Evolutionary analysis of the picornavirus family

An exhaustive evolutionary analysis of the picornavirus family has been carried out using the amino acid sequences of several proteins of the viruses including: the capsid proteins (1D, 1B, and 1C) situated at the 5 end of the genome and responsible for the serotype of the viruses, and the viral polymerase (3D), located at the 3 end of the genome. The evolutionary relationships found among the viruses studied support the new classification, recently suggested, in contrast to the classical one, and the existence of a new genus for the picornavirus family. In the new taxonomic organization, five genera form the picornavirus family: (1) aphthoviruses, (2) cardioviruses, (3) hepatoviruses (previously classified as enteroviruses), (4) renteroviruses (which mainly constitute a combination of the previous genera rhinovirus and enterovirus), and (5) a new genus, with a new and unique representative: the echovirus 22. Our analysis also allowed us, for the first time, to propose the most probable sequence of speciation events to have given rise to the current picornavirus family.The bootstrap procedure was used to check the reliability of the phylogenetic trees obtained. The application of the method of the statistical geometry in distance space to internal branches of the tree revealed a high degree of evolutionary noise, which makes the resolution of some internal branching points difficult. Correspondence to: J. Dopazo     

Microfiltration of yeast suspensions with self-cleaning spiral vortices: possibilities for a new membrane module design

A novel method of producing controlled vortices was used to reduce both concentration polarization and membrane fouling during microfiltration of Saccharomyces cerevisiae broth suspensions. The method involves flow around a curved channel at a sufficient rate so as to produce centrifugal instabilities (called Dean vortices). These vortices depolarize the build-up of suspended particles such as yeast cells at the membrane-solution interface and allow for increased membrane permeation rates. Various operating conditions under which such vortices effectively reduced cake build-up of suspended particles such as yeast cells at the membrane-solution interface and allow for increased membrane permeation rates. Various operating conditions under which such vortices effectively reduced cake build-up during microfiltration of 0 to 0.55 dry wt% yeast broth were investigated. Flux improvements of over 60% for 0.25 dry wt% yeast broth for flow with over that without Dean vortices were observed. This beneficial effect increased with increasing retentate flow rate and increasing transmembrane pressure and decreased with increasing concentration of suspended matter. Similar behavior was observed whether the cells were viable of killed. the improvement in flux in the presence over that in the absence of vortices correlated well with centrifugal force or azimuthal velocity squared. The relative cake resistances increased with reservoir yeast concentration. These values with vortices increased from 62% to 75% of that without vortices with increasing yeast concentration. The ratio of the cake thicknesses in the limiting case (at high feed concentration) was 3.25. These results suggest that self-cleaning spiral vortices could be effective in maintaining good and steady microfiltration performance with cell suspensions other than those tested. (c) 1995 John Wiley & Sons, Inc.     

Microquantification of proteins with low chromophore content

The advantages of the intrinsic fluorescence of the tryptophan and the absorbance of the methionine residues of the 18 kDa-hsp - a recombinant protein from Mycobacterium leprae - was exploited here to develop a sensitive and low costs method for protein assaying. They presented linearity between 3 and 1000 g of protein. The correlations between intrinsic fluorescence or absorbance at 230 nm and protein contents were both superiors to 0.99. These methods can be extended to others proteins with low aromatic residue contents.     

Distribution of transposable elements in neotropical species of Drosophila

The phylogenetic distribution of transposable families, P, gypsy, hobo, I, and mariner has been analyzed in 33 species of 11 groups of neotropical Drosophila and a Drosophilidae species Zygotrica vittimaculosa, using squash blot and dot blot. Genomic DNA of almost all neotropical species tested hybridized with gypsy probe and some species showed a particularly strong hybridization signal, as D. gaucha, D. virilis, and species of flavopilosa group. The hobo element was restricted to melanogaster group and some strains of D. willistoni. Only D. simulans DNA showed hybridization to mariner probe in all species tested and D. simulans and D. melanogaster showed hybridization with I element probe. P element homologous sequence was present in D. melanogaster and all species and strains of the willistoni and saltans groups tested. The presence of at least one P-homologous sequence was detected in Drosophila mediopunctata. This one was the only P-bearing species of all six tested from the tripunctata group. Four different pairs of primers homologous to segments of the canonical sequence of D. melanogaster's P were used to amplify specific sequences from D. mediopunctata DNA, showing the occurrence of seemingly well-conserved P-homologous sequences. This revised version was published online in July 2006 with corrections to the Cover Date.     

Characterization of novel long-chain 1,2-diols in Thermus species and demonstration that Thermus strains contain both glycerol-linked and diol-linked glycolipids.

In this study, we purified and characterized tetra- and triglycosyl glycolipids (GL-1 and GL-2, respectively) from two different colonial forms of Thermus scotoductus X-1, from T. filiformis Tok4 A2, and from T. oshimai SPS-11. Acid hydrolysis of the purified glycolipids liberated, in addition to the expected long-chain fatty acids, two components which were identified by gas chromatography-mass spectrometry as 16-methylheptadecane-1,2-diol and 15-methylheptadecane-1,2-diol. Fast atom bombardment mass spectrometry of the intact glycolipids indicated that a major proportion consisted of components with glycan head groups linked to long-chain 1,2-diols rather than to glycerol, although in all cases glycerol-linked compounds containing similar glycan head groups were also present. As in other Thermus strains, the polar head group of GL-1 from T. filiformis Tok4 A2 and from T. scotoductus X-1 colony type t2 was a glucosylgalactosyl-(N-acyl)glucosaminylglucosyl moiety. However, GL-2 from T. scotoductus X-1 colony type t1 and from T. oshimai SPS-11 was a truncated analog which lacked the nonreducing terminal glucose. Long-chain 1,2-diols have been previously reported in the polar lipids of Thermomicrobium roseum and (possibly) Chloroflexus aurantiacus, but to our knowledge, this is the first report of their detection in other bacteria and the first account of the structural determination of long-chain diol-linked glycolipids.     

Latent Transforming Growth Factor-β Binding Protein Domains Involved in Activation and Transglutaminase-dependent Cross-Linking of Latent Transforming Growth Factor-β

Transforming growth factor-β (TGF-β) is secreted by many cell types as part of a large latent complex composed of three subunits: TGF-β, the TGF-β propeptide, and the latent TGF-β binding protein (LTBP). To interact with its cell surface receptors, TGF-β must be released from the latent complex by disrupting noncovalent interactions between mature TGF-β and its propeptide. Previously, we identified LTBP-1 and transglutaminase, a cross-linking enzyme, as reactants involved in the formation of TGF-β. In this study, we demonstrate that LTBP-1 and large latent complex are substrates for transglutaminase. Furthermore, we show that the covalent association between LTBP-1 and the extracellular matrix is transglutaminase dependent, as little LTBP-1 is recovered from matrix digests prepared from cultures treated with transglutaminase inhibitors. Three polyclonal antisera to glutathione S–transferase fusion proteins containing amino, middle, or carboxyl regions of LTBP-1S were used to identify domains of LTBP-1 involved in crosslinking and formation of TGF-β by transglutaminase. Antibodies to the amino and carboxyl regions of LTBP-1S abrogate TGF-β generation by vascular cell cocultures or macrophages. However, only antibodies to the amino-terminal region of LTBP-1 block transglutaminase-dependent cross-linking of large latent complex or LTBP-1. To further identify transglutaminase-reactive domains within the amino-terminal region of LTBP-1S, mutants of LTBP-1S with deletions of either the amino-terminal 293 (ΔN293) or 441 (ΔN441) amino acids were expressed transiently in CHO cells. Analysis of the LTBP-1S content in matrices of transfected CHO cultures revealed that ΔN293 LTBP-1S was matrix associated via a transglutaminasedependent reaction, whereas ΔN441 LTBP-1S was not. This suggests that residues 294–441 are critical to the transglutaminase reactivity of LTBP-1S.     

Hemagglutination properties of Enterococcus

In total, 86 enterococcal strains including representatives of most of the described species were tested for the ability to agglutinate human, sheep, and rabbit erythrocytes. Five strains did not react with any of the erythrocytes tested, and 81 (94.2%) strains agglutinated only rabbit erythrocytes. The hemagglutination titers ranged from 2 to 64. Loss of the hemagglutination activity was observed when rabbit erythrocytes were treated with trypsin or neuraminidase. Trypsin treatment of the bacterial suspensions also caused loss of the agglutination ability. On the other hand, heat treatment of bacterial suspensions increased the efficiency of the interactions, and higher titers were obtained. Assays for inhibition of hemagglutination were performed with -d-fucose, -d-galactose, -d-galactose, d-glucose, N-acetyl-galactosamine, N-acetyl-glucosamine, N-acetylneuraminic acid, N-acetylneuraminic acid-lactose, and fetuin. Only fetuin was able to inhibit the hemagglutination reactions. The results showed that hemagglutination properties are common to the different enterococcal species tested. They also suggest that enterococci possess hemagglutinins of proteic and non-proteic nature that are involved in the attachment to sialic acid-containing receptors on the surface of rabbit erythrocytes.     

Catecholamine and MHPG plasma levels,platelet MAO activity,and3H-imipramine binding in heroin and cocaine addicts

This work evaluated in a population of heroin and heroin plus cocaine human addicts:

1. Norepinephrine (NE), epinephrine (Epi), and 3-methoxy-4-hydroxyphenylglycol (MHPG) (the principal metabolite of brain NE) plasma levels;
2. Monoamine oxidase (MAO) activity; and
3. ³H-imipramine specific binding to the amine carrier in platelets.

NE plasma levels were significantly lower in the short-term heroin user groups (1–3 and 4–6 yr), a finding not observed in both the long-term heroin user (>6 yr) and heroin plus cocaine user (>6 yr) groups. Epi levels changed in a similar manner, except that a significant increase was noted in heroin plus cocaine abusers. Conversely, dopamine and MHPG plasma levels increased with the duration of heroin use, and even more with cocaine abuse. Platelet MAO activity increased in all groups. Specific³H-imipramine binding sites showed an increase after 3 yr of heroin abuse and in all heroin plus cocaine addicts. In conclusion, short-term use of heroin decreases NE or Epi release, but with prolonged use, a slow adpatation occurs. In contrast, cocaine inhibits the neuronal Epi uptake, even in a situation of long duration of abuse. Probably the amine levels additionally regulate the amine carrier, resulting in changes that show a different pattern from major depression. These drugs of abuse may also influence directly or indirectly related enzymatic systems.     

Synthesis of new azino fused benzimidazolium salts. a new family of DNA intercalating agents. I

A series of new pyrido[1,2-a]- and pyridazino[1,6-a]benzimidazolium salts have been synthesized from readily available 1,3-disubstituted 2-alkylbenzimidazolium salts. Their affinity to DNA and in vitro cytotoxicity versus HT-29 have been tested. The initial results show that the title compounds are a new family of intercalating agents.     

Exudate-feeding byCallithrix jacchus penicillata in semideciduous woodland (Cerradão) in central Brazil

We report the exudate feeding behavior of two groups of marmosets (Callithrix jacchus penicillata) living permanently in Cerradão, a common woodland formation of Central Brazil. Cerradão is an open canopy formation and marmosets must occasionally descend to the ground in order to move from tree to tree. Even in atypical habitat, exudate eating is the predominant foraging activity. Marmosets are engaged in exudate collection over 70% of the total time spent feeding. They were observed gnawing on seven species of trees, and consumed exudates from four of these species. We compared the degree of utilization of the exudate sources, and examined a number of different characteristics of the exudates. Morphological adaptations that allow for the exploitation of the “exudate-eater niche” may be an important component of the adaptability ofCallithrix marmosets.