Stable expression of an active soluble recombinant form of human fucosyltransferase IX in Spodoptera frugiperda Sf9 cells

A secretory form of human α3-fucosyltransferase IX (sFUT9) was overexpressed in Spodoptera frugiperda (Sf9) insect cells using the stable expression vector pIB/V5-His-TOPO and the signal sequence of human interleukin 2 for efficient secretion. sFUT9 was active and its three potential N-glycosylation sites were occupied. sFUT9 efficiently fucosylated the type II acceptors Galbeta4GlcNAC-R and Fucalpha2Galbeta4GlcNAc-R (R = (CH2)3NHCO(CH2)5–NH-biotin) but not the corresponding sialylated acceptor, and only very poorly the type I (Galbeta3GlcNAc-R) related acceptors. sFUT9 showed a clear preference for glycoproteins containing type II acceptors, with values of 121, 113 and 110 microU/million cell for asialofetuin, erythropoietin and asialoerythropoietin, respectively, values approximately 11-fold higher than those obtained for the small acceptors.     

Glucose improves the in vitro viability of Microsporum canis and Trichophyton mentagrophytes var. mentagrophytes

We describe simple and cost-effective methods using carbohydrates to improve the in vitro viability of dermatophytes. Glucose and sucrose in different concentrations (3, 6, 9 and 12%) were used to maintain fifteen strains of M. canis and T. mentagrophytes var. mentagrophytes at 4 and -20 degrees C. The strains were phenotypically analyzed before storage and reevaluated at 1, 3, 6 and 9 months. At 1 and 3 months, any alterations in the viability or phenotype pattern of the stored strains were noted. At 6 months, both dermatophytes were 100% viable, when preserved in glucose (3, 6, 9 and 12%) at -20 degrees C. All T. mentagrophytes strains were also viable in sucrose (12%), at 4 degrees C and -20 degrees C. However, sucrose failed to improve the viability of M. canis at both temperatures. At 9 months, the higher viabilities without pleomorphism were seen for both dermatophytes preserved in glucose (9 and 12%) at -20 degrees C.     

Gas exchange and water relations of three <Emphasis Type="Italic">Vitis vinifera</Emphasis> L. cultivars growing under Mediterranean climate

Optical characteristics, contents of photosynthetic pigments, total soluble sugars, and starch, rates of gas exchange, chlorophyll (Chl) a fluorescence, and leaf water relations were analysed in three Vitis vinifera L. cultivars, Tinto Cão (TC), Touriga Nacional (TN), and Tinta Roriz (TR), grown in Mediterranean climate. Chl content was significantly lower in TC than in TN and TR leaves, while the Chl a/b ratio was higher. TR had the lowest net photosynthetic rate, stomatal conductance, and contents of soluble sugars and starch than TN and TC. In spite of low Chl content, TC showed the lowest photon absorbance and the highest photochemical efficiency of photosystem 2. TC had the lowest predawn and midday leaf water potential. The capability for osmotic adjustment was similar among cultivars and the calculated modulus of elasticity was higher in TC leaves. The typical lighter green leaves of TC seemed to be an adaptive strategy to high irradiance and air temperature associated to water stress.     

The use of Tn5 transposable elements in a gene trapping strategy for the protozoan Leishmania

The use of transposable elements as a gene-trapping strategy is a powerful tool for gene discovery. Herein we describe the development of a transposable system, based on the bacterial Tn5 transposon, which has been used successfully in Leishmania braziliensis. The transposon carries the neomycin phosphotransferase gene, which is expressed only when inserted in-frame with a Leishmania gene present in the target DNA. Four cosmid clones from a L. braziliensis genomic library were used as targets in transposition reactions and four insertional libraries were constructed and transfected in L. braziliensis. Clones resistant to G418 were selected and analysed by immunofluorescence in order to identify the subcellular localisation of the protein coded by the trapped gene. A definitive subcellular localisation for neomycin phosphotransferase/targeted protein fusion was not obtained in any of the four Leishmania clones investigated. However, the constructed transposable element is highly efficient considering the frequency of insertion in large targets and is therefore a useful tool for functional genetic studies in Leishmania. Our data confirm the utility of the Tn5 transposon system for insertion of sequencing priming sites into target DNA. Furthermore, the high frequency of insertion and even distribution are important in studying genomic regions bearing long and polymorphic repetitive sequences.     

Contrasting patterns of mitochondrial and microsatellite genetic structure among Western European populations of tawny owls (Strix aluco)

A recent study of mitochondrial phylogeography of tawny owls (Strix aluco) in western Europe suggested that this species survived the Pleistocene glaciations in three allopatric refugia located in Iberia, Italy, and the Balkans, and the latter was likely the predominant source of postglacial colonization of northern Europe. New data from seven microsatellite loci from 184 individual owls distributed among 14 populations were used to assess the genetic congruence between nuclear and mitochondrial DNA (mtDNA) markers. Microsatellites corroborated the major phylogeographical conclusions reached on the basis of the mtDNA sequences, but also showed important differences leading to novel inferences. Microsatellites corroborated the three major refugia and supported the Balkan origin of northern populations. When corrected for differences in effective population size, microsatellites and mtDNA yielded generally congruent overall estimates of population structure (N*ST=0.12 vs. RST=0.16); however, there was substantial heterogeneity in the RST among the seven nuclear loci that was not correlated with heterozygosity. Populations representing the Balkans postglacial expansion interact with populations from the other two refugia forming two clines near the Alps and the Pyrenees. In both cases, the apparent position of the contact zones differed substantially between markers due to the genetic composition of populations sampled in northern Italy and Madrid. Microsatellite data did not corroborate the lower genetic diversity of northern, recently populated regions as was found with mtDNA; this discrepancy was taken as evidence for a recent bottleneck recovery. Finally, this study suggests that congruence among genetic markers should be more likely in cases of range expansion into new areas than when populations interact across contact zones.     

Environmental Correlates for Species Richness among Amphibians and Reptiles in a Climate Transition Area

Correlations between environmental factors and the distribution of amphibian and reptile species richness were investigated in a climate transition area, Peneda-Gerês National Park (PNPG), in North-Western Portugal. Using presence-data at a local-scale (1 × 1 km), Ecological-Niche Factor Analysis (ENFA) identified a mixture of climatic (precipitation and number of days with fog), topographical (altitude and relief) and habitat factors (number of watercourses and water surfaces, the type of the largest water surface and tree diversity cover), as accurate predictors of species occurrence. Three factors were common for both taxonomic groups, and consistently presented a positive relation with species occurrence: precipitation, number of water surfaces, and tree diversity cover; suggesting a strong coincidence in the environmental correlates that influence amphibian and reptile species richness. Distribution patterns of observed and predicted species richness were compared using a Geographical Information System. Overall, three high species richness areas were predicted in common for both taxonomic groups and two additional areas for amphibians only. These areas matched with the observed species richness but suggested larger areas of high species richness. The location of the PNPG in a biogeographic crossroad, between Euro-Siberian and Mediterranean provinces, emphasised species richness of amphibians and reptiles and suggests a high priority conservation status for this protected area. Most of Central-Northern Portugal is located in a climatic transition area; therefore, increased species richness should be expected for other areas. Local scale studies for other protected areas should be planned as a framework for the development of multi-scale conservation planning by Portuguese authorities.     

The influence of macrofouling on the corrosion behaviour of API 5L X65 carbon steel

Seawater is a complex corrosive system, and biofouling is one of the factors that influences corrosion processes. The behaviour of corrosion associated with the development of macrofouling was investigated during the first 6 months of the successional process. Three treatments were compared: the 'Control' treatment (absence of macrofouling); 'Community' treatment, and 'Barnacle' treatment, where other macroorganisms were excluded. In the Community treatment, the dominant organisms were filamentous macroalgae (23.73%), barnacles (17.51%), hydroids (16.96%) and encrusting bryozoans (9.58%). In the Barnacle treatment, the cover varied between 39.38% and 62.50%. The corrosion potential ranged from -665.75 to -517.50 mV(Ag/AgC l((KCl))) and could not be associated with fouling development. The highest corrosion rate in the control suggests that macrofouling provides a protection against mass loss. The highest percentage of localised attacks was found in the Community treatment. This may indicate that not only barnacles, but also other organisms induce localised corrosion.     

A new species of the interstitial genus <Emphasis Type="Italic">Neopetitia</Emphasis> (Polychaeta,Syllidae, Eusyllinae) from Tenerife,with modified acicular chaetae in males

A new species of Neopetitia San Martín, 2003 is described from intertidal and shallow subtidal soft-bottom stations on the eastern and western coast of Tenerife, Canary Islands. The new species is characterized by the presence in males of a modified acicular chaeta in chaetiger 11. A discussion of known species of the genus is presented.     

Spatial and temporal variability of macroinvertebrate communities in two farmed Mediterranean rice fields

The spatial and temporal variation of macroinvertebrate assemblages was studied in two Portuguese commercial rice agroecosystems under the effect of field management involving the application of pesticides and fertilizers. A faunal succession of organisms was observed on both fields. Grazers were the first to colonize the paddies after a dry period when pesticides were applied, followed by development into nymphs and by an increase in the abundance of the species after the application of fertilizers. At the end of the season when no pesticides or fertilizers were applied, the communities changed with the presence of adult predators as a result of an increase in prey. Insecticide application revealed specific taxa increase due to the lack of competition with the target organism. Macroinvertebrates tended to prefer infested field margins with aquatic, submerged vegetation, revealing a spatial distribution along the paddies. Two different sampling devices were used and proved necessary in documenting the macroinvertebrate communities (grab for benthic and hand-net for pelagic organisms).