The novel NhaE-type Na+/H+ antiporter of the pathogenic bacterium Neisseria meningitidis

Neisseria meningitidis is a pathogenic bacterium responsible for meningitis. The mechanisms underlying the control of Na⁺ transmembrane movement, presumably important to pathogenicity, have been barely addressed. To elucidate the function of the components of the Na⁺ transport system in N. meningitidis, an open reading frame from the genome of this bacterium displaying similarity with the NhaE type of Na⁺/H⁺ antiporters was expressed in Escherichia coli and characterized for sodium transport ability. The N. meningitidis antiporter (NmNhaE) was able to complement an E. coli strain devoid of Na⁺/H⁺ antiporters (KNabc) respecting the ability to grow in the presence of NaCl and LiCl. Ion transport assays in everted vesicles prepared from KNabc expressing NmNhaE from a plasmid confirmed its ability to translocate Na⁺ and Li⁺. Here is presented the characterization of the first NhaE from a pathogen, an important contribution to the comprehension of sodium ion metabolism in this kind of microorganisms.     

Pathogen-derived resistance to Citrus tristeza virus (CTV) in transgenic mexican lime (Citrus aurantifolia (Christ.) Swing.) plants expressing its p25 coat protein gene

The p25 coat protein (CP) gene of Citrustristezavirus (CTV) was incorporated to Mexican lime plants and forty-twotransgeniclines were produced, 25 containing the p25 CP gene of thesevere CTV strain T-305 and 17 with that of the mild strain T-317. When plantspropagated from each transgenic line were graft-inoculated with CTV T-305 oraphidinoculated with T-300, two types of response to viral challenge wereobserved: some lines developed CTV symptoms similar to those of non-transgeniccontrols, whereas others exhibited protection against the virus. Thisprotectionconsisted of a proportion of plants, ranging from 10 to 33%, that wereresistantto CTV, and the rest of them that showed a significant delay in virusaccumulation and symptom onset. Protection was efficient against non-homologousCTV strains and was generally accompanied by high accumulation of p25 CP in theprotected lines, which suggest a CP-mediated protection mechanism in mostcases.This is the first report demonstrating pathogen-derived resistance intransgenicplants against a Closterovirus member in its natural host.     

Diet and trophic position of two mackerel species in the archipelago of Madeira,Portugal

The Atlantic chub mackerel Scomber colias and the blue jack mackerel Trachurus picturatus are two abundant species in the Macaronesia region which includes the archipelago of Madeira, Portugal. Both are key species in the trophic web, being important prey for several local top predators, such as seabirds and marine mammals. Nonetheless, little is known about their feeding ecology in oceanic environments. In this study, the authors describe the seasonal variation in the diet of S. colias and T. picturatus in the oceanic region of Madeira throughout a year. Visual inspection of stomach contents revealed that S. colias fed on a broader range of prey groups than T. picturatus, but for both species, zooplankton (particularly calanoid copepods) and fish were the most important food items. The diet of S. colias included a higher proportion of fish, namely Atlantic saury Scomberesox saurus and S. colias, than that of T. picturatus, that included mostly the longspine snipefish Macroramphosus scolopax. T. picturatus consumed a higher proportion of decapods and other copepods. Seasonal variation was found in the diet of both species, with zooplanktonic species being more important in colder months (February to April) for S. colias and during warm months (May to October) for T. picturatus. Their diet in other seasons was dominated by fish. Although they consume similar prey, carbon and nitrogen stable isotope analysis of muscle of S. colias and T. picturatus showed little overlap in their diets, and T. picturatus showed higher δ¹⁵N and a narrower isotopic niche.     

Functional and structural characterization of synthetic cardosin B-derived rennet

The potential of using a synthetic cardosin-based rennet in cheese manufacturing was recently demonstrated with the development and optimization of production of a recombinant form of cardosin B in Kluyveromyces lactis. With the goal of providing a more detailed characterization of this rennet, we herein evaluate the impact of the plant-specific insert (PSI) on cardosin B secretion in this yeast, and provide a thorough analysis of the specificity requirements as well as the biochemical and structural properties of the isolated recombinant protease. We demonstrate that the PSI domain can be substituted by different linker sequences without substantially affecting protein secretion and milk clotting activity. However, the presence of small portions of the PSI results in dramatic reductions of secretion yields in this heterologous system. Kinetic characterization and specificity profiling results clearly suggest that synthetic cardosin B displays lower catalytic efficiency and is more sequence selective than native cardosin B. Elucidation of the structure of synthetic cardosin B confirms the canonical fold of an aspartic protease with the presence of two high mannose-type, N-linked glycan structures; however, there are some differences in the conformation of the flap region when compared to cardosin A. These subtle variations in catalytic properties and the more stringent substrate specificity of synthetic cardosin B help to explain the observed suitability of this rennet for cheese production.     

Cellulase immobilization on magnetic nanoparticles encapsulated in polymer nanospheres

Immobilization of cellulases on magnetic nanoparticles, especially magnetite nanoparticles, has been the main approach studied to make this enzyme, economically and industrially, more attractive. However, magnetite nanoparticles tend to agglomerate, are very reactive and easily oxidized in air, which has strong impact on their useful life. Thus, it is very important to provide proper surface coating to avoid the mentioned problems. This study aimed to investigate the immobilization of cellulase on magnetic nanoparticles encapsulated in polymeric nanospheres. The support was characterized in terms of morphology, average diameter, magnetic behavior and thermal decomposition analyses. The polymer nanospheres containing encapsulated magnetic nanoparticles showed superparamagnetic behavior and intensity average diameter about 150 nm. Immobilized cellulase exhibited broader temperature stability than in the free form and great reusability capacity, 69% of the initial enzyme activity was maintained after eight cycles of use. The magnetic support showed potential for cellulase immobilization and allowed fast and easy biocatalyst recovery through a single magnet.

     

Ageing increases vulnerability to aβ42 toxicity in Drosophila

Age is the major risk factor for many neurodegenerative diseases, including Alzheimer's Disease (AD), for reasons that are not clear. The association could indicate that the duration or degree of exposure to toxic proteins is important for pathology, or that age itself increases susceptibility to protein toxicity. Using an inducible Drosophila model of AD, we investigated these possibilities by varying the expression of an Aβ42 transgene in neurons at different adult ages and measuring the effects on Aβ42 levels and associated pathological phenotypes. Acute induction of Arctic Aβ42 in young adult flies resulted in rapid expression and clearance of mRNA and soluble Arctic Aβ42 protein, but in irreversible expression of insoluble Arctic Aβ42 peptide. Arctic Aβ42 peptide levels accumulated with longer durations of induction, and this led to a dose-dependent reduction in negative geotaxis and lifespan. For a standardised level of mRNA expression, older flies had higher levels of Arctic Aβ42 peptide and associated toxicity, and this correlated with an age-dependent reduction in proteasome activity. Equalising Aβ42 protein at different ages shortened lifespan in correlation with the duration of exposure to the peptide, suggesting that Aβ42 expression accumulates damage over time. However, the relative reduction in lifespan compared to controls was greater in flies first exposed to the peptide at older ages, suggesting that ageing itself also increases susceptibility to Aβ42 toxicity. Indeed older flies were more vulnerable to chronic Aβ42 toxicity even with a much lower lifetime exposure to the peptide. Finally, the persistence of insoluble Aβ42 in both young and old induced flies suggests that aggregated forms of the peptide cause toxicity in later life. Our results suggest that reduced protein turnover, increased duration of exposure and increased vulnerability to protein toxicity at later ages in combination could explain the late age-of-onset of neurodegenerative phenotypes.     

Extracellular lipid droplets promote hemozoin crystallization in the gut of the blood fluke Schistosoma mansoni

Hemozoin (Hz) is a heme crystal produced upon hemoglobin digestion as the main mechanism of heme disposal in several hematophagous organisms. Here, we show that, in the helminth Schistosoma mansoni, Hz formation occurs in extracellular lipid droplets (LDs). Transmission electron microscopy of adult worms revealed the presence of numerous electron-lucent round structures similar to LDs in gut lumen, where multicrystalline Hz assemblies were found associated to their surfaces. Female regurgitates promoted Hz formation in vitro in reactions partially inhibited by boiling. Fractionation of regurgitates showed that Hz crystallization activity was essentially concentrated on lower density fractions, which have small amounts of pre-formed Hz crystals, suggesting that hydrophilic-hydrophobic interfaces, and not Hz itself, play a key catalytic role in Hz formation in S. mansoni. Thus, these data demonstrate that LDs present in the gut lumen of S. mansoni support Hz formation possibly by allowing association of heme to the lipid-water interface of these structures.     

Genetic demography of Antioquia (Colombia) and the Central Valley of Costa Rica

We report a comparative genetic characterization of two population isolates with parallel demographic histories: the Central Valley of Costa Rica (CVCR) and Antioquia (in northwest Colombia). The analysis of mtDNA, Y-chromosome and autosomal polymorphisms shows that Antioquia and the CVCR are genetically very similar, indicating that closely related parental populations founded these two isolates. In both populations, the male ancestry is predominantly European, whereas the female ancestry is mostly Amerind. In agreement with their isolation, the Amerindian mtDNA diversity of Antioquia and the CVCR is typical of ethnically-defined native populations and is markedly lower than in other Latin American populations. A comparison of linkage disequilibrium (LD) at 18 marker pairs in Antioquia and the CVCR shows that markers in LD in both populations are located at short genetic distances (相似文献   

Enhanced fungal resistance in transgenic cotton expressing an endochitinase gene from Trichoderma virens

Mycoparasitic fungi are proving to be rich sources of antifungal genes that can be utilized to genetically engineer important crops for resistance against fungal pathogens. We have transformed cotton and tobacco plants with a cDNA clone encoding a 42 kDa endochitinase from the mycoparasitic fungus, Trichoderma virens. Plants from 82 independently transformed callus lines of cotton were regenerated and analysed for transgene expression. Several primary transformants were identified with endochitinase activities that were significantly higher than the control values. Transgene integration and expression was confirmed by Southern and Northern blot analyses, respectively. The transgenic endochitinase activities were examined in the leaves of transgenic tobacco as well as in the leaves, roots, hypocotyls and seeds of transgenic cotton. Transgenic plants with elevated endochitinase activities also showed the expected 42 kDa endochitinase band in fluorescence, gel-based assays performed with the leaf extracts in both species. Homozygous T2 plants of the high endochitinase-expressing cotton lines were tested for disease resistance against a soil-borne pathogen, Rhizoctonia solani and a foliar pathogen, Alternaria alternata. Transgenic cotton plants showed significant resistance to both pathogens.