Dynamics of the hydrocarbon-degrading Cycloclasticus bacteria during mesocosm-simulated oil spills

We used catalysed reported deposition - fluorescence in situ hybridization (CARD-FISH) to analyse changes in the abundance of the bacterial groups Alphaproteobacteria, Gammaproteobacteria and Bacteroidetes, and of hydrocarbon-degrading Cycloclasticus bacteria in mesocosms that had received polycyclic aromatic hydrocarbons (PAHs) additions. The effects of PAHs were assessed under four contrasting hydrographic conditions in the coastal upwelling system of the Rías Baixas: winter mixing, spring bloom, summer stratification and autumn upwelling. We used realistic additions of water soluble PAHs (approximately 20-30 microg l(-1) equivalent of chrysene), but during the winter period we also investigated the effect of higher PAHs concentrations (10-80 microg l(-1) chrysene) on the bacterial community using microcosms. The most significant change observed was a significant reduction (68 +/- 5%) in the relative abundance of Alphaproteobacteria. The magnitude of the response of Cycloclasticus bacteria (positive with probe CYPU829) to PAHs additions varied depending on the initial environmental conditions, and on the initial concentration of added PAHs. Our results clearly show that bacteria of the Cycloclasticus group play a major role in low molecular weight PAHs biodegradation in this planktonic ecosystem. Their response was stronger in colder waters, when their background abundance was also higher. During the warm periods, the response of Cycloclasticus was limited, possibly due to both, a lower bioavailability of PAHs caused by abiotic factors (solar radiation, temperature), and by inorganic nutrient limitation of bacterial growth.     

Prediction of folding pathway and kinetics among plant hemoglobins using an average distance map method

Computational methods, such as the ADM (average distance map) method, have been developed to predict folding of homologous proteins. In this work we used the ADM method to predict the folding pathway and kinetics among selected plant nonsymbiotic (nsHb), symbiotic (Lb), and truncated (tHb) hemoglobins (Hbs). Results predicted that (1) folding of plant Hbs occurs throughout the formation of compact folding modules mostly formed by helices A, B, and C, and E, F, G, and H (folding modules A/C and E/H, respectively), and (2) primitive (moss) nsHbs fold in the C-->N direction, evolved (monocot and dicot) nsHbs fold either in the C-->N or N-->C direction, and Lbs and plant tHbs fold in the C-->N direction. We also predicted relative folding rates of plant Hbs from qualitative analyses of the stability of subdomains and classified plant Hbs into fast and moderate folding. ADM analysis of nsHbs predicted that prehelix A plays a role during folding of the N-terminal domain of Ceratodon nsHb, and that CD-loop plays a role in folding of primitive (Physcomitrella and Ceratodon) but not evolved nsHbs. Modeling of the rice Hb1 A/C and E/H modules showed that module E/H overlaps to the Mycobacterium tuberculosis HbO two-on-two folding. This observation suggests that module E/H is an ancient tertiary structure in plant Hbs.     

Net production and consumption of fluorescent colored dissolved organic matter by natural bacterial assemblages growing on marine phytoplankton exudates

An understanding of the distribution of colored dissolved organic matter (CDOM) in the oceans and its role in the global carbon cycle requires a better knowledge of the colored materials produced and consumed by marine phytoplankton and bacteria. In this work, we examined the net uptake and release of CDOM by a natural bacterial community growing on DOM derived from four phytoplankton species cultured under axenic conditions. Fluorescent humic-like substances exuded by phytoplankton (excitation/emission [Ex/Em] wavelength, 310 nm/392 nm; Coble's peak M) were utilized by bacteria in different proportions depending on the phytoplankton species of origin. Furthermore, bacteria produced humic-like substances that fluoresce at an Ex/Em wavelength of 340 nm/440 nm (Coble's peak C). Differences were also observed in the Ex/Em wavelengths of the protein-like materials (Coble's peak T) produced by phytoplankton and bacteria. The induced fluorescent emission of CDOM produced by prokaryotes was an order of magnitude higher than that of CDOM produced by eukaryotes. We have also examined the final compositions of the bacterial communities growing on the exudates, which differed markedly depending on the phytoplankton species of origin. Alteromonas and Roseobacter were dominant during all the incubations on Chaetoceros sp. and Prorocentrum minimum exudates, respectively. Alteromonas was the dominant group growing on Skeletonema costatum exudates during the exponential growth phase, but it was replaced by Roseobacter afterwards. On Micromonas pusilla exudates, Roseobacter was replaced by Bacteroidetes after the exponential growth phase. Our work shows that fluorescence excitation-emission matrices of CDOM can be a helpful tool for the identification of microbial sources of DOM in the marine environment, but further studies are necessary to explore the association of particular bacterial groups with specific fluorophores.     

Linkages between bacterioplankton community composition, heterotrophic carbon cycling and environmental conditions in a highly dynamic coastal ecosystem

We used mesocosm experiments to study the bacterioplankton community in a highly dynamic coastal ecosystem during four contrasting periods of the seasonal cycle: winter mixing, spring phytoplankton bloom, summer stratification and autumn upwelling. A correlation approach was used in order to measure the degree of coupling between the dynamics of major bacterial groups, heterotrophic carbon cycling and environmental factors. We used catalysed reporter deposition-fluorescence in situ hybridization to follow changes in the relative abundance of the most abundant groups of bacteria (Alphaproteobacteria, Gammaproteobacteria and Bacteroidetes). Bacterial carbon flux-related variables included bacterial standing stock, bacterial production and microbial respiration. The environmental factors included both, biotic variables such as chlorophyll-a concentration, primary production, phytoplankton extracellular release, and abiotic variables such as the concentration of dissolved inorganic and organic nutrients. Rapid shifts in the dominant bacterial groups occurred associated to environmental changes and bacterial bulk functions. An alternation between Alphaproteobacteria and Bacteroidetes was observed associated to different phytoplankton growth phases. The dominance of the group Bacteroidetes was related to high bacterial biomass and production. We found a significant, non-spurious, linkage between the relative abundances of major bacterial groups and bacterial carbon cycling. Our results suggest that bacteria belonging to these major groups could actually share a function in planktonic ecosystems.     

Failure to upregulate cell surface PD-1 is associated with dysregulated stimulation of T cells by TGN1412-like CD28 superagonist

The CD28 superagonist (CD28SA) TGN1412 was administered to humans as an agent that can selectively activate and expand regulatory T cells but resulted in uncontrolled T cell activation accompanied by cytokine storm. The molecular mechanisms that underlie this uncontrolled T cell activation are unclear. Physiological activation of T cells leads to upregulation of not only activation molecules but also inhibitory receptors such as PD-1. We hypothesized that the uncontrolled activation of CD28SA-stimulated T cells is due to both the enhanced expression of activation molecules and the lack of or reduced inhibitory signals. In this study, we show that anti-CD3 antibody-stimulated human T cells undergo time-limited controlled DNA synthesis, proliferation and interleukin-2 secretion, accompanied by PD-1 expression. In contrast, CD28SA-activated T cells demonstrate uncontrolled activation parameters including enhanced expression of LFA-1 and CCR5 but fail to express PD-1 on the cell surface. We demonstrate the functional relevance of the lack of PD-1 mediated regulatory mechanism in CD28SA-stimulated T cells. Our findings provide a molecular explanation for the dysregulated activation of CD28SA-stimulated T cells and also highlight the potential for the use of differential expression of PD-1 as a biomarker of safety for T cell immunostimulatory biologics.     

Red tide assemblage formation in an estuarine upwelling ecosystem: Ria de Vigo

Red tides are conspicuous in the upwelling system of Galicia(NW Iberian Peninsula). At present, there are conflicting hypothesesabout the generation site of these phytoplankton assemblages.It is interesting to know whether the rias can be sites of redtide formation or if they act only as accumulation sites ofpopulations advected from shelf waters. A study in the Ra deVigo, carried out during late September 1990, showed the developmentof a red tide assemblage, composed of Alexandrium affinis, Ceraiiumfusus and Gymnodinium catenaium, during a 2 week upwelling-downwellingcycle. Growth occurred at the bottom of the thermocline-topof the nutricline. Above this assemblage, a diatom assemblage(large diatoms) was blooming. Prior to the formation of thered tide, a subsurface chlorophyll maximum made up of smalldiatoms (Nilzschia f. seriaia, Chaeloceros socialis), smallflagellates (<30 µm) and small gymnodinid forms (<30µm) was observed. In the nutrient-depleted upper layer,several autotrophic and large heterotrophic dinoflagellatesdominated. It is suggested that the ratio between the velocityof upward water movement and the depth of the stratified upperlayer (flushing rate, day^–1) is the critical parameterwhich triggers active phytoplankton growth. It can be concludedthat upward water velocities of {small tilde}2.5 m day^–1and a stratified upper layer of 10 m depth (flushing rate 0.25day^–1) are the main physical constraints for red tidedevelopment.