Biomass Production and Assimilation of Dissolved Organic Matter by SAR11 Bacteria in the Northwest Atlantic Ocean

Members of the SAR11 clade often dominate the composition of marine microbial communities, yet their contribution to biomass production and the flux of dissolved organic matter (DOM) is unclear. In addition, little is known about the specific components of the DOM pool utilized by SAR11 bacteria. To better understand the role of SAR11 bacteria in the flux of DOM, we examined the assimilation of leucine (a measure of biomass production), as well as free amino acids, protein, and glucose, by SAR11 bacteria in the Northwest Atlantic Ocean. We found that when SAR11 bacteria were >25% of total prokaryotes, they accounted for about 30 to 50% of leucine incorporation, suggesting that SAR11 bacteria were major contributors to bacterial biomass production and the DOM flux. Specific growth rates of SAR11 bacteria either equaled or exceeded growth rates for the total prokaryotic community. In addition, SAR11 bacteria were typically responsible for a greater portion of amino acid assimilation (34 to 61%) and glucose assimilation (45 to 57%) than of protein assimilation (≤34%). These data suggest that SAR11 bacteria do not utilize various components of the DOM pool equally and may be more important to the flux of low-molecular-weight monomers than to that of high-molecular-weight polymers.     

Contribution of SAR11 bacteria to dissolved dimethylsulfoniopropionate and amino acid uptake in the North Atlantic ocean

SAR11 bacteria are abundant in marine environments, often accounting for 35% of total prokaryotes in the surface ocean, but little is known about their involvement in marine biogeochemical cycles. Previous studies reported that SAR11 bacteria are very small and potentially have few ribosomes, indicating that SAR11 bacteria could have low metabolic activities and could play a smaller role in the flux of dissolved organic matter than suggested by their abundance. To determine the ecological activity of SAR11 bacteria, we used a combination of microautoradiography and fluorescence in situ hybridization (Micro-FISH) to measure assimilation of (3)H-amino acids and [(35)S]dimethylsulfoniopropionate (DMSP) by SAR11 bacteria in the coastal North Atlantic Ocean and the Sargasso Sea. We found that SAR11 bacteria were often abundant in surface waters, accounting for 25% of all prokaryotes on average. SAR11 bacteria were typically as large as, if not larger than, other prokaryotes. Additionally, more than half of SAR11 bacteria assimilated dissolved amino acids and DMSP, whereas about 40% of other prokaryotes assimilated these compounds. Due to their high abundance and activity, SAR11 bacteria were responsible for about 50% of amino acid assimilation and 30% of DMSP assimilation in surface waters. The contribution of SAR11 bacteria to amino acid assimilation was greater than would be expected based on their overall abundance, implying that SAR11 bacteria outcompete other prokaryotes for these labile compounds. These data suggest that SAR11 bacteria are highly active and play a significant role in C, N, and S cycling in the ocean.     

Contribution of SAR11 Bacteria to Dissolved Dimethylsulfoniopropionate and Amino Acid Uptake in the North Atlantic Ocean

SAR11 bacteria are abundant in marine environments, often accounting for 35% of total prokaryotes in the surface ocean, but little is known about their involvement in marine biogeochemical cycles. Previous studies reported that SAR11 bacteria are very small and potentially have few ribosomes, indicating that SAR11 bacteria could have low metabolic activities and could play a smaller role in the flux of dissolved organic matter than suggested by their abundance. To determine the ecological activity of SAR11 bacteria, we used a combination of microautoradiography and fluorescence in situ hybridization (Micro-FISH) to measure assimilation of ³H-amino acids and [³⁵S]dimethylsulfoniopropionate (DMSP) by SAR11 bacteria in the coastal North Atlantic Ocean and the Sargasso Sea. We found that SAR11 bacteria were often abundant in surface waters, accounting for 25% of all prokaryotes on average. SAR11 bacteria were typically as large as, if not larger than, other prokaryotes. Additionally, more than half of SAR11 bacteria assimilated dissolved amino acids and DMSP, whereas about 40% of other prokaryotes assimilated these compounds. Due to their high abundance and activity, SAR11 bacteria were responsible for about 50% of amino acid assimilation and 30% of DMSP assimilation in surface waters. The contribution of SAR11 bacteria to amino acid assimilation was greater than would be expected based on their overall abundance, implying that SAR11 bacteria outcompete other prokaryotes for these labile compounds. These data suggest that SAR11 bacteria are highly active and play a significant role in C, N, and S cycling in the ocean.     

Light-stimulated bacterial production and amino acid assimilation by cyanobacteria and other microbes in the North Atlantic ocean

We examined the contribution of photoheterotrophic microbes--those capable of light-mediated assimilation of organic compounds--to bacterial production and amino acid assimilation along a transect from Florida to Iceland from 28 May to 9 July 2005. Bacterial production (leucine incorporation at a 20 nM final concentration) was on average 30% higher in light than in dark-incubated samples, but the effect varied greatly (3% to 60%). To further characterize this light effect, we examined the abundance of potential photoheterotrophs and measured their contribution to bacterial production and amino acid assimilation (0.5 nM addition) using flow cytometry. Prochlorococcus and Synechococcus were abundant in surface waters where light-dependent leucine incorporation was observed, whereas aerobic anoxygenic phototrophic bacteria were abundant but did not correlate with the light effect. The per-cell assimilation rates of Prochlorococcus and Synechococcus were comparable to or higher than those of other prokaryotes, especially in the light. Picoeukaryotes also took up leucine (20 nM) and other amino acids (0.5 nM), but rates normalized to biovolume were much lower than those of prokaryotes. Prochlorococcus was responsible for 80% of light-stimulated bacterial production and amino acid assimilation in surface waters south of the Azores, while Synechococcus accounted for on average 12% of total assimilation. However, nearly 40% of the light-stimulated leucine assimilation was not accounted for by these groups, suggesting that assimilation by other microbes is also affected by light. Our results clarify the contribution of cyanobacteria to photoheterotrophy and highlight the potential role of other photoheterotrophs in biomass production and dissolved-organic-matter assimilation.     

Uptake of Dissolved Organic Carbon by Gammaproteobacterial Subgroups in Coastal Waters of the West Antarctic Peninsula

Heterotrophic bacteria are well known to be key players in the turnover of dissolved organic material (DOM) in the oceans, but the relationship between DOM uptake and bacterial clades is still not well understood. Here we explore the turnover and single-cell use of glucose, an amino acid mixture, N-acetylglucosamine (NAG), and protein by gammaproteobacterial clades in coastal waters of the West Antarctic Peninsula in summer and fall. More than 60% of the cells within two closely related gammaproteobacterial clades, Ant4D3 and Arctic96B-16, were active in using the amino acid mixture, protein, and NAG. In contrast, an average of only 7% of all SAR86 cells used amino acids and protein even in summer when DOM use was high. In addition to DOM uptake within a group, we explored the contribution of the three gammaproteobacterial groups to total community uptake of a compound. SAR86 contributed 5- to 10-fold less than the other gammaproteobacterial subgroups to the uptake of all compounds. We found that the overall contribution of the Ant4D3 clade to DOM uptake was highest, whereas the SAR86 clade contributed the least to DOM turnover in West Antarctic Peninsula waters. Our results suggest that the low growth activity of a bacterial clade leads to low abundance, fewer active cells and a low contribution to the turnover of DOM components.     

High Contribution of SAR11 to Microbial Activity in the North West Mediterranean Sea

We investigated the abundance and activity of SAR11 on a monthly time scale between January 2008 and October 2008 in the oligotrophic NW Mediterranean Sea. Applying MICRO-CARD-FISH, we observed that SAR11 had a large contribution to bulk abundance (37 ± 6% of DAPI-stained cells) and to bulk bacterial heterotrophic production (BHP), as estimated from leucine incorporation (55 ± 15% of DAPI-cells assimilating leucine) in surface waters (5 m) throughout the study period. SAR11 contributed also substantially to the assimilation of glucose, ATP, and a combination of amino acids (44 ± 17%, 37 ± 14%, and 43 ± 12% of DAPI cells assimilating these compounds, respectively), organic compounds that provide either single or combined sources of C, P, and N. Temporal changes in the abundance of SAR11 cells that assimilated leucine, glucose, amino acids, and ATP revealed a pattern consistent with that of substrate-active DAPI cells, suggesting that the activity of SAR11 can explain to a large extent the variability in total cells contributing to the utilization of these compounds. Short-term nutrient enrichment experiments performed on each sampling date revealed a strong co-limitation of at least two of the three elements analyzed (C, N, P), in particular, during summer and early autumn. The in situ abundance of SAR11 cells assimilating leucine appeared to increase with P limitation as determined in the nutrient enrichment experiments (r = 0.81, p = 0.015). Our results demonstrate that SAR11 is an important component of the active bacterial community in the NW Mediterranean Sea. Our observations further indicate that the activity of the bulk bacterial community is linked to the activity of SAR11, possibly due to its adaptation to nutrient limitation.     

Bacterial Consumption of Humic and Non-Humic Low and High Molecular Weight DOM and the Effect of Solar Irradiation on the Turnover of Labile DOM in the Southern Ocean

The decomposition of dissolved organic matter (DOM) in pelagic ecosystems is mediated primarily by heterotrophic bacteria, but transformation by short-wave solar radiation may play an important role in surface waters, in particular when humic substances constitute a substantial fraction of the DOM pool. Most of the studies examining bacterial decomposition and photochemical transformation of DOM stem from limnetic and coastal marine systems and much less information is available from oceanic environments. To examine the bacterial decomposition of humic and non-humic DOM in the Southern Ocean we carried out microcosm experiments in which we measured bacterial growth on isolated fractions of humic and non-humic DOM of the size classes <3 kDa and >3 kDa. Experiments carried out at the Polar Front showed a preferential bacterial growth on non-humic DOM and in particular on the size fraction <3 kDa. Bacterial growth, measured as bacterial biomass production, on non-humic DOM accounted for 74% to 88% of the total growth on all four DOM fractions. In experiments in the Antarctic circumpolar current and the coastal current under pack ice, bacterial growth was 6× lower than at the Polar Front, and humic and non-humic DOM was consumed to equal amounts. The size fraction <3 kDa was always preferred. Experiments examining the effect of solar radiation on the release of dissolved amino acids (DAA) and carbohydrates (DCHO) and their subsequent bacterial utilization showed a stimulating effect on glucose uptake and the release of DAA at the Polar Front but an inhibition in the eastern Weddell Sea. Ultraviolet-B was the most effective component of the solar radiation spectrum tested. Effects of UV-B on glucose uptake and release of DAA were positively correlated with concentrations of humic-bound DAA. The data imply that at low concentrations, e.g., <100 nM (amino acid equivalent), UV-irradiation reduces, whereas at concentrations >100 nM UV-irradiation stimulates glucose uptake and release of DAA as compared to dark conditions.     

Regulation of Bacterial Growth Rates by Dissolved Organic Carbon and Temperature in the Equatorial Pacific Ocean

Abstract The effect of dissolved organic matter (DOM) and temperature on bacterial production was examined in the equatorial Pacific Ocean. Addition of glucose, glucose plus ammonium, or free amino acids stimulated bacterial production ([³H]thymidine incorporation), whereas changes in bacterial abundance were either negligible or much less than changes in bacterial production. The average bacterial growth rate also greatly increased following DOM additions, whereas in contrast, addition of ammonium alone never affected production, bacterial abundance, or growth rates. Since the large glucose effect was not observed in previous studies of cold oceanic waters, several experiments were conducted to examine DOM-temperature interactions. These experiments suggest that bacteria respond more quickly and to a greater extent to DOM additions at higher temperatures, which may explain apparently conflicting results from previous studies. We also examined how temperate affects the kinetic parameters of sugar uptake. Maximum uptake rates (V_max) of glucose and mannose increased with temperature (Q₁₀= 2.4), although the half-saturation constant (K_m) was unaffected; K_m+ S was roughly equal to glucose concentrations (S) measured by a high pressure liquid chromographic technique. Bacterial production and growth rates appear to be limited by DOM in the equatorial Pacific, and thus bacterial production follows primary production over large spatial and temporal scales in this oceanic regime, as has been observed in other aquatic systems. Although temperature may not limit bacterial growth rates in the equatorial Pacific and similar warm waters, it could still affect how bacteria respond to changes in DOM supply and help set steady-state DOM concentrations. Received: 26 July 1995; Revised: 19 January 1996     

Light-Stimulated Bacterial Production and Amino Acid Assimilation by Cyanobacteria and Other Microbes in the North Atlantic Ocean

We examined the contribution of photoheterotrophic microbes—those capable of light-mediated assimilation of organic compounds—to bacterial production and amino acid assimilation along a transect from Florida to Iceland from 28 May to 9 July 2005. Bacterial production (leucine incorporation at a 20 nM final concentration) was on average 30% higher in light than in dark-incubated samples, but the effect varied greatly (3% to 60%). To further characterize this light effect, we examined the abundance of potential photoheterotrophs and measured their contribution to bacterial production and amino acid assimilation (0.5 nM addition) using flow cytometry. Prochlorococcus and Synechococcus were abundant in surface waters where light-dependent leucine incorporation was observed, whereas aerobic anoxygenic phototrophic bacteria were abundant but did not correlate with the light effect. The per-cell assimilation rates of Prochlorococcus and Synechococcus were comparable to or higher than those of other prokaryotes, especially in the light. Picoeukaryotes also took up leucine (20 nM) and other amino acids (0.5 nM), but rates normalized to biovolume were much lower than those of prokaryotes. Prochlorococcus was responsible for 80% of light-stimulated bacterial production and amino acid assimilation in surface waters south of the Azores, while Synechococcus accounted for on average 12% of total assimilation. However, nearly 40% of the light-stimulated leucine assimilation was not accounted for by these groups, suggesting that assimilation by other microbes is also affected by light. Our results clarify the contribution of cyanobacteria to photoheterotrophy and highlight the potential role of other photoheterotrophs in biomass production and dissolved-organic-matter assimilation.     

Abundance and single-cell activity of heterotrophic bacterial groups in the western Arctic Ocean in summer and winter

Environmental conditions in the western Arctic Ocean range from constant light and nutrient depletion in summer to complete darkness and sea ice cover in winter. This seasonal environmental variation is likely to have an effect on the use of dissolved organic matter (DOM) by heterotrophic bacteria in surface water. However, this effect is not well studied and we know little about the activity of specific bacterial clades in the surface oceans. The use of DOM by three bacterial subgroups in both winter and summer was examined by microautoradiography combined with fluorescence in situ hybridization. We found selective use of substrates by these groups, although the abundances of Ant4D3 (Antarctic Gammaproteobacteria), Polaribacter (Bacteroidetes), and SAR11 (Alphaproteobacteria) were not different between summer and winter in the Beaufort and Chukchi Seas. The number of cells taking up glucose within all three bacterial groups decreased significantly from summer to winter, while the percentage of cells using leucine did not show a clear pattern between seasons. The uptake of the amino acid mix increased substantially from summer to winter by the Ant4D3 group, although such a large increase in uptake was not seen for the other two groups. Use of glucose by bacteria, but not use of leucine or the amino acid mix, related strongly to inorganic nutrients, chlorophyll a, and other environmental factors. Our results suggest a switch in use of dissolved organic substrates from summer to winter and that the three phylogenetic subgroups examined fill different niches in DOM use in the two seasons.     

Effects of different molecular weight fractions of dissolved organic matter on the growth of bacteria,algae and protozoa from a highly humic lake

Effects of different molecular size fractions (< 1000 MW, < 10 000 MW, < 100 000 MW and <0.1 μm) of dissolved organic matter (DOM) on the growth of bacteria, algae and protozoa from a highly humic lake were investigated. DOM from catchment drainage water as well as from the lake consisted mostly (59–63%) of high molecular weight (HMW) compounds (> 10 000 MW). With excess inorganic nutrients, the growth rate and yield of bacteria were almost identical in all size fractions. However, in < 1000 MW fractions and with glucose added, a longer lag phase occurred. Without added nutrients both the growth rates and biomasses of bacteria decreased towards the smaller size fractions and the percentage of dissolved organic carbon (DOC) used during the experiment and the growth efficiency of bacteria were lower than with excess nutrients. The growth efficiency of bacteria was estimated to vary between 3–66% in different MW fractions, largely depending on the nutrient concentrations, but the highest growth efficiencies were observed in HMW fractions and with glucose. The growth of algae was clearly lowest in the < 1000 MW fraction. In dim light no net growth of algae could be found. In contrast, added nutrients substantially enhanced algal growth and in deionized water with glucose, algae achieved almost the same growth rate and biomass as in higher MW fractions of DOM. The results suggested that bacteria and some algae were favoured by DOM, but protozoans seemed to benefit only indirectly, through bacterial grazing. The utilization of DOM by bacteria and algae was strongly affected by the availability of phosphorus and nitrogen.     

Formation and distribution of free amino acids in Cunninghamella elegans mycelium during growth on media with glucose and dodecane]

The growth, assimilation of organic nitrogen compounds, and production and distribution of free amino acids were studied during the growth of Cunninghamella elegans (--) 1204 on the defined medium 12 containing glucose and dodecane and without an additional carbon source (control). The culture utilized all leucine, glycine, and asparagine after three days of the growth, irrespective of the source of carbon in the medium. Glutamic acid was assimilated only in the presence of glucose. Some 15 free amino acids and 2 amides were detected in the cells, and also some correlations in the composition of the pool in the presence of different sources of carbon in the medium. The substitution of dodecane for glucose decelerates metabolism of free intracellular amino acids and stimulates their accumulation in the cells, especially if there is a deficiency of carbon sources: the overall content of amino acids and amides on the medium with glucose is from 2.2 to 4.1 mg/g; with dodecane, from 9.8 to 16.5 mg/g; in the control, from 16.4 to 25.8 mg/g.     

An annual cycle of dimethylsulfoniopropionate-sulfur and leucine assimilating bacterioplankton in the coastal NW Mediterranean

The contribution of major phylogenetic groups to heterotrophic bacteria assimilating sulfur from dissolved dimethylsulfoniopropionate (DMSP) and assimilating leucine was analysed in surface seawaters from Blanes Bay (NW Mediterranean) over an annual study between March 2003 and April 2004. The percentage of bacteria assimilating DMSP-S showed a strong seasonal pattern, with a steady increase from winter (8 +/- 5%) to summer (23 +/- 3%). The same seasonal pattern was observed for the rate of DMSP-S assimilation. The annual average percentage of DMSP-S-assimilating bacteria (16 +/- 8%) was lower than the corresponding percentage of leucine-assimilating cells (35 +/- 16%), suggesting that not all bacteria synthesizing protein incorporated DMSP-S. Smaller differences between both percentages were recorded in summer. Members of the Alphaproteobacteria (Roseobacter and SAR11) and Gammaproteobacteria groups accounted for most of bacterial DMSP-S-assimilating cells over the year. All major bacterial groups showed an increase of the percentage of cells assimilating DMSP-S during summer, and contributed to the increase of the DMSP-S assimilation rate in this period. In these primarily P-limited waters, enrichment with P + DMSP resulted in a stimulation of bacterial heterotrophic production comparable to, or higher than, that with P + glucose in summer, while during the rest of the year P + glucose induced a stronger response. This suggested that DMSP was more important a S and C source for bacteria in the warm stratified season. Overall, our results suggest that DMSP-S assimilation is controlled by the contribution of DMSP to S (and C) sources rather than by the phylogenetic composition of the bacterioplankton.     

Limitation of bacterial growth by dissolved organic matter and iron in the Southern ocean

The importance of resource limitation in controlling bacterial growth in the high-nutrient, low-chlorophyll (HNLC) region of the Southern Ocean was experimentally determined during February and March 1998. Organic- and inorganic-nutrient enrichment experiments were performed between 42 degrees S and 55 degrees S along 141 degrees E. Bacterial abundance, mean cell volume, and [(3)H]thymidine and [(3)H]leucine incorporation were measured during 4- to 5-day incubations. Bacterial biomass, production, and rates of growth all responded to organic enrichments in three of the four experiments. These results indicate that bacterial growth was constrained primarily by the availability of dissolved organic matter. Bacterial growth in the subtropical front, subantarctic zone, and subantarctic front responded most favorably to additions of dissolved free amino acids or glucose plus ammonium. Bacterial growth in these regions may be limited by input of both organic matter and reduced nitrogen. Unlike similar experimental results in other HNLC regions (subarctic and equatorial Pacific), growth stimulation of bacteria in the Southern Ocean resulted in significant biomass accumulation, apparently by stimulating bacterial growth in excess of removal processes. Bacterial growth was relatively unchanged by additions of iron alone; however, additions of glucose plus iron resulted in substantial increases in rates of bacterial growth and biomass accumulation. These results imply that bacterial growth efficiency and nitrogen utilization may be partly constrained by iron availability in the HNLC Southern Ocean.     

Metabolic fluxes in Schizosaccharomyces pombe grown on glucose and mixtures of glycerol and acetate

Growth on glycerol has already been a topic of research for several yeast species, and recent publications deal with the regulatory mechanisms of glycerol assimilation by the fission yeast Schizosaccharomyces pombe. We investigated glycerol metabolism of S. pombe from a physiological point of view, characterizing growth and metabolism on a mixture of glycerol and acetate and comparing it to growth on glucose under respirative growth conditions in chemostat experiments. On glycerol/acetate mixtures, the cells grew with a maximum specific growth rate of 0.11 h^?1 where 46 % of the carbon was channeled into biomass and the key fermentation product ethanol was not detectable. ¹³C-assisted metabolic flux analysis resolved substrate distributions through central carbon metabolism, proving that glycerol is used as a precursor for glycolysis, gluconeogenesis, and the pentose phosphate pathway, while acetate enters the tricarboxylic acid cycle via acetyl-CoA. Considering compartmentalization between cytosol and mitochondria in the metabolic model, we found compartmentalization of biosynthesis for the amino acids aspartate and leucine. Balancing of redox cofactors revealed an abundant production of cytosolic NADPH that must be finally regenerated via the respiratory chain shown by the simulated and measured CO₂ production and oxygen consumption rates which were in good agreement.     

Competition for dissolved glucose between meiobenthos and sediment microbes

Meiobenthos, small invertebrates inhabiting the surface layers of marine sediments, can absorb dissolved organic matter (DOM). Experiments were performed to test if meiobenthos can compete with sediment microbes for uptake of small amounts of [¹⁴C]glucose. Meiofaunal glucose uptake rates were measured by themselves and in the presence of sediment microbes. Glucose uptake by meiofauna was not inhibited by the presence of bacteria, nor did it appear that bacterial uptake was inhibited by meiofauna. Thus, there was no direct or interference competition. Uptake rates by 1 cm³ of sediment (bacteria) were four orders of magnitude greater than those of individual meiofauna, but on a biomass specific basis, meiofaunal uptake was in the same range if not higher than that of sediment bacteria. Thus, the potential for indirect or resource competition exists. Since bacterial biomass dominated the system studied, uptake of glucose was dominated by bacteria. The results support the hypothesis that in natural sediments, where the biomass of bacteria is higher than that of meiofauna, heterotrophic uptake is primarily a microbial process. However, resource competition between meiofauna and bacteria for DOM in sediments probably exists where bacterial biomass is low relative to meiofaunal biomass.     

Metabolic analysis of the synthesis of high levels of intracellular human SOD in Saccharomyces cerevisiae rhSOD 2060 411 SGA122

The synthesis of human superoxide dismutase (SOD) in batch cultures of a Saccharomyces cerevisiae strain using a glucose-limited minimal medium was studied through metabolic flux analysis. A stoichiometric model was built, which included 78 reactions, according to metabolic pathways operative in these strains during respirofermentative and oxidative metabolism. It allowed calculation of the distribution of metabolic fluxes during diauxic growth on glucose and ethanol. Fermentation profiles and metabolic fluxes were analyzed at different phases of diauxic growth for the recombinant strain (P+) and for its wild type (P-). The synthesis of SOD by the strain P+ resulted in a decrease in specific growth rate of 34 and 54% (growth on glucose and ethanol respectively) in comparison to the wild type. Both strains exhibited similar flux of glucose consumption and ethanol synthesis but important differences in carbon distribution with biomass/substrate yields and ATP production 50% higher in P-. A higher contribution of fermentative metabolism, with 64% of the energy produced at the phosphorylation level, was observed during SOD production. The flux of precursors to amino acids and nucleotides was higher in the recombinant strain, in agreement with the higher total RNA and protein levels. Lower specific growth rates in strain P+ appear to be related to the decrease in the rate of synthesis of nonrecombinant protein, as well as a decrease in the activities of the pentose phosphate (PP) pathway and TCA cycle. A very different way of entry into the stationary phase was observed for each strain: in the wild-type strain most metabolic fluxes decreased and fluxes related to energy reserve synthesis increased, while in the P+ strain the flux of 22 reactions (including PP pathway and amino acids biosynthesis) related to SOD production increased their fluxes. Changes in SOD production rates at different physiological states appear to be related to the differences in building blocks availability between respirofermentative and oxidative metabolism. Using the present expression system, ideal conditions for SOD synthesis are represented by either active growth during respirofermentative metabolism or transition from a growing to a nongrowing state. An increase in SOD flux could be achieved using an expression system nonassociated to growth and potentially eliminating part of the metabolic burden.     

Effect of amino acid supplement on cell yield and gene product in Escherichia coli harboring plasmid

Escherichia coli harboring a recombinant plasmid was cultivated in fed-batch culture to enhance production of a gene product. Expression of the leucine gene from Thermus thermophilus in the recombinant plasmid was examined by the assay of beta-isopropylmalate dehydrogenase activity at 75 degrees C. When E. coli was cultivated in medium without leucine, biomass concentration reached 15 g/L and the specific activity became 0.082 U/mg protein. When leucine was fed in the medium throughout cultivation, although biomass concentration reached 63 g/L, the specific activity decreased to 0.016 U/mg protein. When E. coli was cultivated in medium containing 1 g leucine/L, the specific activity remained virtually constant (about 0.13 U/mg protein) and biomass concentration reached 32 g dry cells/L. In these cultivations, growth yields of several amino acids and glucose were examined. When leucine was not added to the medium, growth yields except for histidine were lowest. When leucine was fed throughout the cultivation, growth yields of glucose and tryptophan were highest. The pH-stat was useful for feeding amino acids.     

Contribution of Bacterial Cell Wall Components to DOM in Alkaline,Hypersaline Mono Lake,California

Soda lakes are often characterized by high densities of prokaryotes and high concentrations of dissolved organic carbon. Since bacterial cell walls are less degradable than most other cell constituents, accumulation of cell wall material may occur in these lakes and contribute to the DOM pool, but composition of DOM in soda lakes has rarely been examined. Here we report concentrations of DOM components likely originating from bacterial cell walls, including D amino acids, glucosamine (GluA) and muramic acid (MurA), in depth profiles of stratified, alkaline, hypersaline Mono Lake, CA. Concentrations of cell wall components were related to total pools of dissolved free and combined amino acids (DFAA and DCAA), and bacterial density and production. In the free pool, total DFAA ranged from 50 to 3250 nM and typically increased with depth, while GluA (5 to 140 nM) and MurA (< 0.5 nM and only detected in 2005) fluctuated with depth. In the combined pool, DCAA varied between 5000 and 15000 nM and did not show clear depth-related trends. GluA ranged from 1000 to 5000 nM and tended to increase in the hypolimnion, while MurA varied between 25 and 75 nM. Free D isomers in the DFAA pool either made up < 13% (Asp and Ser) or varied from 10 to 57% (Glu and Ala). In the combined pool, D isomers of Asp, Glu, Ser and Ala made up 24-48% of these DCAA and typically showed minor changes with depth. In 2005, lysozyme activity had highest rates in the surface and correlated negatively with most D isomers among the combined amino acids. Our observations demonstrate that the pool of dissolved combined amino compounds in the lake was about 5-fold higher than in other eutrophic lakes and that a substantial portion of these amino compounds originated from bacterial cell walls.     

Temperature affects the response of heterotrophic bacteria and mixotrophic algae to enhanced concentrations of soil extract

To investigate the consequences of increased temperature and enhanced input of dissolved organic matter (DOM) into lakes for heterotrophicic bacteria and for mixotrophic algae which use DOM in addition to photosynthesis, the hypotheses were tested whether (1) both bacteria and mixotrophic algae benefit from increased input of DOM, or (2) increased DOM input enhances bacterial biomass and thereby decreases algal biomass. Growth experiments in batch cultures, exudation measurements, and competition experiments in chemostats were performed at two temperature levels. Increased temperature stimulated the autotrophic growth rate of Chlorella protothecoides. Bacteria and Chlorella increased their heterotrophic growth rates at higher DOM concentration at lower temperature whereas enhanced DOM concentration hardly stimulated their growth at higher temperature. In chemostats, enhanced input of soil extract increased both bacterial and algal biomass at lower temperature whereas bacterial biomass increased only slightly and algal biomass decreased at higher temperature. Thus, the temperature determines the response of microorganisms to enhanced DOM concentration.