Growth, nitrogen utilization and biodiesel potential for two chlorophytes grown on ammonium, nitrate or urea

Nitrogen removal from wastewater by algae provides the potential benefit of producing lipids for biodiesel and biomass for anaerobic digestion. Further, ammonium is the renewable form of nitrogen produced during anaerobic digestion and one of the main nitrogen sources associated with wastewater. The wastewater isolates Scenedesmus sp. 131 and Monoraphidium sp. 92 were grown with ammonium, nitrate, or urea in the presence of 5 % CO₂, and ammonium and nitrate in the presence of air to optimize the growth and biofuel production of these chlorophytes. Results showed that growth on ammonium, in both 5 % CO₂ and air, caused a significant decrease in pH during the exponential phase causing growth inhibition due to the low buffering capacity of the medium. Therefore, biological buffers and pH controllers were utilized to prevent a decrease in pH. Growth on ammonium with pH control (synthetic buffers or KOH dosing) demonstrated that growth (rate and yield), biodiesel production, and ammonium utilization, similar to nitrate- and urea-amended treatments, can be achieved if sufficient CO₂ is available. Since the use of buffers is economically limited to laboratory-scale experiments, chemical pH control could bridge the gap encountered in the scale-up to industrial processes.     

Depletion of nucleoporins from HeLa nuclear pore complexes to facilitate the production of ghost pores for in vitro reconstitution

During cell division, Nuclear Pore Complexes (NPCs) are broken down into protein subcomplexes that are the basis for reassembly in daughter cells. This is the driving force for the establishment of an in vitro reconstitution system to study aspects of NPC reassembly. In this study, nuclear envelope (NE) was isolated from HeLa cells. NE was treated with increasing concentrations of heparin to extract nucleoporins (Nups) for the production of “ghost pores” which are pores severely deficient in Nups, while still containing Pore Membrane proteins (POM) needed to anchor the NPC. Ghost pores have been subjected to incubation with previously stripped Nups and some re-binding has been shown to occur by western blot analysis. This in vitro assay provides a powerful tool to investigate the protein–protein interactions of NPC reassembly from a human cell line. Through a better understanding of the process of NPC reassembly, we can continue to piece together the puzzle of this macromolecular structure. It is most advantageous to establish a straightforward reconstitution procedure at the mammalian level.     

Heat stress but not inbreeding affects offensive sperm competitiveness in Callosobruchus maculatus

Environmental and genetic stress have well‐known detrimental effects on ejaculate quality, but their concomitant effect on male fitness remains poorly understood. We used competitive fertilization assays to expose the effects of stress on offensive sperm competitive ability in the beetle Callosobruchus maculatus, a species where ejaculates make up more than 5% of male body mass. To examine the effects of environmental and genetic stress, males derived from outcrosses or sib matings were heat shocked at 50°C for 50 min during the pupal stage, while their siblings were maintained at a standard rearing temperature of 28°C. Heat‐shocked males achieved only half the offensive paternity success of their siblings. While this population exhibited inbreeding depression in body size, sperm competitiveness was unaffected by inbreeding, nor did the effect of heat shock stress on sperm competitiveness depend on inbreeding status. In contrast, pupal emergence success was increased by 34% among heat‐stressed individuals, regardless of their inbreeding status. Heat‐shocked males' ejaculate size was 19% reduced, but they exhibited 25% increased mating duration in single mating trials. Our results highlight both the importance of stress in postcopulatory sexual selection, and the variability among stressors in affecting male fitness.     

Determining indicator taxa across spatial and seasonal gradients in the Columbia River coastal margin

Bacterioplankton communities are deeply diverse and highly variable across space and time, but several recent studies demonstrate repeatable and predictable patterns in this diversity. We expanded on previous studies by determining patterns of variability in both individual taxa and bacterial communities across coastal environmental gradients. We surveyed bacterioplankton diversity across the Columbia River coastal margin, USA, using amplicon pyrosequencing of 16S rRNA genes from 596 water samples collected from 2007 to 2010. Our results showed seasonal shifts and annual reassembly of bacterioplankton communities in the freshwater-influenced Columbia River, estuary, and plume, and identified indicator taxa, including species from freshwater SAR11, Oceanospirillales, and Flavobacteria groups, that characterize the changing seasonal conditions in these environments. In the river and estuary, Actinobacteria and Betaproteobacteria indicator taxa correlated strongly with seasonal fluctuations in particulate organic carbon (ρ=−0.664) and residence time (ρ=0.512), respectively. In contrast, seasonal change in communities was not detected in the coastal ocean and varied more with the spatial variability of environmental factors including temperature and dissolved oxygen. Indicator taxa of coastal ocean environments included SAR406 and SUP05 taxa from the deep ocean, and Prochlorococcus and SAR11 taxa from the upper water column. We found that in the Columbia River coastal margin, freshwater-influenced environments were consistent and predictable, whereas coastal ocean community variability was difficult to interpret due to complex physical conditions. This study moves beyond beta-diversity patterns to focus on the occurrence of specific taxa and lends insight into the potential ecological roles these taxa have in coastal ocean environments.     

Reduction of product-related species during the fermentation and purification of a recombinant IL-1 receptor antagonist at the laboratory and pilot scale

Through a parallel approach of tracking product quality through fermentation and purification development, a robust process was designed to reduce the levels of product-related species. Three biochemically similar product-related species were identified as byproducts of host-cell enzymatic activity. To modulate intracellular proteolytic activity, key fermentation parameters (temperature, pH, trace metals, EDTA levels, and carbon source) were evaluated through bioreactor optimization, while balancing negative effects on growth, productivity, and oxygen demand. The purification process was based on three non-affinity steps and resolved product-related species by exploiting small charge differences. Using statistical design of experiments for elution conditions, a high-resolution cation exchange capture column was optimized for resolution and recovery. Further reduction of product-related species was achieved by evaluating a matrix of conditions for a ceramic hydroxyapatite column. The optimized fermentation process was transferred from the 2-L laboratory scale to the 100-L pilot scale and the purification process was scaled accordingly to process the fermentation harvest. The laboratory- and pilot-scale processes resulted in similar process recoveries of 60 and 65%, respectively, and in a product that was of equal quality and purity to that of small-scale development preparations. The parallel approach for up- and downstream development was paramount in achieving a robust and scalable clinical process.