Toxins from anaerobic bacteria: specificity and molecular mechanisms of action

Major advances have been made in the past five years in the identification of cellular targets of toxins produced by anaerobic bacteria. These targets include the vesicular membrane docking and fusion apparatus, the actin cytoskeleton, the signal transduction machinery and the cell membrane. The recent discovery that large clostridial toxins (Clostridium difficile A and B toxins, C. sordellii lethal and hemorrhagic toxins, and alpha C. novyi toxin) are monoglucosyltransferases, together with the establishment of the perfringolysin crystal structure, has led to new insights in the field of toxins from anaerobic bacteria.     

Kinetics of the hydrolysis of lean meat protein by alcalase: Derivation of two alternative rate equations and their fit to experimental data

Two rate equations have been developed to model the hydrolysis of ground lean meat protein by Alcalase. The first equation was based on classical Michaelis-Menten kinetics and the second on the adsorption of enzyme to the protein prior to reaction. It was assumed that this adsorption could be modelled by a Langmuir-type adsorption isotherm. Each equation considered the enzyme to be competitively inhibited by reaction product, and considered enzyme inactivation to be first order. Both rate equations have been fitted to experimental data obtained from the hydrolysis of meat protein by Alcalase. Initial rate data indicated that the adsorption model was more appropriate. However, both equations gave satisfactory fits to 11 reaction progress curves determined over a wide range of enzyme and substrate concentrations.     

Steroid hormones and agonistic behavior in a cichlid teleost, Aequidens pulcher

The effects of three steroid hormones on the agonistic behavior of female Aequidens pulcher have been evaluated. Testosterone, estradiol, and cortisol were tested using an immersion technique to minimize trauma, and we also examined metyrapone, a blocker of cortisol biosynthesis. Two different experimental protocols were employed, the first investigating agonistic interactions within groups of fish, and the second examining the responses of isolated fish to models and mirrors. Differences between replicates were small, and both protocols supported similar conclusions. Each of the three hormones produced a characteristically different spectrum of behaviors when compared to the controls. Testosterone increased agonistic behavior in all experimental situations, while estradiol had a generally opposite effect; this may reflect the natural modulation of behavior by hormones during the reproductive cycle of A. pulcher. Cortisol also had distinct behavioral effects; available evidence suggests that this steroid increases submissive components of agonistic behavior, and that observed increases in some aggressive components are an indirect consequence, dependent upon the feedback of social information received by each fish. Metyrapone treatment greatly reduced all agonistic behaviors, groups of fish forming shoals typical of juveniles. This was not reversed by replacement therapy with cortisol, which suggests that metyrapone affects behavior by an alternative, possibly toxic, mechanism.     

Bill morphology reflects adaptation to a fibrous diet in the kākāpō (Strigops: Psittaciformes)

We describe the upper portion of the bill sheath (rhinotheca) of the kākāpō (Strigops habroptilus) from three adult female specimens. The external buccal surface of the rhinotheca is deeply concave with a prominent palatal stop and hardened chevrons creating a ‘milling apparatus’ that the kākāpō uses to grind food. The palatal stop presents a working face of 40–50?mm². The internal surface of the rhinotheca mirrors the overlying premaxilla and provides a distinct thickened abutment consistent with resistance against the increased workload of the mandibles (gnathotheca) due to the kākāpō’s fibrous diet and chewing style. Along the midline, the rhinotheca at the abutment is up to 5.6?mm thick, compared with as thin as 2.1?mm elsewhere on the midline. The closely related Nestor parrots have less developed palatal stops, chevrons and abutments on their rhinothecas consistent with their lower preference for fibrous plant material. The form of the rhinotheca agrees with the kākāpō’s feeding ecology as a generalist herbivore that grinds locally available fibrous material to assist digestion.     

Molecular evolution modeled as a fractal Poisson process in agreement with mammalian sequence comparisons

The fractal doubly stochastic Poisson process (FDSPP) model of molecular evolution, like other doubly stochastic Poisson models, agrees with the high estimates for the index of dispersion found from sequence comparisons. Unlike certain previous models, the FDSPP also predicts a positive geometric correlation between the index of dispersion and the mean number of substitutions. Such a relationship is statistically proven herein using comparisons between 49 mammalian genes. There is no characteristic rate associated with molecular evolution according to this model, but there is a scaling relationship in rates according to a fractal dimension of evolution. The FDSPP is a suitable replacement for the homogeneous Poisson process in tests of the lineage dependence of rates and in estimating confidence intervals for divergence times. As opposed to other fractal models, this model can be interpreted in terms of Darwinian selection and drift.      

A Retrospective Analysis of the Establishment and Dispersal of the Introduced Australian Parasitoids Xanthopimpla rhopaloceros (Krieger) (Hymenoptera: Ichneumonidae) and Trigonospila brevifacies (Hardy) (Diptera: Tachinidae) within New Zealand

Two Australian parasitoids, Xanthopimpla rhopaloceros (Krieger) and Trigonospila brevifacies (Hardy), were introduced to New Zealand to control the light-brown apple moth (Epiphyas postvittana Walker). Dispersal by the parasitoids has since occurred naturally and with the aid of releases in fruit-growing areas. The present geographical range of the parasitoids includes all the North Island and some offshore islands to latitude 41 20 S. X. rhopaloceros is also present to latitude 41 48 S in the South Island. Comparisons of these distributions with those in Australia indicate that climatic conditions may have played a major role in the areas of establishment of both species in New Zealand. The mean winter temperature may be a limiting factor in the dispersal of T. brevifacies and X. rhopaloceros in New Zealand. Other factors that have probably aided the successful dispersal of the parasitoids include the wide distribution of host Tortricidae and the occurrence of tortricid host plants. The areas of New Zealand that appear suitable for further colonization by T. brevifacies include northern areas of the South Island, and both parasitoids could disperse further into suitable climate areas of the east and west coasts of the central South Island. The rate of dispersal for X. rhopaloceros was estimated at 13-24 km/year, and for T. brevifacies at 8-15 km/year.     

Molecular evidence for the presence of novel actinomycete lineages in a temperate forest soil

PCR primers were designed to selectively recover partial (1100 bp) actinomycete 16S ribosomal DNA sequences from a temperate forest soil. A gene library was made and colony PCR was used to identify clones containing inserts. Unique clones were identified and partial or complete insert sequences were determined for 53 clones. Phylogenetic analyses revealed that 46 (87%) of the clones sampled contained 16S rDNA sequences which fell within the actinomycete radiation. The largest group of 34 sequences formed two closely related monophyletic groups in the 16S rRNA tree, which in turn formed a weakly supported sister group with the sequence fromActinomadura madurae. Four novel 16S rDNA lineages were detected inMycobacterium, one inPropionibacterium and one inCorynebacterium. Three novel sequences weakly grouped withSporichthya polymorpha. Two sequences formed an isolated lineage not closely related to any of the reference actinomycetes. Our results lend strong support to the hypothesis that cultured (and sequenced) actinomycetes do not adequately describe the diversity of this group in the environment.     

Sucrose-derived exopolymers have site-dependent roles in Streptococcus mutans-promoted dental decay

Abstract We have constructed a panel of mutants of S. mutans V403 which are defective in one or more of the glucosyltransferase genes ( gtfB,C or D ) or the fructosyltransferase gene ( ftf ). These strains have been tested for virulence in a gnotobiotic rat caries model with reference to both buccal (smooth surface) and sulcal (pit and fissure) carious lesions. Our data suggest differing roles for degradable and non-degradable polymers at buccal and sulcal sites. Non-degradable polymers (made by products of the gtfB and C genes) contributed significantly to the severity of smooth surface lesions. However, our studies suggested their role in pit and fissure lesions was not as important as the role of degradable polymers. Specifically, a mutant deficient in the major insoluble glucan synthesizing activity (product of the gtfB gene) was 25% more cariogenic on sulcal surfaces than was the wild-type V403 strain. We propose that extracellular glucosyltransferases and fructosyltransferase compete for sucrose and that this competition influences pathogenicity at differing tooth sites.     

Infectivity of a glucan synthesis-defective mutant of Streptococcus gordonii (Challis) in a rat endocarditis model

Abstract Streptococcus gordonii , a member of the human indigenous oral microflora, colonizes smooth tooth surfaces and contributes to dental plaque formation. Although it is not recognized as being a cariogenic pathogen, it may cause endocarditis following invasion of the bloodstream. Using allelic exchange mutagenesis, we have constructed a mutant of S. gordonii (Challis) which is defective in its single functional glucosyltransferase gene and, hence, is unable to synthesize glucan exopolymers from sucrose. When examined in a rat endocarditis model, the sucrose-grown mutant did not differ significantly from S. gordonii wild-type, suggesting that glucan polymers did not contribute to infectivity. This result was in striking contrast to that previously observed with a polymer-defective S. mutans mutant.     

Prenatal and postnatal changes in the content and species of ferritin in rat liver

The iron and ferritin content of rat liver and the species of ferritin present were examined from 4 days before to 3 weeks after birth. 1. Total iron and ferritin iron accumulated rapidly during the last days of gestation and from the second postnatal day underwent a steady depletion. 2. The amount of iron deposited before birth in the liver of each pup varied inversely with litter size and could be increased moderately by injection of iron into the mother before mating. 3. Intraperitoneal injection of iron 1 day after birth doubled the concentration of total iron, ferritin iron and ferritin protein in the liver over the next 24h, but at 3 weeks after birth it raised the very low concentrations of iron and ferritin severalfold. 4. As shown by electrophoretic migration, ferritin and dissociated ferritin subunits prepared from the livers of rats from 4 days before to 3 weeks after birth differed from those of adult liver ferritin and were indistinguishable from those of adult kidney and spleen ferritin. Treatment with iron at 3 weeks of age induced formation of a ferritin with electrophoretic properties resembling those of adult liver. It is concluded that iron given at this stage of development may activate the genetic cistron for adult liver ferritin.