinity than xanomeline were discovered. Results indicate that persistent binding may arise not only from hydrophobic interactions but also from ionic interactions with a secondary M(1) receptor binding site. Herein, a qualitative model that accounts for both binding scenarios is proposed and applied to understand the structural basis to wash-resistant binding and long-acting effects of xanomeline-based compounds.     

Biosynthesis and Regulation of the Yeast Vacuolar H+-ATPase

The yeast V-ATPase is highly similar to V-ATPases of higher organismsand has proved to be a biochemically and genetically accessible model formany aspects of V-ATPase function. Like other V-ATPases, the yeast enzymeconsists of a complex of peripheral membrane proteins, the V₁sector, attached to a complex of integral membrane subunits, theV₀ sector. Multiple pathways for biosynthetic assembly of theenzyme appear to be available to cells containing a full complement ofsubunits and enzyme activity may be further controlled during biosynthesis bya protease activity localized to the late Golgi apparatus. Surprisingly, theassembled V-ATPase is not a static structure. Instead, fully assembledV₁V₀ complexes appear to exist in a dynamic equilibriumwith inactive cytosolic V₁ and membrane-bound V₀complexes and this equilibrium can be rapidly shifted in response to changesin carbon source. The reversible disassembly of the yeast V-ATPase may be anovel regulatory mechanism, common to V-ATPases, that works in vivoin coordination with many other regulatory mechanisms.     

Role of Hpn and NixA of Helicobacter pylori in Susceptibility and Resistance to Bismuth and Other Metal Ions

Background. Helicobacter pylori produces Hpn, a 60-amino acid, histidine-rich protein that avidly binds nickel and zinc ions, and NixA, a high-affinity nickel transporter in the cytoplasmic membrane. We tested the hypothesis that Hpn and NixA govern susceptibility to metal ions in H. pylori. Materials and Methods. Hpn-negative mutants of four H. pylori strains were constructed by standard allelic exchange techniques to yield isogenic Hpn⁺/Hpn-deficient pairs. A metal concentration that inhibited growth by 50% (IC₅₀) was calculated for Ni²⁺, Zn²⁺, Cu²⁺, and Co²⁺ by comparing OD₆₀₀ of cultures in metal-supplemented and control media. Results. Among all four pairs of isogenic strains, the tolerance for Ni²⁺ was reduced significantly (p < .001) in the Hpn mutants; the mean IC₅₀ value for wild-type strains was 1.9 mM; for the mutant, it was 0.8 mM. In  contrast, growth inhibition by Zn²⁺ was identical within the fours pairs, as was Cu²⁺ and Co²⁺ tolerance in one pair tested. We also found that deletion of the hpn gene increases susceptibility to therapeutic forms of bismuth by testing a mutant and wild-type pair with ranitidine bismuth citrate, bismuth citrate, and four antibiotics. Minimal inhibitory concentrations of ranitidine bismuth citrate dropped from 9.2 to 2.3 μg/ml, and those of bismuth citrate dropped from 7.4 to 3.2 μg/ml (p < .05 for both comparisons), while susceptibility to the antibiotics was unaffected. Disruption of the nixA gene encoding the specific Ni²⁺ transport protein of H. pylori did not change susceptibility to bismuth. Conclusion. We concluded that bacteria lacking Hpn, cultured in vitro, are more susceptible than is the wild type to bismuth and Ni²⁺.     

Optimisation of the P2 pharmacophore in a series of thrombin inhibitors: ion-dipole interactions with lysine 60G.

The optimisation of the P2 pharmacophore in a series of thrombin inhibitors is described. The interaction of a number of piperidine P2 functionalities with lysine 60G of thrombin is explored with reference to the crystal structure of inhibitor enzyme complexes. A primary ion-dipole interaction between the terminal P2 side chain group and lysine 60G is evoked to explain the SAR in this series.