Hydrodynamic, structural and magnetic properties of Megasphaera elsdenii Fe hydrogenase reinvestigated

Megasphaera elsdenii hydrogenase has been purified to homogeneity using an FPLC procedure as the final step. The protein gives a single band in SDS/PAGE with an apparent molecular mass of 57-59 kDa. There is no second hydrogenase activity in the soluble fraction of M. elsdenii. The hydrodynamics of the enzyme have been compared to those of the two-subunit Fe hydrogenase from Desulfovibrio vulgaris (Hildenborough) in the analytical ultracentrifuge using the absorption of the intrinsic iron-sulfur clusters as the monitor. Sedimentation-velocity experiments indicate the M. elsdenii enzyme (s20,w = 4.95 S) to be essentially globular, while the D. vulgaris enzyme (s20,w = 4.1 S) has a less symmetric shape. From the sedimentation equilibrium measurements under a variety of conditions an average molecular mass is calculated of 58 kDa (M. elsdenii) and 54 kDa (D. vulgaris), respectively. Pure, maximally active M. elsdenii hydrogenase has A405/A280 = 0.36 and has a specific H2-production activity of 400 mumol H2.min-1.(mg protein)-1 at 30 degrees C and pH 8.0. The enzyme contains some 13-18 iron and acid-labile sulfur ions/58-kDa monomer. Eight of these Fe-S are present as two electron-transferring ferredoxin-like cubanes with Em approximately greater than -0.3 V, as indicated by pH-dependent EPR spectroscopy on the H2-reduced enzyme. In the (re)oxidized state the remainder iron gives rise to a single S = 1/2 rhombic EPR signal. Hydrogen-production activity, content of remainder iron and rhombic EPR signal intensity are mutually correlated. Purified hydrogenase appears to exist as a mixture of fully active holoenzyme and inactive protein still carrying the two cubanes but deficient in active-site iron.     

Advances in transgenic rat production

Predictable and reproducible production of transgenic rats from a standardized input of egg donors and egg recipients is essential for routine rat model production. In the course of establishing a transgenic rat service, transgenic founders were produced from three transgenes in outbred Sprague-Dawley (SD) rats and four transgenes in inbred Fischer 344 (F344) rats. Key parameters that affect transgenesis efficiency were assessed, including superovulation treatments, methods to prepare pseudopregnant recipients, and microinjection technique. Five superovulation regimens were compared and treatment with 20 IU PMSG and 30 IU HCG was selected for routine use. Four methods to prepare pseudopregnant egg recipients were compared and estrus synchronization with LHRHa and mating to vasectomized males was selected as most effective. More than 80% of eggs survived microinjection when modified pronuclear microinjection needles and DNA buffers were used. The efficiencies of transgenic production in rats and C57BL/6J (B6J) mice were compared to provide a context for assessing the difficulty of transgenic rat production. Compared to B6J mice, SD rat transgenesis required fewer egg donors per founder, fewer pseudopregnant egg recipients per founder, and produced more founders per eggs microinjected. Similar numbers of injection days were required to produce founders. These results suggest that SD rat transgenesis can be more efficient than B6J mouse transgenesis with the appropriate technical refinements. Advances in transgenic rat production have the potential to increase access to rat models.     

Zinc induces caspase-dependent mitochondrial pathway of the programmed cell death in haemocytes of Drosophila melanogaster

Zinc is an essential trace element in cells. However, its high level in cytoplasm promotes activation of stress signaling pathways and may lead to cell death. In the present study we used Drosophila melanogaster blood cells (haemocytes), obtained from the third instar larvae, to study the effects of high concentrations of Zn(2+) on programmed cell death (PCD). We analyzed the activity of caspases, the level of caspase inhibitor protein DIAP1 and metallothioneins, as well as calcium concentrations and activity of mitochondria in haemocytes exposed to 0.35 and 1.7 mM concentrations of Zn. The obtained results showed that rapid increase of [Zn(2+)]( i ) in the cytoplasm up-regulates metallothionein MtnB but not MtnA gene expression in cells treated with Zn(2+) in both concentrations. Excess of Zn(2+) also induced activation of the initiator caspase Dronc, associated with the mitochondrial pathway of PCD, and the effector caspase DrICE. In turn, the activity of receptor-regulated Dredd caspase was not changed. The level of DIAP1 decreased significantly in haemocytes in the presence of high Zn(2+) concentration in comparison to untreated cells. Moreover, mitochondrial membrane potential was significantly decreased after exposure to Zn ions. These results indicate that high concentration of Zn(2+) in the cytoplasm of haemocytes induces PCD via a mitochondrial pathway and that caspases play a pivotal role in this process.     

Caution on the assessment of intestinal parasitic load in studying parasite-mediated sexual selection: The case of Blackbirds coccidiosis

The parasite-mediated sexual selection (PMSS) theory has led to an increasing number of experimental studies essentially focussed on blood parasites. Currently, more research is being carried out on intestinal parasites in relationship to this theory. Before testing the theory with gastrointestinal parasites, it is important: (i) to determine an optimal research methodology to obtain an accurate assessment of parasite burden and (ii) to have information about life-history traits of the parasite to interpret data appropriately. In this study, we present data on oocyst output of Isosporaturdi in the faeces of blackbirds (Turdus merula) that illustrate the importance of developing methods that are relevant for a particular model system, instead of relying on existing methods that may work in other systems. Our results show that: (i) a single droplet of faeces will accurately indicate the parasitic load in blackbirds, (ii) oocyst shedding varies greatly within and between days, (iii) the course of infection is characterised with two successive peaks of oocyst shedding, (iv) infection lasts approximately 1 month and (iv) there is no effect of sex, size of infective dose or re-infection on the course of infection. We discuss the practical implications of these results in determining the reliability of, and in avoiding erroneous conclusions about, PMSS when using intestinal parasites as models. In particular, we emphasise that numerous measurements must be performed on different days. In addition, faeces must be collected at a particular time of day because there is a strong temporal variation in oocyst shedding. Finally, a standardised methodology that has been developed for a particular host-parasite assemblage does not necessarily work with other biological models. Together, our results should encourage future investigators to identify an accurate methodology for assessing parasitic load as a first step before testing hypotheses associated with the PMSS theory.     

Nutritional dynamics during the development of xylophagous beetles related to changes in the stoichiometry of 11 elements

The present study examines the adaptive strategy used by wood‐boring beetles to compensate for the lack of nutrients in dead wood. The contents of nutritional elements in growing wood‐boring beetles (Stictoleptura rubra L. and Chalcophora mariana Dejean) are compared with the elemental composition of decaying dead wood (pine stumps), showing changes during the beetles' ontogenetic (i.e. larval) development. The stoichiometric ratios of C and other nutritional elements (N, P, K, Na, Ca, Mg, Fe, Zn, Mn and Cu) are investigated to identify the most important nutrients for larval development. The degree of nutritional mismatch that is encountered by the beetle larvae changes dramatically over 3–4 years of simultaneous larval growth and wood decay. Excluding C, the relative contents of nutritional elements increase substantially in decaying wood, whereas the opposite tendency is found in larvae, most likely because of carbon deposition in fat. The elements limiting larval development because of their scarcity in dead wood are N, P, K, Na, Mg, Zn and Cu. Fungal activity (i.e. the transport of nutrients from the surrounding environment to decaying stumps) can explain the observed mitigation of the original mismatch, although prolongation of the larval development time is still necessary to compensate for the scarcity of some of the required elements in food.     

Palynostratigraphy of the Frasnian-Famennian boundary deposits from the Central Devonian Field, western Russia and comparisons with adjacent areas

Detailed palynological research carried out on samples from the three quarries at Khlevnoye and Kamenka (Voronezh region) permitted recognition of the Frasnian-Famennian boundary in the lower Kamenka quarry. Two miospore zones, Cristatisporites deliquescens-Verrucosisporites evlanensis (DE) from the Frasnian and Cyrtospora cristifera-Diaphanospora zadonica (CZ) from the Famennian, have been recognized. Some faunistic data were also obtained from the F-F boundary deposits. Research confirmed the existence of a sedimentological gap between the Livny and Zadonsk horizons, including all of the Volgograd horizon. These new palynological results are compared with data from adjacent areas: Pripyat Depression (Belarus), Timan-Pechora Province and the Volgograd Volga region (Russia), western Pomerania and the Holy Cross Mountains (Poland).     

Polymorphism of the serum proteins of wild pigs

One hundred and ninety five wild pigs from two different regions of Poland were investigated for transferrin, amylase and ceruloplasmin polymorphism. A new tranferrin phenotype Tf PB was detected. This phenotype differed from Tf AB in the electrophoretic mobility of the more anodal transferrin. Tf P is assumed to be the product of a new allele Tf P at the Tf locus. Two amylase phenotypes Am 1-2 and Am 2 were observed. The Am 1 allele was absent from the pigs in the Poznan region. Only one ceruloplasmin phenotype, Cp B, was found.     

A comprehensive phylogeographic study of Arion vulgaris Moquin-Tandon, 1855 (Gastropoda: Pulmonata: Arionidae) in Europe

Organisms Diversity & Evolution - Arion vulgaris Moquin-Tandon, 1855 is regarded as one of the 100 most invasive species in Europe. The native distribution range of this species is uncertain,...     

The combined impact of mechanical factors on the wall stress of the human ascending aorta – a finite elements study

Background

Biomechanical factors influence stress in the aortic wall. The aim of this study was to assess how the diameter and shape of the vessel, blood pressure and longitudinal systolic aortic stretching (SAS) caused by the contraction of the myocardium influence stress in the aortic wall.

Methods

Three computational models of the non-dilated aorta and aneurysms of the ascending aorta and aortic root were created. Then, finite elements analyses were carried out. The models were subjected to blood pressure (120 mmHg and 160 mmHg) and longitudinal systolic aortic stretching (0 mm, 5 mm, 10 mm and 15 mm). The influence of wall elasticity was examined too.

Results

Blood pressure had a smaller impact on the stress than the SAS. An increase in blood pressure from 120 mmHg to 160 mmHg increased the peak wall stress (PWS) on average by 0.1 MPa in all models. A 5 mm SAS caused a 0.1–0.2 MPa increase in PWS in all the models. The increase in PWS caused by a 10 mm and 15 mm SAS was 0.2 MPa and 0.4 MPa in the non-dilated aorta, 0.2–0.3 MPa and 0.3–0.5 MPa in the aneurysm of the ascending aorta, and 0.1–0.2 MPa and 0.2–0.3 MPa in the aortic root aneurysm model, respectively. The loss of elasticity of the aneurysmal wall resulted in an increase of PWS by 0.1–0.2 MPa.

Conclusions

Aortic geometry, wall stiffness, blood pressure and SAS have an impact on PWS. However, SAS had the biggest impact on wall stress. The results of this study may be useful in future patient-specific computational models used to assess the risk of aortic complications.