MARE - IPleiria

Disease assessment and management in cultured aquatic animals is a major concern in commercial aquaculture. Disease outbreaks have direct effects on fish production, causing serious economic losses in this industry. This can be overcome by early detection through molecular high sensitivity tools such as PCR (polymerase chain reaction). One of the most critical steps in the study of bacterial fish diseases is the precise identification of the infectious agent. Considering the damage that some bacteria can cause to fish and humans, the development of a rapid detection method for the four target species that demonstrates to be simple, accurate and low cost is an essential step for the prevention and early treatment of these diseases. Edwardsiella tarda (ACC 36.1), Vibrio harveyi (DSM 19023), Vibrio anguillarum (AQV 55.1) and Vibrio alginolyticus (CECT 521) were selected as targets of a multiplex PCR tool. The multiplex PCR reactions were performed in various reaction conditions, including different annealing temperatures (between 49°C and 55°C) and changes in the MgCl 2 concentration from 2 mM to 8 mM. Best results were obtained at 51°C and MgCl 2 concentration from 4 mM to 6 mM. Primers were tested using purified DNA from the respective bacterial strains, yielding bands at expected sizes, which can be identified on agarose gels with clearness and without overlapping sizes. Results indicate that this multiplex PCR tool is suitable for the detection of target pathogens and may, in the future, have extended practical application in commercial aquaculture.

The effect of salinity and growth temperature on the accumulation of intracellular organic solutes was examined in the hyperthermophilic archaeon Palaeococcus ferrophilus. The genus Palaeococcus represents a deep-branching lineage of the order Thermococcales, which diverged before Thermococcus and Pyrococcus. Palaeococcus ferrophilus accumulated mannosylglycerate, glutamate, and aspartate as major compatible solutes. Unlike members of the genera Pyrococcus and Thermococcus, Palaeococcus ferrophilus did not accumulate di-myo-inositol phosphate, a canonical solute of hyperthermophiles. The level of mannosylglycerate increased in response to both heat and salt stress; glutamate increased at supraoptimal growth temperatures, whereas aspartate increased at supraoptimal salt concentration. Proline, alanine, and trehalose were also found in lesser amounts, but their levels did not respond significantly to any of the stresses imposed. Additionally, the genes involved in the synthesis of mannosylglycerate in Palaeococcus ferrophilus and Thermococcus litoralis were identified. In both organisms the synthesis proceeds via the two-step pathway comprising mannosyl-3-phosphoglycerate synthase (MPGS) (EC 2.4.1.217) and mannosyl-3-phosphoglycerate phosphatase (MPGP) (EC 3.1.3.70). The mpgS and mpgP genes of Palaeococcus ferrophilus were expressed in Escherichia coli and the proteins were characterized. MPGS had maximal activity at 90°C and pH near 7.0, and was strictly dependent on Mg 2؉ . MPGP had optimal activity at 90°C and pH 6.0 and was barely dependent on Mg 2؉ . The half-life values for inactivation of MPGS and MPGP at 83°C were 18 and 25 min, respectively. A comparative discussion of the osmo-and thermoadaptation strategies in these three genera of the Thermococcales is presented.

The random amplified polymorphic DNA (RAPD; Williams et al., 1990) marker system has been used in many different applications involving the detection of DNA sequence polymorphism, but most often in construction of linkage maps and in bulk segregant analysis (BSA) for identification of markers linked to genes of interest. The sequence-related amplified polymorphism (SRAP) technique is relatively a new and highly efficient PCR-based technique, based on amplification using two primers. The primers are designed to preferentially amplify exonic and intronic regions, and create polymorphisms that depend on variation of the length of introns, promoters and spacers (Li and Quiros, 2001). In this report we present results in the development of fast and inexpensive RAPD and SRAP marker protocols and discuss their potential use in identification of markers linked to gene(s) for resistance to sugar beet cyst nematode (Heterodera schachtii Schmidt) using BSA.

The aim of our study is the identification of the critical points in the rooting process at the molecular level for possible improvement of the propagation of cork oak. The rhizogenic potential of woody plants depends on both genotype and developmental stage. To investigate the role of cyclin transcription in the regulation of cell division during adventitious root induction, we have analysed the temporal expression of a member of the cyclin gene family. The histological modifications that occur during the root induction and formation processes as a consequence of auxin treatments can be detected by the technique of tissue printing for hydrogen peroxide (H 2 O 2 ). Thus a H 2 O 2 printing assay was performed in parallel with cyclin expression studies. We report the cloning and sequencing of a B-type cyclin genomic fragment from micropropagated cork oak using gene-specific and genotype-independent oligonucleotide primers to conserved regions of the gene. After 48 h of auxin induction, the expression of this B-type cyclin increased, possibly in association with the first cell divisions. The expression observed in control shoots may reflect unorganised cell divisions associated with callus formation.

Citation: Mouga, T.; Mendes, S.; Franco, I.; Fagundes, A.I.; Oliveira, N.; Crisóstomo, P.; Morais, L.; Afonso, C. Recent Efforts to Recover Armeria berlengensis, an Endemic Species from Berlengas Archipelago, Portugal. Plants 2021, 10, 498. https://doi.

Saffron is a triploid and sterile crop that propagated by replanting rest corms thereby it is difficult that we can improvement properties of its agronomic by old methods. Protoplast culture is an efficient instrument for improvement properties of its agronomic. In monocots such as saffron plantlet regeneration from protoplast that isolated directly from the plant's organs is difficult therefore embryogenic callus proved to be the best source for protoplast isolation. Protoplast isolation was directly from embryogenic callus for 3 h in an enzyme solution consisting of MS medium, 0.2% Pectolyase Y 2-3, 2% Cellulase RS, 0.5% Deriselase, 0.05% MES, and 0.3 M mannitol in the darkness at 22 • C. Isolated protoplast was mixed slowly with steril sodium alginate and was added drop by drop with a sterile pasteur pipette to liquid MS medium which was contained CaCl 2 1% and 0.3 M manitol to create Ca-alginate beads. The beads were transferred to MS liquid medium culture with different concentration of growth regulators. After 4-5 weeks of culture, microcalli appeared on the surface of the Ca-alginate beads. Plantlet regeneration was occurred when beads transferred onto solid MS medium without growth regulators with 6% sucrose after 6 months.

Motivated by our interest in micro and biological air vehicles, Navier-Stokes simulations for fluid flow around a hovering elliptic airfoil have been conducted to investigate the effects of Reynolds number, reduced frequency, and flapping kinematics on the flow structure and aerodynamics. The Reynolds number investigated ranges from 75 to 1700, and the reduced frequency from 0.36 to 2.0. Two flapping modes are studied, namely, the "water treading" hovering mode, and the "normal" hovering mode. While the delayed-stall mechanism is found to be responsible for generating the maximum lift peaks in both hovering modes, the wake-capturing mechanism is identified only in the normal hovering mode. In addition to the strong role played by the kinematics, the Reynolds number's role has also been clearly identified. In the low Reynolds number regime, O(100), the viscosity dissipates the vortex structures quickly and leads to essentially symmetric flow structure and aerodynamics force between the forward stroke and backward strokes. At higher Reynolds numbers (300 and larger), the history effect is influential resulting in distinctly asymmetric phenomena between the forward and backward strokes.