Validez de constructo de la escala motivos de abandono de centros deportivos / Construct validity of the scale attrition sport centres

El abandono de los centros deportivos es un problema social y económico. Disponer de instrumentos válidos y fiables permite la realización de investigaciones rigurosas. El objetivo del estudio fue constatar la validez de la escala de abandono de centros deportivos. Se ha realizado un análisis factorial confirmatorio con el programa AMOS. 20. Posteriormente se ha constatado la validez discriminante mediante tres procedimientos diferentes con los programas SPSS. 20 y AMOS. 20: cálculo de correlaciones entre factores, construcción de intervalos de confianza de la correlación de factores al 95% de confianza y estimación de modelos alternativos. El resultado final ha sido una escala de cinco factores y doce ítems que cumple con los requisitos de validez y fiabilidad exigibles en un instrumento de investigación (AU)

Withdrawal from sport centers is a social and economic problem. Having valid and reliable tools allows rigorous research. The aim of this study was to confirm the validity of the withdrawal rate from sport centers. A confirmatory factor analysis was carried out using AMOS. 20 software. Later the discriminant validity was verified by means of three different procedures with SPSS. 20 and AMOS. 20 software programs: calculating correlation between factors, constructing confidence intervals of the factor correlation at 95% confidence level and estimating alternative models. The final result consists of twelve questions (items) and answers. The answers are rated on five levels (factors) that meet the validity and reliability requirements needed in a research tool (AU)

Utilidad de la serie blanca en el diagnóstico diferencial de la mononucleosis infecciosa / Utility of the white series in the differential diagnosis of infectious mononucleosis

Introducción: la mononucleosis infecciosa (MI) es una enfermedad frecuente en la infancia. Nos planteamos comparar la serie blanca de niños con sospecha de MI, en función de la serología positiva/negativa para virus Epstein-Barr (VEB), citomegalovirus (CMV) y Paul-Bunnell. Material y métodos: estudio descriptivo transversal. Se revisaron niños atendidos en Urgencias en 2010-2011, con diagnóstico de síndrome mononucleósico y serología positiva para VEB o CMV e igual número de niños con serologías negativas como grupo de control. Se compararon variables epidemiológicas, clínicas y serológicas. Resultados: se obtuvieron 50 niños con serologías positivas y 50 niños con serologías negativas (edad media de 5,81 años). Tuvieron serología positiva para VEB 44 niños, 2 para CMV y 4 para ambos. De los 48 niños con serología positiva para VEB, 26 eran Paul-Bunnell negativos y 22 positivos, siendo estos 22 niños el total de Paul-Bunnell positivos. La media de linfocitos, monocitos y basófilos fue mayor en niños con serología positiva para VEB y los neutrófilos fueron más bajos. En los dos casos con CMV positivo encontramos cifras de neutrófilos totales mayores. Ninguna edad se asoció con mayor probabilidad de VEB y Paul-Bunnell positivos. Conclusiones: existe predominio de linfocitos, monocitos y basófilos en niños con MI por VEB. El descenso de neutrófilos es la única variación analítica en los niños con MI por CMV. Estos valores analíticos pueden orientarnos en el diagnóstico de MI. Todos los niños con Paul-Bunnell positivo tenían positividad para el VEB sin relación con la edad (AU)

Introduction: the infectious mononucleosis (IM) is a common disease in childhood. We propose to compare the white series of children with suspected IM, based on serology positive/negative for Epstein -Barr Virus (EBV), Cytomegalovirus (CMV) and Paul- Bunnell. Material and methods: descriptive study. Children were reviewed, taken to hospital in 2010-2011, diagnosed with mononucleosis syndrome and positive serology for EBV or CMV, and equal number of children who were seronegative control group. Epidemiology, clinical and serological variables were compared. Results: there were 50 children with positive serology and 50 negative children (mean age 5.81 years ). EBV serology were 44 children, 2 and 4 both CMV. Of the 48 children with positive serology for EBV, 26 were negative Paul- Bunnell and 22 positive, and these 22 children total positive Paul-Bunnell. The average number of lymphocytes, monocytes and basophils was higher in children with positive serology for EBV and neutrophils were lower. Children were CMV negative but elevated neutrophils. No age was associated with increased likelihood of EBV and Paul- Bunnell positive. Conclusions: there is a predominance of lymphocytes, monocytes and basophils in children with EBV IM. The increase in neutrophils is the only analytical variation in children with CMV IM. These analytical values can guide the diagnosis of IM. All children with positive Paul- Bunnell positive for EBV had no relation with age (AU)

Endogenous cytokinin profiles and their relationships to between-family differences during adventitious caulogenesis in Pinus pinea cotyledons.

Caulogenesis in mature stone pine (Pinus pinea L.) cotyledons is promoted, to varying degrees depending on genotype, by exogenous application of the cytokinin (CK) benzyladenine (BA). In the present study, endogenous CK profiles of cotyledons from open-pollinated plants and two families of stone pine with widely differing organogenic capacities were monitored during caulogenesis and linked to previously characterized BA uptake and induction phases. Changes in levels of free bases, ribosides, ribotides and glucosides of both isoprenoid and aromatic CKs were followed. Before BA application, the pool of endogenous CKs in all sets of cotyledons was dominated by isoprenoid ribotides, but 1h after BA exposure, aromatic CKs (mainly active free bases and ribosides of topolins) accounted for more than 90% of the pool. BA N-glucosides were also observed, levels of which (and topolins) rose from 2d until the end of the (six-day) culture period. The CK profiles of the two selected pine families also differed, although the general trends were similar. During the first 6h, levels of BA and meta-topolin were highest in cotyledons from the family with the strongest caulogenic responses, while levels of ribotides and aromatic glucosides were highest in cotyledons from the other family.

Cloning and characterization of a type-A response regulator differentially expressed during adventitious shoot formation in Pinus pinea L.

Type-A response regulators play an important role in cytokinin-induced adventitious shoot formation, acting as negative regulators of cytokinin signal transduction. In this work, we obtained the full-length cDNA clone of a type-A response regulator from the conifer Pinus pinea, designated PipiRR1. The derived peptide sequence showed all the characteristic motifs found in angiosperms. Gene expression analysis showed that the gene was differentially expressed during adventitious shoot formation in P. pinea cotyledons, suggesting that PipiRR1 may play a role in caulogenesis in conifers. This is the first type-A response regulator identified in gymnosperms.

Organogenic responses of Pinus pinea cotyledons to hormonal treatments: BA metabolism and cytokinin content.

Isolated cotyledons from mature Pinus pinea L. embryos were cultured in vitro in a factorial combination of 4.4, 10 and 44.4 microM N6-benzyladenine (BA) for 2, 4, 8, 16 and 35 days to optimize shoot regeneration. Incubation of explants in 44.4 microM BA for 4 days, in place of the standard incubation in 4.4 microM BA for 35 days, reduced the entire culture period to 4 weeks. Shortening the culture period had no significant effect on the caulogenic response or the number of buds formed per cotyledon. To establish the relationship between key moments in the caulogenic process induced by 4.4 microM BA and the endogenous concentrations of the active forms of BA and other isoprenoid-type cytokinins (CKs), we examined uptake, metabolism and amount of BA, as well as the amounts of zeatin, dihydrozeatin and their ribosides in P. pinea cotyledons after 1, 2, 6, 12 and 24 h, and 2, 4, 8, 16 and 35 days of exposure to 8-[14C]BA. Uptake and release of BA were associated with water movement between explants and the medium during the first 8 days of culture. The interconvertible forms of BA were the main metabolites formed in the tissues. Inactivation of BA as a result of conjugation or oxidation was insignificant. The endogenous concentration of BA + N6-benzyladenosine was 20-fold higher than the exogenously applied BA during the competence acquisition phase (Days 0-3). The concentration of isoprenoid-type CKs also increased 16-fold and then decreased during this time. Induction of shoot buds (Days 4-8) was characterized by a second peak of BA uptake by explants that triggered the synthesis of N6-benzyladenosine-5 -monophosphate and by the maintenance of isoprenoid-type CKs. Reestablishment of CK homeostasis marked the shift from the induction phase to the shoot development phase in this organogenic process (Days 8-12).

Genetic transformation of selected mature cork oak (Quercus suber L.) trees.

A transformation system for selected mature cork oak (Quercus suber L.) trees using Agrobacterium tumefaciens has been established. Embryos obtained from recurrent proliferating embryogenic masses were inoculated with A. tumefaciens strains EHA105, LBA4404 or AGL1 harbouring the plasmid pBINUbiGUSint [carrying the neomycin phosphotransferase II (nptII) and beta-glucuronidase (uidA) genes]. The highest transformation efficiency (4%) was obtained when freshly isolated explants were inoculated with A. tumefaciens strain AGL1. Evidence of stable transgene integration was obtained by PCR for the nptII and uidA genes, Southern blotting and expression of the uidA gene. The transgenic embryos were germinated and successfully transferred to soil.

Induction of a virus-specific antibody response to foot and mouth disease virus using the structural protein VP1 expressed in transgenic potato plants.

We have recently communicated the oral and parental immunogenicity of the structural protein VP1 of foot and mouth disease virus (FMDV) expressed in different transgenic plants. Those results clearly indicated the necessity of increasing the expression of the foreign genes in the transgenic plant to avoid additional steps toward the purification and/or concentration of the antigen of interest. Here, we report the production of transgenic potatoes plants containing the VP1 gene cloned under the regulatory activity of either a single (pRok2) or a double (pRok3) copy of the S35 cauliflower mosaic virus (CaMV 35S) promoter, as a strategy for increasing the level of VP1 gene expression. The presence of the VP1 gene in the plants was confirmed by polymerase chain reaction (PCR) and its specific transcription activity was demonstrated by reverse transcriptase-polymerase chain reaction (RT-PCR). The results showed that, although the immunized animals presented a FMDV VP1 specific antibody response and protection against the experimental challenge, no significant differences were demonstrated in the immunizing activity of plant extracts obtained from the pRok2 or pRok3 transformed plants. These results confirm those previously obtained using other plant species allowing the possibility of using plants as antigen expression vectors, and demonstrated that at least in the potato system, the use of double CaMV 35S promoter does not cause a significant increase in the level of the VP1 expressed.

Oral immunogenicity of the plant derived spike protein from swine-transmissible gastroenteritis coronavirus.

Transgenic plants represent an inexpensive alternative to classical fermentation systems for production of recombinant subunit vaccines. Transgenic potato plants were created that express the N-terminal domain of the glycoprotein S (N-gS) from Transmissible gastroenteritis coronavirus (TGEV), containing the major antigenic sites of the protein. Extracts from potato tubers expressing N-gS were inoculated intraperitoneally to mice, and the vaccinated mice developed serum IgG specific for TGEV. Furthermore, when potato tubers expressing N-gS were fed directly to mice, they developed serum antibodies specific for gS protein, demonstrating the oral immunogenicity of the plant derived spike protein from TGEV.

Immunization with potato plants expressing VP60 protein protects against rabbit hemorrhagic disease virus.

The major structural protein VP60 of rabbit hemorrhagic disease virus (RHDV) has been produced in transgenic potato plants under the control of a cauliflower mosaic virus 35S promoter or a modified 35S promoter that included two copies of a strong transcriptional enhancer. Both types of promoters allowed the production of specific mRNAs and detectable levels of recombinant VP60, which were higher for the constructs carrying the modified 35S promoter. Rabbits immunized with leaf extracts from plants carrying this modified 35S promoter showed high anti-VP60 antibody titers and were fully protected against the hemorrhagic disease.

Agrobacterium tumefaciens-mediated transformation of Pinus pinea L. cotyledons: an assessment of factors influencing the efficiency of uidA gene transfer.

This study is the first report of a protocol for transfer and expression of foreign chimeric genes into cotyledons excised from Pinus pinea L. embryos. Agrobacterium tumefaciens EHA105 harbouring the plasmid p35SGUSint was more infective than LBA4404 or C58 GV3850, as determined by the percentage of cotyledons showing uidA expression. Factors which significantly affected the T-DNA transfer included: (1) preinduction and concentration of bacteria, (2) days of coculture and (3) the wounding procedure applied. More efficient transfer of the uidA gene was achieved growing the bacteria in YEP medium at pH 7, infecting the cotyledons according to the sonication-assisted Agrobacterium-mediated transformation procedure with a bacterial density of 1 (OD600 nm) for 5 min, and coculture for 72 h. Using this protocol, 49.7% of the cotyledons showed a diffuse blue staining 7 days after infection. However, all were necrotic 30 days after inoculation. Since a decrease in bacterial density to 0.01 allowed the recovery of about 4% of cotyledons forming buds 1 month after inoculation, we conclude that the high mortality associated with the infection may be related to the hypersensitive response of the plant to bacterial infection.

N-acetyl-D-neuraminic acid lyase generates the sialic acid for colominic acid biosynthesis in Escherichia coli K1.

Colominic acid is a capsular homopolymer from Escherichia coli K1 composed of alpha (2-8)-linked N-acetyl-D-neuraminic acid (NeuAc) residues. Recently, we have described that NeuAc synthesis in this bacterium occurs through the action of NeuAc lyase (EC 4.1.3.3) [ Rodríguez-Aparicio, Ferrero and Reglero (1995) Biochem. J.308, 501-505]. In the present work we analysed and characterized this enzyme. E. coli K1 NeuAc lyase is detected from the early logarithmic phase of growth, is induced by NeuAc and is not repressed by glucose. The enzyme was purified to apparent homogeneity (312-fold) using two types of hydrophobic chromatographies (butyl-agarose and phenyl-Sepharose CL-4B), gel filtration on Sephacryl S-200, and anion-exchange chromatography on DEAE-FPLC. The pure enzyme, whose amino acid composition and N-terminal amino acid sequence are also established, has a native molecular mass, estimated by gel filtration, of 135 +/- 3 kDa, whereas its molecular mass in SDS/PAGE was 33 +/- 1 kDa. The enzyme was able to synthesize and cleave NeuAc in a reversible reaction. The maximal rate of catalysis was achieved in 125 mM Tris/HCl buffer, pH 7.8, at 37 degrees C. Under these conditions, the K(m) values calculated for N-acetyl-D-mannosamine and pyruvate (condensation direction), and NeuAc (hydrolysis direction) were 7.7, 8.3 and 4.8 mM respectively. NeuAc synthesis by the pure enzyme was activated by Ca2+ and inhibited by Mn2+ and NeuAc, whereas the enzyme cleavage direction was inhibited by Ca2+, Mn2+ and pyruvate. The reaction products, NeuAc and pyruvate, and Ca2+ are able to regulate the direction of this enzyme (synthesis or cleavage of sialic acid) and, accordingly, to modulate colominic acid biosynthesis.

'Phytoantibodies': a general vector for the expression of immunoglobulin domains in transgenic plants.

Sequences encoding the immunoglobulin heavy-chain variable (VH) domains were engineered in a new general purpose vector to transform plants via Agrobacterium. The expression of an isolated VH domain (IVD) after introduction into the plant genome has been monitored by northern, western and immunohistochemical analysis. Immunoblotting showed that the polypeptide was stably expressed and accounted for up to 1% of the soluble protein fraction. It is therefore proposed that single immunoglobulin domains of suitable specificity expressed in plants may constitute an effective system to inhibit the activity of molecules involved in plant pathology or plant development.