Development and characterization of 11 microsatellite loci for the Mona Island iguana (Cyclura cornuta stejnegeri)

We isolated and characterized 11 microsatellite loci in the Mona Island iguana (Cyclura cornuta stejnegeri). Eleven loci exhibit moderate to high allelic diversity (two to 12 alleles, mean = 4.5) and polymorphism (mean observed heterozygosity, 0.56; range, 0.26 to 0.78) in 41 adults. This marker set has low probability of identity and high parentage exclusion power and will be suitable for studies of paternity, social organization and relatedness in this species.     

Synthesis and structure-activity relationships of uracil derived human GnRH receptor antagonists: (R)-3-[2-(2-amino)phenethyl]-1-(2,6-difluorobenzyl)-6-methyluracils containing a substituted thiophene or thiazole at C-5

The synthesis of a series of (R)-3-[2-(2-amino)phenethyl]-1-(2,6-difluorobenzyl)-6-methyluracils containing a substituted thiophene or thiazole at C-5 is described. SAR around C-5 of the uracil led to the discovery that a 2-thienyl or (2-phenyl)thiazol-4-yl group is required for optimal receptor binding. The best compound from the series had a binding affinity of 2 nM (K(i)) for the human GnRH receptor. A novel and convenient preparation of N-1-(2,6-difluorobenzyl)-6-methyluracil is also described.     

Stress Reactivity to Repeated Low-Level Challenges: A Pilot Study

The purpose of this study was to examine the effects of a mental challenge on cardiovascular and endocrine [epinephrine (EPI), norepinephrine (NE), and cortisol (CORT)] responses to subsequent low-intensity physical exertion. Twelve males (23.25 ± 0.45 years) completed three sessions, including a graded exercise test on a cycle ergometer and two counter-balanced mental stress trials. In the mental challenge-control condition (MC), participants sat quietly for 20 min following a 20 min mental challenge whereas in the mental challenge-exercise condition (MEC) subjects cycled at 35% of maximal oxygen consumption (VO_2max) following the mental challenge. Repeated-measures ANOVAs were used to assess state anxiety (SAI), cardiovascular variables, EPI, NE, and CORT levels across time between conditions. Participants reported significantly greater increases in SAI scores immediately after the mental challenge, which then decreased post-challenge in both conditions. Neither EPI or NE demonstrated an alteration in levels in either condition, but CORT significantly increased after the mental challenge in both conditions and then maintained a significantly greater level during the MEC compared to the MC condition from midexercise through 15 min of recovery. Area-under-the-curve calculations for CORT was significantly greater in the MEC compared to the MC. Results suggest that the initial mental challenge may have acted to enhance the overall adrenal response to the subsequent anticipation of and actual participation in the low-level physical challenge.     

Epidemiological risk factors of Aujeszky’s disease in wild boars (<Emphasis Type="Italic">Sus scrofa</Emphasis>) and domestic pigs in Spain

An Aujeszky’s disease (AD) control and eradication campaign started in Spain in 1995, and as a result, AD virus (ADV) seroprevalences have notably diminished in Spanish domestic pig herds, but eradication has not yet been achieved. Since the presence of ADV in Spanish wild boar populations can impede schemes to eradicate ADV in domestic pig, we conducted analyses of risk factors and investigated associations between the patterns of ADV seroprevalence at the municipal level in the wild boar and the domestic pig, respectively, in south-central Spain. There was a clear influence of individual factors on the risk of wild boar to test positive to ADV. Concerning the domestic pig, higher ADV seroprevalences were observed with increasing numbers of pigs per farm. Indoor pig farms showed higher seroprevalences than open-air farms. Our results show that, at least at the study scale, the level of ADV seroprevalence in domestic pig farms is not influenced by the level of ADV seroprevalence in the coexisting wild boar populations. Also, there was no influence of domestic pigs ADV seroprevalence on the risk of wild boars to test seropositive. Finally, there was no evidence for a statistical association between ADV seroprevalence levels in wild boar and domestic pig at the municipal level in the study region. These findings are of interest for both veterinary and public health authorities. Nevertheless, more effort is needed in order to elucidate the nature of particular ADV outbreaks in domestic pigs in the study region due to possible contacts with wild boars, which will probably require a molecular approach.     

Genetic mosaic in a marine species flock

We used molecular approaches to study the status of speciation in coral reef fishes known as hamlets (Serranidae: Hypoplectrus). Several hamlet morphospecies coexist on Caribbean reefs, and mate assortatively with respect to their strikingly distinct colour patterns. We provide evidence that, genetically, the hamlets display characteristics common in species flocks on land and in freshwaters. Substitutions within two mitochondrial DNA (mtDNA) protein-coding genes place hamlets within a monophyletic group relative to members of two related genera (Serranus and Diplectrum), and establish that the hamlet radiation must have been very recent. mtDNA distances separating hamlet morphospecies were slight (0.6 +/- 0.04%), yielding a coalescent estimate for the age of the hamlet flock of approximately 430 000 years. Morphospecies did not sort into distinct mtDNA haplotype phylogroups, and alleles at five hypervariable microsatellite loci were shared broadly across species boundaries. None the less, molecular variation was not distributed at random. Analyses of mtDNA haplotype frequencies and nested clades in haplotype networks revealed significant genetic differences between geographical regions and among colour morphospecies. We also observed significant microsatellite differentiation between geographical regions and in Puerto Rico, among colour morphospecies; the latter providing evidence for reproductive isolation between colour morphospecies at this locale. In our Panama collection, however, colour morphospecies were mostly genetically indistinguishable. This mosaic pattern of DNA differentiation implies a complex interaction between population history, mating behaviour and geography and suggests that porous boundaries separate species in this flock of brilliantly coloured coral reef fishes.     

Improvement of biomaterials used in tissue engineering by an ageing treatment

Biomaterials based on crosslinked sponges of biopolymers have been extensively used as scaffolds to culture mammal cells. It is well known that single biopolymers show significant change over time due to a phenomenon called physical ageing. In this research, it was verified that scaffolds used for skin tissue engineering (based on gelatin, chitosan and hyaluronic acid) express an ageing-like phenomenon. Treatments based on ageing of scaffolds improve the behavior of skin-cells for tissue engineering purposes. Physical ageing of dry scaffolds was studied by differential scanning calorimetry and was modeled with ageing kinetic equations. In addition, the physical properties of wet scaffolds also changed with the ageing treatments. Scaffolds were aged up to 3 weeks, and then skin-cells (fibroblasts) were seeded on them. Results indicated that adhesion, migration, viability, proliferation and spreading of the skin-cells were affected by the scaffold ageing. The best performance was obtained with a 2-week aged scaffold (under cell culture conditions). The cell viability inside the scaffold was increased from 60 % (scaffold without ageing treatment) to 80 %. It is concluded that biopolymeric scaffolds can be modified by means of an ageing treatment, which changes the behavior of the cells seeded on them. The ageing treatment under cell culture conditions might become a bioprocess to improve the scaffolds used for tissue engineering and regenerative medicine.     

Antigenic and molecular characterization of Vibrio ordalii strains isolated from Atlantic salmon Salmo salar in Chile

Biochemical, serological and molecular properties of a group of 14 Vibrio ordalii strains isolated from cultured Atlantic salmon Salmo salar in Chile in recent years were studied. The characteristics of isolates were compared with the type strain V. ordalii ATCC 33509T. The Chilean V. ordalii represented a biochemically homogenous group; however, some minor differences with the type strain were observed. The serological relationships among isolates, as well as the study of their antigenic determinant (LPS) revealed a strong reaction with antisera raised against Atlantic salmon strains and the antiserum raised against Listonella anguillarum serotype O2. However, LPS electrophoretic patterns were completely different from the V. ordalii type strain, regardless of the serum employed, suggesting the possibility that the Chilean strains constitute a new serological subgroup within this bacterial species. Genetic analyses by PFGE, RAPD, REP-PCR and ERIC-PCR demonstrated that all V. ordalii strains were genetically homogenous, displaying similar DNA patterns, regardless of the techniques used. Moreover, the analysis of DNA banding patterns generated by ERIC-PCR and REP-PCR also clearly separated the type strain from the Chilean strains. This is the first report of characterization of V. ordalii strains from the Southeastern Pacific area, the results of which should facilitate the development of vaccines for protecting cultured Atlantic salmon against vibriosis in this area.     

The genes that encode the gonococcal transferrin binding proteins,TbpB and TbpA,are differentially regulated by MisR under iron‐replete and iron‐depleted conditions

Neisseria gonorrhoeae produces two transferrin binding proteins, TbpA and TbpB, which together enable efficient iron transport from human transferrin. We demonstrate that expression of the tbp genes is controlled by MisR, a response regulator in the two‐component regulatory system that also includes the sensor kinase MisS. The tbp genes were up‐regulated in the misR mutant under iron‐replete conditions but were conversely down‐regulated in the misR mutant under iron‐depleted conditions. The misR mutant was capable of transferrin‐iron uptake at only 50% of wild‐type levels, consistent with decreased tbp expression. We demonstrate that phosphorylated MisR specifically binds to the tbpBA promoter and that MisR interacts with five regions upstream of the tbpB start codon. These analyses confirm that MisR directly regulates tbpBA expression. The MisR binding sites in the gonococcus are only partially conserved in Neisseria meningitidis, which may explain why tbpBA was not MisR‐regulated in previous studies using this related pathogen. This is the first report of a trans‐acting protein factor other than Fur that can directly contribute to gonococcal tbpBA regulation.     

Temperature Response and Effect of Ca2+ and Mg2+ on ATPases from Roots of Oats and Wheat as Influenced by Growth Temperature and Nutritional Status

Activation by Ca²⁺, Mg²⁺, Zn²⁺, or Mn²⁺ of adenosine triphosphatases in a microsomal fraction from wheat roots depends upon the growth temperature when the plants are grown under low salt conditions, but not when the plants get a full-strength culture medium. At low ionic strength, cultivation at 25°C gives only half as high activation as cultivation at 18°C or at high ionic strength at both temperatures. Corresponding data for activation of ATPases from oats also show that low ionic strength during growth gives the highest temperature dependence. Low temperature together with low salt conditions during growth gives the highest ATPase activity after stimulation with divalent cations. High growth temperature and full-strength medium decrease the ATPase activity. Activation energies (Ea) were calculated for the two temperature intervals 35–20°C and 20–5°C. The dominating ATPase stimulation (Ca²⁺ in wheat, Mg²⁺ in oats) is characterized by high specific activity combined with a low Ea value. The differences in ATPase activity between oats and wheat can be correlated with different cultivation requirements known from agriculture.