Native Plant Regeneration on Abandoned Desert Farmland: Effects of Irrigation, Soil Preparation, and Amendments on Seedling Establishment

Direct seeding methods to revegetate abandoned farmland were tested at a desert site west of Phoenix, Arizona. Native seeds were broadcast onto plots prepared by mulching, imprinting, chiseling, and fertilizing with phosphorous in a split-plot design. Each main plot was split into subplots that were not irrigated, irrigated with saline (3.25 dS/m) well water, or irrigated and hand weeded of Salsola iberica . Native seeds germinated poorly on all treatments, and three annual exotic weeds ( Brassica nigra , S. iberica , and Schismus spp.) dominated the plots. None of the main plot treatments (mulching, imprinting, chiseling, or fertilizing) had a significant effect on seed germination or canopy cover. Irrigation increased plant cover on plots, but weeds dominated the cover (<4% native species, up to 50% weeds). Near the end of the second growing season a seed bank study was conducted in the greenhouse. Undisturbed desert soil had relatively few weed seeds and more native plant seeds than the disturbed agricultural soil samples, which had few viable native seeds and were dominated by Schismus spp., B. nigra , and S. iberica . The results illustrate the difficulty of establishing native plants on abandoned desert farmland due to the dominance of weedy species, the presence of salts in the soil, and the lack of adequate soil moisture in the treatments without supplemental irrigation.     

Successful transduction of liver in hemophilia by AAV-Factor IX and limitations imposed by the host immune response

We have previously shown that a single portal vein infusion of a recombinant adeno-associated viral vector (rAAV) expressing canine Factor IX (F.IX) resulted in long-term expression of therapeutic levels of F.IX in dogs with severe hemophilia B. We carried out a phase 1/2 dose-escalation clinical study to extend this approach to humans with severe hemophilia B. rAAV-2 vector expressing human F.IX was infused through the hepatic artery into seven subjects. The data show that: (i) vector infusion at doses up to 2 x 10(12) vg/kg was not associated with acute or long-lasting toxicity; (ii) therapeutic levels of F.IX were achieved at the highest dose tested; (iii) duration of expression at therapeutic levels was limited to a period of approximately 8 weeks; (iv) a gradual decline in F.IX was accompanied by a transient asymptomatic elevation of liver transaminases that resolved without treatment. Further studies suggested that destruction of transduced hepatocytes by cell-mediated immunity targeting antigens of the AAV capsid caused both the decline in F.IX and the transient transaminitis. We conclude that rAAV-2 vectors can transduce human hepatocytes in vivo to result in therapeutically relevant levels of F.IX, but that future studies in humans may require immunomodulation to achieve long-term expression.     

Ethnobotanical ground‐truthing: indigenous knowledge,floristic inventories and satellite imagery in the upper Rio Negro,Brazil

Aim To assess the utility of indigenous habitat knowledge in studies of habitat diversity in Amazonia. Location Baniwa indigenous communities in Rio Içana, upper Rio Negro, Brazil. Methods Six campinarana vegetation types, recognized and named by a consensus of Baniwa indigenous informants according to salient indicator species, were studied in 15 widely distributed plots. Floristic composition (using Baniwa plant nomenclature only, after frustrated attempts to obtain botanical collection permits), quantitative measures of forest structure and GPS waypoints of the 4‐ha composite plot contours were registered, permitting their location on Landsat satellite images. Non‐metric multidimensional scaling (NMDS) ordination was carried out using pc‐ord software. Results The NMDS ordinations of the plot data revealed a clear gradient of floristic composition that was highly correlated with three quantitative measures of forest structure: basal area, canopy height and satellite reflectance. Main conclusions Baniwa‐defined forest types are excellent predictors of habitat diversity along the structural gradient comprising distinctive white‐sand campinarana vegetation types. Indigenous ecological knowledge, as revealed by satellite imagery and floristic analyses, proves to be a powerful and efficient shortcut to assessing habitat diversity, promoting dialogue between scientific and indigenous worldviews, and promoting joint study and conservation of biodiversity.     

Evidence for polynuclear iron(III) clusters in the root nodule bacterium, Rhizobium leguminosarum bv. viciae WSM710

Cells of the root nodule bacterium Rhizobium leguminosarum bv. viciae WSM710 were cultured in a medium containing 20 M ⁵⁷Fe. Mössbauer spectra of the cells at 5.5 and 3.7 K indicated that the major form of iron present in the cells was in the form of polynuclear iron(III) clusters. At 5.5 K the spectral component associated with these clusters was in the form of a superposition of a broad feature (large magnetic hyperfine field distribution) and a doublet. On lowering the temperature of the cells to 3.7 K, the spectral component was transformed into resolved magnetic hyperfine field splitting which yielded a magnetic hyperfine field of 42.4 T when fitted with broad Lorentzian peaks. These spectral characteristics are typical of the hydrated iron(III) phosphate cores of several bacterioferritins. A small fraction (11%) of the Mössbauer spectral area of the cells was in the form of a doublet which yielded parameters ( = 1.35 mm/s; E_Q = 3.15 mm/s) indicative of iron(II). The parameters are very similar to those of a spectral component previously observed in several other microbes (R. Böhnke and B.F. Matzanke (1995) BioMetals 8, 223-230) and which has been associated with a 2.2 kDa oligomeric iron(II) carbohydrate phosphate     

A selective protein sensor for heparin detection

No clinical assays for the direct detection of heparin in blood exist. To create a heparin sensor, the hyaluronan (HA)-binding domain (HABD) of a protein that binds heparin and HA was engineered. GST fusion proteins containing one to three HABD modules were cloned, expressed, and purified. The affinities of each construct for heparin and for HA were determined by a competitive enzyme-linked immunosorbent assay using immobilized HA or heparin. Each of the constructs showed modest affinity for immobilized HA. However, heparin was 100-fold more potent than HA as a competing ligand. With immobilized heparin, affinity increased as the HABD copy number increased. The three-copy construct, GST-HB3, detected unfractionated free heparin (UFH) as low as 39ng/ml (equivalent to approximately 0.1U/ml) with a signal-to-noise ratio of 5.6. GST-HB3 also showed 100-fold selectivity for heparin in preference to other glycosaminoglycans. The plot of logKd vs log [Na+] showed 2.5 ionic interactions per heparin-HB3 interaction. GST-HB3 showed a linear detection of both UFH (15kDa) and low-molecular-weight heparin (LMWH; 6kDa) added to human plasma. For UFH, the range examined was 78 to over 2000ng/ml (equivalent to 0.2 to 5.0U/ml). For LMWH, the useful range was 312 to over 2000ng/ml. The coefficient of variance for the assay was < 9% for six serial heparin dilutions and <12% for three plasma samples. In clinical use, GST-HB3 could accurately measure therapeutic heparin levels in plasma (0.2 to 2U/ml).     

Differential hydrolysis of erythrocyte and mitochondrial membrane phospholipids by two phospholipase A2 isoenzymes (NK-PLA2-I and NK-PLA2-II) from the venom of the Indian monocled cobra Naja kaouthia

We previously demonstrated that venom from the Indian monocled cobra Naja kaouthia is a rich source of phospholipase A2 enzymes, and we purified and characterized a major PLA2 isoenzyme (NK-PLA2-I) from N. kaouthia venom. In the present study, we report the purification and biochemical characterization of a second PLA2 isoenzyme (NK-PLA2-II) from the same venom. A comparison of the membrane phospholipid hydrolysis patterns by these two PLA2s has revealed that they cause significantly more damage to mitochondrial membranes (NK-PLA2-I > NK-PLA2-II) as compared to erythrocyte membranes due to more efficient binding of the enzymes to mitochondrial membranes. Fatty acid release patterns by these PLA2s from the membrane phospholipid PC-pools indicate that NK-PLA2-I does not discriminate between saturated and unsaturated fatty acids whereas NK-PLA2-II shows a preference for unsaturated fatty acids during the initial phase of attack. The current investigation provides new insight into the molecular arrangement of NK-PLA2-sensitive domains in erythrocyte and mitochondrial membranes and highlights the contribution of polar, but uncharged, amino acids such as serine and cysteine in NK-PLA2 induced membrane damage.     

Extracellular proteolytic activity plays a central role in swarming motility in Bacillus subtilis

Natural isolates of Bacillus subtilis exhibit a robust multicellular behavior known as swarming. A form of motility, swarming is characterized by a rapid, coordinated progression of a bacterial population across a surface. As a collective bacterial process, swarming is often associated with biofilm formation and has been linked to virulence factor expression in pathogenic bacteria. While the swarming phenotype has been well documented for Bacillus species, an understanding of the molecular mechanisms responsible remains largely isolated to gram-negative bacteria. To better understand how swarming is controlled in members of the genus Bacillus, we investigated the effect of a series of gene deletions on swarm motility. Our analysis revealed that a strain deficient for the production of surfactin and extracellular proteolytic activity did not swarm or form biofilm. While it is known that surfactin, a lipoprotein surfactant, functions in swarming motility by reducing surface tension, this is the first report demonstrating that general extracellular protease activity also has an important function. These results not only help to define the factors involved in eliciting swarm migration but support the idea that swarming and biofilm formation may have overlapping control mechanisms.