Genotype Analysis of Escherichia coli Strains Isolated from Children and Chickens Living in Close Contact

Escherichia coli isolates from rectal swabs from 62 chickens and stools from 42 children living in close contact with chickens on the same farms in Kiambu district, Kenya, were compared for their genetic relatedness. Antibiotic susceptibility profiles broadly categorized isolates from the children and from the chickens into two separate clusters: the majority (144; 85.5%) of the E. coli isolates from children were multidrug resistant, while the majority (216; 87.1%) of the E. coli isolates from chickens were either fully susceptible or resistant only to tetracycline. Sixty- and 100- to 110-MDA plasmids were found to encode the transferable resistance to co-trimoxazole and tetracycline. HindIII restriction endonuclease digestion of the 60- and 100- to 110-MDA plasmids produced four distinct patterns for isolates from children and three distinct patterns for isolates from chickens. XbaI digestion of genomic DNA followed by pulsed-field gel electrophoresis (PFGE) analysis produced 14 distinct clusters. There were six distinct PFGE clusters among the isolates from children, while among the isolates from chickens there were seven distinct clusters. Only one PFGE cluster contained isolates from both children and chickens, with the isolates displaying an approximately 60% coefficient of similarity. This study showed that although several different genotypes of E. coli were isolated from children and chickens from the same farms, the E. coli strains from these two sources were distinct.     

Histamine metabolism in the visual system of the horseshoe crab Limulus polyphemus

There is now strong evidence that arthropod photoreceptors use histamine as a neurotransmitter. The synthesis, storage and release of histamine from arthropod photoreceptors have been demonstrated, and the postsynaptic effects of histamine and the endogenous neurotransmitter are similar. However, a full understanding of these photoreceptor synapses also requires knowledge of histamine inactivation and metabolism. Relatively little is known about histamine metabolism in the nervous system of arthropods, and mechanisms appear to differ with the species. This study focuses on histamine metabolism in visual tissues of the horseshoe crab Limulus polyphemus, a chelicerate. We present two major findings: (1) histamine is metabolized to imidazole acetic acid and to gamma-glutamyl histamine. (2) relatively low levels of histamine metabolites accumulate in Limulus visual tissues.     

Prevalence and genetic diversity of Campylobacter spp. in environmental water samples from a 100-square-kilometer predominantly dairy farming area

Water samples were taken systematically from a 100-km2 area of mainly dairy farmland in northwestern England and examined for Campylobacter spp. Pulsed-field gel electrophoresis-restriction fragment length polymorphism (PFGE-RFLP) and flaA strain typing of Campylobacter jejuni and Campylobacter coli isolates were done. Data on the water source and the adjacent environment were recorded and examined as explanatory variables. Campylobacter spp. were isolated from 40.5% (n = 119) of the water samples tested. C. jejuni was isolated from 14.3%, C. coli was isolated from 18.5%, and Campylobacter lari was isolated from 4.2% of the samples. Campylobacter hyointestinalis was not isolated from any water source. The difference in prevalence between water types (trough, running, and standing) was significant (P = 0.001). C. jejuni was the species most commonly isolated from trough-water and running-water sources, while C. coli was the most frequently isolated from standing water (P < 0.001). No association was found between the presence of Escherichia coli and that of Campylobacter spp. The final multivariable logistic regression model for Campylobacter spp. included the following variables: water source, soil type, aspect, and amount of cattle fecal material in the environment (fecal pat count). Strain typing demonstrated a diverse population of C. jejuni and the presence of a common C. coli flaA type that was widely distributed throughout the area. Most of the isolates within the common flaA type were discriminated by PFGE-RFLP. These findings suggest a possible role for environmental water in the epidemiology of Campylobacter spp. in a farming environment.     

Evolutionary stability of egg trading and parceling in simultaneous hermaphrodites: the chalk bass revisited

Several species of simultaneously hermaphroditic seabasses living on coral reefs mate by alternating male and female roles with a partner. This is known as egg trading, one of the classic and most widely cited examples of social reciprocity among animals. Some of the egg-trading seabass species, including the chalk bass, Serranus tortugarum, switch mating roles repeatedly, having subdivided their clutch of eggs into parcels offered to the partner for fertilization. Here we attempt to understand these dynamics as a pair of evolutionary games, modifying some previous approaches to better reflect the biological system. We find that the trading of egg clutches is evolutionarily stable via byproduct mutualism and resistant to invasion by rare individuals that take the male role exclusively. We note why and how parceling may reflect sexual conflict between individuals in the mating pair. We estimate evolutionarily stable parcel numbers and show how they depend on parameter values. Typically, two or more sequential parcel numbers are evolutionarily stable, though the lowest of these yields the highest fitness. Assuming that parcel numbers are adjusted to local conditions, we predict that parcel numbers in nature are inversely related both to mating group density (except at low density) and predation risk.     

Glycoproteins bound to ion channels mediate detection of electric fields: a proposed mechanism and supporting evidence

The mechanism by which animals detect weak electric and magnetic fields has not yet been elucidated. We propose that transduction of an electric field (E) occurs at the apical membrane of a specialized cell as a consequence of an interaction between the field and glycoproteins bound to the gates of ion channels. According to the model, a glycoprotein mass (M) could control the gates of ion channels, where M > 1.4 x 10(-18)/E, resulting in a signal of sufficient strength to overcome thermal noise. Using the electroreceptor organ of Kryptopterus as a mathematical and experimental model, we showed that at the frequency of maximum sensitivity (10 Hz), fields as low as 2 microV/m could be detected, and that the observation could be explained if a glycoprotein mass of 0.7 x 10(-12) kg (a sphere 11 microm in diameter) were bound to channel gates. Antibodies against apical membrane structures in Kryptopterus blocked field transduction, which was consistent with the proposal that it occurred at the membrane surface. Although the target of the field was hypothesized to be an ion channel, the proposed mechanism can easily be extended to include other kinds of membrane proteins.     

Voltammetric behaviour of DNA bases at a screen-printed carbon electrode and its application to a simple and rapid voltammetric method for the determination of oxidative damage in double stranded DNA

Screen-printed carbon electrodes (SPCEs) have been investigated as possible sensors to identify gamma-irradiation induced oxidative damage in double stranded (ds) DNA. Studies were undertaken to explore the possibility of using both cyclic voltammetry and differential pulse voltammetry to identify changes due to oxidative damage. Initially, guanine, adenine and 8-oxoguanosine were examined and it was found possible to differentiate them from their voltammetric responses. The voltammetric response of 8-oxoguanosine was found to be linear over the concentration range 1-400 microM, with a slope of 0.0296 microA microM(-1) (R2 value of 0.9984), in the presence of 2mM concentrations of guanine and adenine. Investigations were made into harnessing these findings to identify oxidative damage in gamma-irradiated dsDNA. The presence of oxidative damage in these samples was readily identifiable, and the magnitude of the voltammetric response was found to be dose dependant (R2=0.9919). A simple sample preparation step involving only the dissolution of double stranded DNA sample in the optimised electrolyte (0.1M acetate buffer pH 4.5) was required. This report appears to be first describing the use of a SPCE to detect DNA damage which can be related to the dose of gamma-radiation used.     

Biophysical characterization of an integrin-targeted lipopolyplex gene delivery vector

Nonviral gene delivery vectors now show good therapeutic potential: however, detailed characterization of the composition and macromolecular organization of such particles remains a challenge. This paper describes experiments to elucidate the structure of a ternary, targeted, lipopolyplex synthetic vector, the LID complex. This consists of a lipid component, Lipofectin (L) (1:1 DOTMA:DOPE), plasmid DNA (D), and a dual-function, cationic peptide component (I) containing DNA condensation and integrin-targeting sequences. Fluorophore-labeled lipid, peptide, and DNA components were used to formulate the vector, and the stoichiometry of the particles was established by fluorescence correlation spectroscopy (FCS). The size of the complex was measured by FCS, and the sizes of LID, L, LD, and ID complexes were measured by dynamic light scattering (DLS). Fluorescence quenching experiments and freeze-fracture electron microscopy were then used to demonstrate the arrangement of the lipid, peptide, and DNA components within the complex. These experiments showed that the cationic portion of the peptide, I, interacts with the plasmid DNA, resulting in a tightly condensed DNA-peptide inner core; this is surrounded by a disordered lipid layer, from which the integrin-targeting sequence of the peptide partially protrudes.     

Targeting lipopolyplexes using bifunctional peptides incorporating hydrophobic spacer amino acids: synthesis, transfection, and biophysical studies

We have developed efficient synthetic routes to two hydrophobic amino acids, suitably protected for solid-phase peptide synthesis, and have successfully synthesized peptides containing these or other hydrophobic amino acids as spacers between a Lys16 moiety and an integrin-targeting motif. These peptides have in turn been used to formulate a range of lipopolyplex vectors with Lipofectin and plasmid DNA. The transfection efficiencies of these vectors and their aggregation behavior in buffers and in serum have been studied. We have shown that vectors containing peptides incorporating long linkers that are entirely hydrophobic are less efficient transfection agents. However, linkers of equivalent length that are in part hydrophobic show improved transfection properties, which is probably due to the improved accessibility of the integrin-binding motif.