Chip calorimetry and biomagnetic separation: Fast detection of bacterial contamination at low cell titers

We present a new chip calorimeter for fast and quantitative measurement of metabolic heat rates of microorganisms attached to magnetic beads. In biomagnetic separation (BMS) experiments, Escherichia coli K12 immobilized on nonspecifically functionalized beads has a specific heat rate of around 1 pW per cell at 37°C. Therefore, at least 2 × 10⁴ bacteria are required to exceed the calorimetric signal resolution of 20 nW. If the samples to be analyzed have the original volume of 4 mL, bacteria at less than 10⁴ cells mL^?1 should be detectable. In practice, we achieved the detection of approximately 2 × 10⁴ cells mL^?1. The method presented here might also find some applications in the investigation of biofilms and study of biomolecular interactions.     

Condition and feeding behaviour of subadult burbot (<Emphasis Type="Italic">Lota lota</Emphasis>) in riverine and lacustrine environments

The condition and feeding behaviour of burbot, a widespread potamodromous species in riverine and lacustrine environments, were compared in order to evaluate the importance of both in three artificial systems. Subadult burbot were sampled in three temperate reservoirs in spring, and one of them also in summer and autumn. Standardised abundance and sizes of burbot were comparable between the reservoirs, but the conditions were significantly different. The Clark’s condition coefficient and index of fullness were independent of burbot size and individuals. Feeding behaviour in terms of abundance and composition of consumed food was environment-dependent. The most common prey category was aquatic insect larvae, dominating in the riverine environment. Permanent water invertebrates, microcrustaceans, beetles and crayfish were less common food and eaten more often in the lacustrine environment. Prey-fish were usually the most common species of suitable size. The food analyses demonstrated opportunistic feeding behaviour with selection of prey associated with benthic habitat and suitable size. Burbot is therefore flexible not only in environments utilisation, but also feeding strategy.     

Murine H-2Dd-reactive monoclonal antibodies recognize shared antigenic determinant(s) on human HLA-B7 or HLA-B27 molecules or both

We have evaluated the serological relationships between the murine H-2Dd and human HLA molecules using four H-2Dd-reactive monoclonal antibodies (mAbs) produced in the A.BY (KbIbDb) anti-A.TL (KsIkDd) combination. In the mouse, these reagents exhibited three distinct reactivity patterns: Dd, Ks, and H-2u (mAb 81.L); Dd, H-2p, and H-2u (mAb 81.R); and Dd, Kd, H-2p, H-2u, and H-2v (mAbs 97.G and 97.H). Sequential immunoprecipitation and cross-competitive mAb binding experiments revealed that these mAbs recognized determinants in two spatially distinct polymorphic domains on the H-2Dd molecule of B10.A(5R) cells (defined by mAbs 81.L and 81.R, 97.H, and 97.G, respectively). MAbs 81.R, 97.G, and 97.H, but not 81.L, also defined an HLA-linked polymorphism in the human, the main characteristics of which can be summarized as follows: (i) on B lymphoblastoid cell lines, mAbs 81.R and 97.H bound to cells expressing the HLA-B7, HL-B27 or Bw40 cross-reacting specificities, (ii) on peripheral blood lymphocyte (PBL) panel mAb 81.R exerted C dependent cytotoxicity to 118 of 400 cells tested, including almost all HLA-B7 or HLA-B27 cells or both (r: 0.952), (iii) the expression of the 81.R cross-reacting determinant segregated in an informative family with the parental haplotype carrying the HLA-B7 allele, and (iv) mAbs 81.R, 97.G, and 97.H recognized topologically related determinants on the same class I molecule(s) of the human B lymphoblastoid cells JY (HLA-A2,2, -B7,7). These data support the view that some, but not all H-2Dd allotopes have been conserved throughout evolution and are associated in the human with the HLA-B7, -B27 cross-reacting specificities.     

Tissue damage in organic rainbow trout muscle investigated by proteomics and bioinformatics

The response to tissue damage is a complex process, which involves the coordinated regulation of multiple proteins to ensure tissue repair. In order to investigate the effect of tissue damage in a lower vertebrate, samples were taken from rainbow trout (Oncorhynchus mykiss) at day 7 after damage and proteins were separated using 2DE. The experimental design included two groups of rainbow trout, which were fed organic feed either with or without astaxanthin. In total, 96 proteins were found to be affected by tissue damage, clearly demonstrating in this lower vertebrate the complexity and magnitude of the cellular response, in the context of a regenerative process. Using a bioinformatics approach, the main biological function of these proteins were assigned, showing the regulation of proteins involved in processes such as apoptosis, iron homeostasis, and regulation of muscular structure. Interestingly, it was established that exclusively within the astaxanthin feed group, three members of the annexin protein family (annexin IV, V, and VI) were regulated in response to tissue damage.     

Comparative morphology of surface ultrastructure of diphyllobothriidean tapeworms (Cestoda: Diphyllobothriidea)

Microtriches on the scolices and adjacent strobila of seven species of diphyllobothriidean cestodes (Bothridium pithonis, Cephalochlamys namaquensis, Dibothriocephalus latus, Duthiersia expansa, D. fimbriata, Ligula intestinalis, and Schistocephalus solidus) from different hosts (frogs, snakes, lizards, birds, and mammals) and biogeographic areas were examined using scanning and transmission electron microscopy. The basic structure of the tegument of the seven species studied does not differ markedly from that found in other cestodes. The main characteristic is the presence of electron‐dense bodies and vesicles in the distal cytoplasm. However, this study has shown differences in the morphology of microtriches even among species of the same family. Two different types of microtriches were found, filitriches and spinitriches, with the latter represented by two forms. Our study reveals that capilliform filitriches are most commonly found in Diphyllobothriidea. They were observed mainly on the strobila and the scolices of all but one studied species; individuals of L. intestinalis bore only coniform spinitriches on their surface. The same type of microtriches was found on the cirrus in D. latus. Gladiate spinitriches covered the scolex in both species of Duthiersia, and gladiate spinitriches interspersed with capilliform filitriches were observed on the anterior part of the strobila in D. fimbriata and the posterior part of the scolex in B. pithonis. Individuals of C. namaquensis were covered only by small acicular filitriches. No obvious pattern in the type and distribution of microtriches was observed among species that belong to different families and parasitize distantly related definitive hosts.     

Genetic homology and crossing over in the X and Y chromosomes of mammals

Summary The X-Y crossover model described in this paper postulates that (1) the pairing observed between the X and the Y chromosome at zygotene is a consequence of genetic homology, (2) there is a single obligatory crossover between the X and Y pairing segments, and (3) the segment of the X which pairs with the Y is protected from subsequent X inactivation. Genes distal to the proposed crossover (pseudoautosomal genes) will appear to be autosomally inherited because they will be transmitted to both male and female offspring. Some criteria for identifying pseudoautosomal genes are outlined.The existence of a single obligatory crossover between the X and Y of the mouse is strongly supported by a recent demonstration that the sex-reversing mutation Sxr, which is passed equally to XX and XY offspring by male carriers, is transmitted on the sex chromosomes. Pseudoautosomally inherited genes may also be responsible for XX sex reversal in goats and familial XX sex reversal in man.     

The effects of NMDA subunit composition on calcium influx and spike timing-dependent plasticity in striatal medium spiny neurons

Calcium through NMDA receptors (NMDARs) is necessary for the long-term potentiation (LTP) of synaptic strength; however, NMDARs differ in several properties that can influence the amount of calcium influx into the spine. These properties, such as sensitivity to magnesium block and conductance decay kinetics, change the receptor's response to spike timing dependent plasticity (STDP) protocols, and thereby shape synaptic integration and information processing. This study investigates the role of GluN2 subunit differences on spine calcium concentration during several STDP protocols in a model of a striatal medium spiny projection neuron (MSPN). The multi-compartment, multi-channel model exhibits firing frequency, spike width, and latency to first spike similar to current clamp data from mouse dorsal striatum MSPN. We find that NMDAR-mediated calcium is dependent on GluN2 subunit type, action potential timing, duration of somatic depolarization, and number of action potentials. Furthermore, the model demonstrates that in MSPNs, GluN2A and GluN2B control which STDP intervals allow for substantial calcium elevation in spines. The model predicts that blocking GluN2B subunits would modulate the range of intervals that cause long term potentiation. We confirmed this prediction experimentally, demonstrating that blocking GluN2B in the striatum, narrows the range of STDP intervals that cause long term potentiation. This ability of the GluN2 subunit to modulate the shape of the STDP curve could underlie the role that GluN2 subunits play in learning and development.     

Consciousness is Cheap, Even if Symbols are Expensive; Metabolism and the Brain’s Dark Energy

Use of symbols, the key to the biosemiotics field as to many others, required bigger brains which implied a promissory note for greater energy consumption; symbols are obviously expensive. A score years before the current estimate of 18–20% for the human brain’s metabolic demand on the organism, it was known that neural tissue is metabolically dear. This paper first discusses two evolutionary responses to this demand, on both of which there is some consensus. The first, assigning care of altricial infants with burgeoning brains (and in human infants the metabolic demand peaks at 65% of the total) to “allomothers” is not unique to humans. The second, using relatively small neurons as primates do, risks misfires past a certain minimal value. Moreover, in apparent paradox, there is an increasing consensus that large “Von Economo” neurons are critical for communication. This paper’s main contribution is the discussion of two further evolutionary tricks. The first is the use of self-similarity in the cortex, both in structure and process, to allow the cortex readily—and in energetic terms, parsimoniously—to shift between states in a high-dimensional space. This leads to discussion of the kind of formalism appropriate to model these shifts, a formalism which—it is tentatively suggested—may do double duty for the modeling of symbolic thought. The second trick is the superimposition on the background “white noise” of neural firing of EEG-detected waves like gamma. The paper describes a method, using the Hilbert transform, of calculating the dips in energy consumption as the brain is transitioned by gamma waves. It is hypothesized that consciousness may be a spandrel, the incidental result of a neurodynamic imperative that the brain enter a maximally sensitive (in sensory terms) “zero power” state a few times a second. If that is the case, then there are obvious benefits for health in meditation, which can be viewed as a state of consciousness extended over time by limiting afferent stimuli.     

Structural determination of the O-specific polysaccharide from Aeromonas hydrophila strain A19 (serogroup O:14) with S-layer

Bacteria belonging to the genus Aeromonas are Gram-negative mesophilic and essentially ubiquitous in the microbial biosphere; moreover they are considered very important pathogens in fish and responsible for a great variety of human infections.The virulence of Gram-negative bacteria is often associated with the structure of lipopolysaccharides, which consist of three regions covalently linked: the glycolipid (lipid A), the oligosaccharide region (core region) and the O-specific polysaccharide (O-chain, O-antigen).The O-chain region seems to play an important role in host-pathogen interaction. In the case of Aeromonas hydrophila the majority of pathogenic strains belongs to serogroups O:11, O:16, O:18 and O:34. In this paper, we report the complete structure of the O-chain of A. hydrophila strain A19 (serogroup O:14), a pathogenic strain isolated from European eels, which showed high virulence when tested in trout or mice. Dried cells were extracted by the PCP (phenol/chloroform/petroleum ether) method obtaining the lipopolysaccharide. After mild acid hydrolysis the lipid A was removed by centrifugation and the obtained polysaccharide was fully characterized by means of chemical analysis and one- and two-dimensional NMR spectroscopy. All the data collected are directed towards the following structure:     

The role of the S-S bridge in retroviral protease function and virion maturation

Retroviral proteases are translated as a part of Gag-related polyproteins, and are released and activated during particle release. Mason-Pfizer monkey virus (M-PMV) Gag polyproteins assemble into immature capsids within the cytoplasm of the host cells; however, their processing occurs only after transport to the plasma membrane and subsequent release. Thus, the activity of M-PMV protease is expected to be highly regulated during the replication cycle. It has been proposed that reversible oxidation of protease cysteine residues might be responsible for such regulation. We show that cysteine residues in M-PMV protease can form an intramolecular S-S bridge. The disulfide bridge shifts the monomer/dimer equilibrium in favor of the dimer, and increases the proteolytic activity significantly. To investigate the role of this disulfide bridge in virus maturation and replication, we engineered an M-PMV clone in which both protease cysteine residues were replaced by alanine (M-PMV(PRC7A/C106A)). Surprisingly, the cysteine residues were dispensable for Gag polyprotein processing within the virus, indicating that even low levels of protease activity are sufficient for polyprotein processing during maturation. However, the long-term infectivity of M-PMV(PRC7A/C106A) was noticeably compromised. These results show clearly that the proposed redox mechanism does not rely solely on the formation of the stabilizing S-S bridge in the protease. Thus, in addition to the protease disulfide bridge, reversible oxidation of cysteine and/or methionine residues in other domains of the Gag polyprotein or in related cellular proteins must be involved in the regulation of maturation.