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The in vitro infection of the hematopoietic stroma of trout kidney by hemorrhagic septicemia rhabdovirus

  • Diago, M. L.
  • Estepa, A.
  • López-Fierro, P.
  • Villena, A.
  • Coll, J. M.
Viral hemorrhagic septicaemia virus (VHSV) infected the hematopoietic stromal cells (7,8) derived from pronephritic tissue of the rainbow trout, Oncorhynchuss mykiss, W.;at their ninth passage in vitro. Viral infection resulted in the development of lytic cytopathic effects on confluent in vitro tridimensional network stromal cell cultures. Replication of VHSV in the stromal cell cultures was demonstrated by the increase in infectivity by epithelioma papulosum cyprini (EPC) cell culture assays and by the increase of the nucleoprotein antigen of VHSV by ELISA. By using anti-VHSV monoclonal antibodies (MAbs), flow cytometry studies demonstrated that only the infected stromal cells contained cytoplasmic viral antigens. The lytic infection of trout hematopoietic stromal cells in vitro could be relevant to the hemorrhagic pathology seen in the kidney of fish infected with VHSV.

The gene encoding glutathione-dependent formaldehyde dehydrogenase/GSNO reductase is responsive to wounding, jasmonic acid and salicylic acid

  • Díaz, M.
  • Achkor, H.
  • Titarenko, E.
  • Martínez, M. C.
It has recently been discovered that glutathione-dependent formaldehyde dehydrogenase (FALDH) exhibits a strong S-nitrosoglutathione reductase activity. Plants use NO and S-nitrosothiols as signaling molecules to activate defense mechanisms. Therefore, it is interesting to investigate the regulation of FALDH by mechanical wounding and plant hormones involved in signal transduction. Our results show that the gene encoding FALDH in Arabidopsis (ADH2) is down-regulated by wounding and activated by salicylic acid (SA). In tobacco, FALDH levels and enzymatic activity decreased after jasmonate treatment, and increased in response to SA. This is the first time that regulation of FALDH in response to signals associated with plant defense has been demonstrated. © 2003 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.

Patterns of population structure and environmental associations to aridity across the range of loblolly pine (Pinus taeda L.;Pinaceae)

  • Eckert, A. J.
  • Van Heerwaarden, J.
  • Wegrzyn, J. L.
  • Nelson, C. D.
  • Ross-Ibarra, J.
  • González-Martínez, S. C.
  • Neale, D. B.
Natural populations of forest trees exhibit striking phenotypic adaptations to diverse environmental gradients, thereby making them appealing subjects for the study of genes underlying ecologically relevant phenotypes. Here, we use a genome-wide data set of single nucleotide polymorphisms genotyped across 3059 functional genes to study patterns of population structure and identify loci associated with aridity across the natural range of loblolly pine (Pinus taeda L.). Overall patterns of population structure, as inferred using principal components and Bayesian cluster analyses, were consistent with three genetic clusters likely resulting from expansions out of Pleistocene refugia located in Mexico and Florida. A novel application of association analysis, which removes the confounding effects of shared ancestry on correlations between genetic and environmental variation, identified five loci correlated with aridity. These loci were primarily involved with abiotic stress response to temperature and drought. A unique set of 24 loci was identified as FST outliers on the basis of the genetic clusters identified previously and after accounting for expansions out of Pleistocene refugia. These loci were involved with a diversity of physiological processes. Identification of nonoverlapping sets of loci highlights the fundamental differences implicit in the use of either method and suggests a pluralistic, yet complementary, approach to the identification of genes underlying ecologically relevant phenotypes. Copyright © 2010 by the Genetics Society of America.

A quantitative trait locus genome scan for porcine muscle fiber traits reveals overdominance and epistasis

  • Estellé, J.
  • Gil, F.
  • Vázquez, J. M.
  • Latorre, R.
  • Ramírez, G.
  • Barragán, M. C.
  • Folch, J. M.
  • Noguera, J. L.
  • Toro, M. A.
  • Pérez-Enciso, M.
Muscle histochemical characteristics are decisive determinants of meat quality. The relative percentage and diameters of the different muscular fiber types influence crucial aspects of meat such as color, tenderness, and ultimate pH. Despite its relevance, however, the information on muscle fiber genetic architecture is scant, because histochemical muscle characterization is a laborious task. Here we report a complete QTL scan of muscle fiber traits in 160 animals from a F2 cross between Iberian and Landrace pigs using 139 markers. We identified 20 genome regions distributed along 15 porcine chromosomes (SSCI, 2, 3, 4, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, and X) with direct and(or) epistatic effects. Epistasis was frequent and some interactions were highly significant. Chromosomes 10 and 11 seemed to behave as hubs; they harbored 2 individual QTL, but also 6 epistatic regions. Numerous individual QTL effects had cryptic alleles, with opposite effects to phenotypic pure breed differences. Many of the QTL identified here coincided with previous reports for these traits in the literature, and there was overlapping with potential candidate genes and previously reported meat quality QTL. © 2008 American Society of Animal Science. All rights reserved.

Enhancement of fish mortality by rhabdovirus infection after immunization with a viral nucleoprotein peptide

  • Estepa, A.
  • Coll, J. M.
A similar sequence to a mouse immunodominant CTL peptide (SYVLQGN, single-letter amino acid code, conserved amino acids underlined) identified in the nucleoproteins of several strains of vesicular stomatitis virus (VSV) (37) was found in the nucleoproteins of viral hemorrhagic septicemia virus (VHSV) of salmonid fish (GYVYQGL in VHSV 07.71 and GYVYQGS in VHSV Makah) and not in the nucleoproteins of other rhabdoviruses. The in vivo immunization of fingerling salmonid fish (rainbow trout Onchorynchus mykiss, W) with this VHSV peptide and their subsequent challenge with VHSV resulted in the enhancement rather than in the reduction of fingerling trout mortality. Possible implications for the development of subunit vaccines against VHSV are discussed.

Sex and testosterone effects on growth, immunity and melanin coloration of nestling Eurasian kestrels

  • Fargallo, J. A.
  • Martínez-Padilla, J.
  • Toledano-Díaz, A.
  • Santiago-Moreno, J.
  • Dávila, J. A.
1. Sex differences in testosterone levels and sex-biased sensitivity to testosterone are the basis of some ideas postulated to account for sex-linked environmental vulnerability during early life. However, sex variation in circulating testosterone levels has been scarcely explored and never manipulated at post-natal stages of birds in the wild. 2. We measured and experimentally increased circulating testosterone levels in nestling Eurasian kestrels Falco tinnunculus. We investigated, possible sexual differences in testosterone levels and the effect of this hormone on growth (body mass and tarsus length) and cell-mediated immunity in males and females. We also explored testosterone effects on rump coloration, a highly variable melanin-based trait in male nestlings. We analysed data on circulating testosterone levels of nestlings in 15 additional bird species. 3. Increased levels of testosterone tended to negatively affect body condition, reduced cell-mediated immune responses in male and female nestlings and also diminished the expression of grey rump coloration in male nestlings. No sex differences were observed in testosterone levels in either control or increased testosterone group nestlings, and no interactions were found between sex and treatment. However, male nestlings showed a lower cell-mediated immune response than females in both groups. 4. Our results indicate first, that a high level of testosterone in all nestlings in a brood entails costs, at least in terms of immunity, coloration and probably growth. Secondly, sex differences in post-natal cell-mediated immunity, and consequently in the capacity to prevent diseases, cannot be explained by sex differences in circulating testosterone levels. Finally, by comparing published data at an interspecific level, contradictory sex patterns in circulating testosterone levels have been found, supporting the idea that circulating testosterone might not be a proximate factor causing sex-dependent vulnerability in bird species. © 2006 The Authors.

Divergence time estimates of East Asian and European pigs based on multiple near complete mitochondrial DNA sequences

  • Fernández, A. I.
  • Alves, E.
  • Óvilo, C.
  • Rodríguez, M. C.
  • Silió, L.
Summary The time since the divergence of European and East Asian domestic pigs and wild boars has been estimated in several phylogenetic analyses, generally based on partial mitochondrial sequences or on a small number of complete mtDNA sequences. In the present study, we obtained a refined estimate of this divergence time based on a set of 32 near-complete mtDNA sequences from wild and domestic pigs of European and Asian types, including 14 new and 18 previously published sequences. A weighted average for different functional mtDNA components resulted in an estimate of 746 000 YBP for the divergence of Asian-type from European-type pigs. In addition, our data allowed us to estimate a divergence time between wild and domestic European pigs of 8500 YBP. However, it must be considered cautiously, as most of the estimated values of this sequence divergence were not different from zero, and isolation between wild and domestic pigs has never been complete. © 2010 The Authors, Animal Genetics © 2010 Stichting International Foundation for Animal Genetics.

An immunohistochemical study of the tonsils in pigs with acute African swine fever virus infection

  • Fernández de Marco, M.
  • Salguero, F. J.
  • Bautista, M. J.
  • Núñez, A.
  • Sánchez-Cordón, P. J.
  • Gómez-Villamandos, J. C.
An immunohistochemical study of the tonsils was carried out to gain further insight in the pathogenesis of acute African swine fever (ASF). Twenty-one pigs were inoculated by intramuscular route with a highly virulent isolate of ASF virus and painlessly killed at 1-7 dpi. Viral antigen was highly distributed in the tonsil from 3 to 4 dpi and an increase in the number of monocyte-macrophages was very evident at the same days post inoculation. This phenomenon was observed together with an increase of the expression of proinflammatory cytokines (Tumour necrosis factor alpha and Interleukin-1 alpha) and the apoptosis of lymphocytes studied by the terminal deoxynucleotidyltransferase-mediated dUTP nick end labelling (TUNEL) technique and haemorrhages. With these results, we can conclude that the tonsil is suffering similar lesions than those observed in other lymphoid organs in acute African swine fever, even when the route of inoculation is the intramuscular and not oral-nasal. © 2006 Elsevier Ltd. All rights reserved.

Purification and characterization of an extracellular cysteine proteinase produced by Micrococcus sp. INIA 528

  • Fernández, J.
  • Mohedano, A. F.
  • Polanco, M. J.
  • Medina, M.
  • Nuñez, M.
Micrococcus sp. INIA 528, a micro-organism isolated from raw ewe's milk Manchego cheese, produced an extracellular proteinase. This enzyme was purified to homogeneity from culture supernatant fluid in two chromatographic steps, with a 29-fold increase of specific activity and a 28% recovery of proteinase activity. The homogeneous protein was characterized biochemically. The molecular weight of the enzyme was determined to be 19.4 kDa by mass spectrometry. The purified enzyme was inhibited by E-64, PMSF and iodoacetamide and activated by cysteine, glutathione, dithiothreitol and β-mercaptoethanol. These results suggest that the enzyme is a cysteine proteinase. Optimal conditions for activity on azocasein were 34°C and a pH of 70. The proteinase preferentially degraded β-casein, while after a longer incubation period α(s1)-casein was also extensively hydrolysed. The proteinase had a K(m) value of 6.12 g l-1 for casein and 2.20 g l-1 for azocasein.

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