Extracellular dissolved organic carbon released from phytoplankton as a source of carbon for heterotrophic bacteria in lakes of different humic content

The quantitative importance of photosynthetically produced dissolved organic carbon (PDOC) released from phytoplankton as a source of carbon for pelagic, heterotrophic bacteria was investigated in four temperate Swedish lakes, of which two had low (≈20 mg Pt 1⁻¹), and two moderately high (60–80 mg Pt 1⁻¹) humic content. The bacterial assimilation of PDOC was estimated with the ¹⁴C method, and the total production of the heterotrophic bacteria was estimated with the [3H]thymidine incorporation method. The release of PDOC from natural communities of phytoplankton was not restricted to periods of photosynthesis, but often continued during periods of darkness. Heterotrophic bacteria often assimilated the labile components of the PDOC at high rates (up to 73% of the released PDOC was assimilated during the incubation in our experiments). The contribution of PDOC to bacterial production exhibited large within-lake seasonal variations, but PDOC was at certain times, both in humic and non-humic lakes, a quantitatively very important carbon source for the heterotrophic bacteria. Under periods of comparatively low primary production, heterotrophic bacteria in humic lakes appear to utilize allochthonous, humic substances as a substrate.     

Excretion of photosynthetically fixed organic carbon by metalimnetic phytoplankton

The effects of light intensity, oxygen concentration, and pH on the rates of photosynthesis and net excretion by metalimnetic phytoplankton populations of Little Crooked Lake, Indiana, were studied. Photosynthetic rates increased from 1.42 to 3.14 mg C·mg^–1 chlorophylla·hour^–1 within a range of light intensities from 65 to 150E·m^–2·sec^–1, whereas net excretion remained constant at 0.05 mg C·mg^–1 chlorophylla·hour^–1. Bacteria assimilated approximately 50% of the carbon released by the phytoplankton under these conditions. Excreted carbon (organic compounds either assimilated by bacteria or dissolved in the lake water) was produced by phytoplankton at rates of 0.02–0.15 mg C·mg^–1 chlorophylla·hour^–1. These rates were 6%–13% of the photosynthetic rates of the phytoplankton. Both total excretion of carbon and bacterial assimilation of excreted carbon increased at high light intensities whereas net excretion remained fairly constant. Elevated oxygen concentrations in samples incubated at 150E· m^–2·sec^–1 decreased rates of both photosynthesis and net excretion. The photosynthetic rate increased from 3.0 to 5.0 mg C·mg^–1 chlorophylla· hour^–1 as the pH was raised from 7.5 to 8.8. Net excretion within this range decreased slightly. Calculation of total primary production using a numerical model showed that whereas 225.8 g C·m^–2 was photosynthetically fixed between 12 May and 24 August 1982, a maximum of about 9.3 g C·m^–2 was released extracellularly.     

Characterization of extracellular particles released from continuous cell cultures derived from human leukemia.

Extracellular particles, with a density of 1.18-1.22 g/cm3 in sucrose, were detected in the culture medium of a continuous cell line (JIII) derived from a patient with monocytic leukemia. These particles contained RNA, DNA, and a DNA polymerase. They synthesized DNA with endogenous templates and primers and also used exogenous DNA but not poly(rC) oligo(dG) as a template. Pretreatment with Nonidet P-40 stimulated DNA polymerase activity while treatment with ribonuclease partially inhibited the enzyme activity. Fluorescent antibodies made to the particles stained both JIII and Z-597 cells derived from human leukemias but not other types of human or nonhuman cultured cells tested. The particles do not appear to be oncornaviruses but may be a particulate antigen associated with malignant cells of hemopoietic and lymphoid origin.     

Molecular weight fractionation of dissolved organic matter (DOM) released by phytoplankton

The application of an ultrafiltration procedure for fractionation of molecular weight of dissolved organic matter (DOM) extracellularly released by phytoplankton is described. Seven ultrafiltration membranes Diaflo (Amicon Corp. USA), with range 500--300,000 molecular weight (MW), were used for separation of different molecular weight compounds released by phytoplankton in Rhode River estuary of Chesapeake Bay, and their composition was determined. Percentage of extracellular release of DOM by phytoplankton varied from 3.92--68.07% of total carbon fixed in photosynthesis. The composition of algal extracellular products varied with different phytoplankton populations. However, two fractions of molecular weight compounds dominated in the composition of DOM released, i.e. a low molecular weight fraction of less than 1,000 MW and the fraction between 10,000--30,000 MW. The ultrafiltration procedure is effective for studying the composition of DOM released by phytoplankton in natural waters.     

Copper binding ability of the extracellular organic matter released by Skeletonema costatum

Abstract

The copper complexing ability of the exudates produced during the exponential growth phase by Skeletonema costatum has been investigated by a ligand-competition technique involving copper sorption onto C-18 Sep-Pak cartridges. Two ligands with different affinity for copper were required for the best fit of the copper complexation data in seawater with and without exudates: a strong ligand with a log K close to 13 and a weaker ligand with a log K close to 9. The culture increased both L₁ and L₂ ligand concentrations, already present in seawater, by a factor close to 4 after the first 72 hours of growth. The presence of class 1 stronger ligands in copper binding organics produced by the diatom is discussed in relation to natural copper speciation in the sea.     

Rates of microbial degradation of dissolved organic carbon from phytoplankton cultures

Dissolved organic carbon (DOC) decay was measured for samplesfrom cultures of the diatoms Chaetoceros gracilis and Phaeodactylumtricornutum, the flagellate Isochrysis galbana, the dinoflagellateAlexandrium tamarense, and a natural algal assemblage from theNorthwest Arm, Nova Scotia, Canada, by a high-temperature catalyticoxidation (HTCO) method. Decay rate constants were determinedusing first-order reaction kinetics in the multi-G model ofBerner (In Early Diagenesis, a Theoretical Approach, PrincetonUniversity Press, 1980). Decay rates as high as 0.37 day^–1wereobtained, which demonstrated that DOC released by phytoplanktonmight be highly labile to bacterial utilization and could bedegraded significantly within hours. Decay rates for most speciestested followed much the same pattern, with K₀₁ values around0.3–0.4, K₀₂ values around 0.03, and K₀₃ and K₀₄ valuesaround 10^–3 day^–1. DOC released by the senescentcells of A.tamarense was found to be essentially bacteria resistant,in contrast to that of the other species tested. The decay ofDOC was directly temperature dependent over the 10–20°Crange. Six methods for DOC preservation were tested. Acidificationwith HCl and refrigerated storage was demonstrated to be themost convenient and practical method. This method can be usedfor both short- and long-term preservation of DOC samples containinghighly labile organic compounds.     

Production of new organic carbon and its distribution between autotrophic picoplankton, bacteria, extracellular organic carbon and phytoplankton in an upland lake

• 1 Picoplankton community production (0.2–2μm) was investigated over 3 months, June-September 1991, in Llyn Padarn, a mesotrophic upland lake in north Wales.
• 2 The picoplankton was differentiated into autotrophic algae (<1–3μm) and heterotrophic bacteria (<0.2–1 μm) using differential filtration through a 1 μm pore size Nuclepore filter.
• 3 Efficient separation of these distinct metabolic constituents of picoplankton was obtained. A good correlation (r= 0.81, P < 0.001) was found between physical separation of bacterial and picoalgal cells from fluorescence microscopy and the distribution of heterotrophic metabolic activity between different cell size fractions measured by uptake of ¹⁴C-glucose.
• 4 Picoplankton community production was differentiated into the ‘absolute’ autotrophic production by picoalgae, corrected for overestimation due to retention of bacteria with the picoalgae, and the heterotrophic component, bacterial uptake of ‘extracellular organic carbon’ (EOC), derived from the entire phytoplankton community.
• 5 The heterotrophic contribution to picoplankton community production ranged from 88 to 1%, mean value 55% of total. Autotrophic picoplankton production was dominant in June and July, but in August and September heterotrophic uptake of EOC was the major input to picoplankton community production.
• 6 During the 3 months, the mean contributions to plankton production were autotrophic picoplankton 10.3%, heterotrophic bacterial uptake of EOC 9.7%, EOC in lake water 11.6% and phytoplankton (>3μm) 68.3%.
• 7 Bacteria accounted for about half the picopfankton community production via uptake of EOC. Thus although autotrophic picoplankton were ubiquitous, it is likely that their contribution via primary production to the carbon balance of planktonic environments has been overestimated in previous studies.

     

Bacterial response to extracellular dissolved organic carbon released from healthy and senescent Fragilaria crotonensis (Bacillariophyceae) in experimental systems

Responses of bacteria to dissolved organic carbon (DOC) released from healthy and senescent Fragilaria crotonensis (Bacillariophyceae) were examined in experimental systems. The alga released DOC actively, although the concentration fluctuated greatly in both the axenic (the alga alone) and the mixed (the alga plus the enriched bacteria) cultures. In the control (the bacteria alone) cultures, both DOC concentration and bacterial density were low and almost constant throughout the experiment: 5.0 mg C 1^–1 and 1.1 × 10⁵ cells ml^–1, respectively. In the mixed cultures, bacterial growth was negligible during the exponential growth phase of the alga, but rapid proliferation of the bacteria occurred after the onset of the stationary growth phase. As the bacterial population grew, the density of senescent algal cells also increased. When the bacteria were fed on the DOC from healthy algae, their growth rate was relatively low (0.44 d^–1), but the maximum cell density was high (6.4 × 10⁵ cells ml^–1). Conversely, when the bacteria fed on the DOC of senescent algae, they grew at a relatively high rate (0.51 d^–1), but the maximum cell density was low (2.8 × 10⁵ cells ml^–1). These results suggest that DOCs released from dominant phytoplankton species in different physiological states affect the biomass and activity of bacteria.     

Biodegradation of photosynthetically produced extracellular organic carbon from intertidal benthic algae

14C-labeled extracellular products of a natural microphytobenthic community and two species of benthic diatoms (Nitzschia hybridaeformis and Amphora coffeaeformis) were fractionated into extracellular dissolved organic carbon (14C-EDOC), organic carbon extracted with EDTA (14C-EDTA-extractable OC) and extracellular polymeric substances (14C-EPS). The biodegradation of this labeled extracellular organic carbon by bacteria in sediments was examined to determine the processes of enzymatic degradation of photosynthetically-produced extracellular organic carbon from microphytobenthos in an intertidal flat ecosystem. In addition, primary production as well as extracellular enzyme activities (beta- and alpha-glucosidase) were measured to evaluate the possible relationship between organic carbon production and microbiological degradation at the Isshiki intertidal flat in Mikawa Bay, Japan. With all three 14C-fractions extracted from a natural microphytobenthic assemblage and two species of benthic diatoms, more than 50% of the added substrates were mineralized within 24 h by the bacterial community in sediments. At that time, the percentage of high-molecular-weight compounds (>5 K MW) to total MW compounds of 14C-EDTA-extractable OC and 14C-EPS fractions decreased within 24 h from 50.9 to 6.6% and 74.5 to 11.1%, respectively. In situ, beta- and alpha-glucosidase activity in sediment was higher than in the seawater column (at a depth of 1 m), though the photosynthetic production of microphytobenthos was equal to that of phytoplankton. Based on our previous studies that microphytobenthos produced much more extracellular products than phytoplankton, it is assumed from these results that carbon flowing into the microbial loop through the mediation of enzymatic degradation of extracellular products in a benthic system exceeds that in the overlying water column.     

Mutualistic relationships between phytoplankton and bacteria caused by carbon excretion from phytoplankton

For the competition system of phytoplankton and bacteria through inorganic phosphorus, our mathematical model showed that mutualistic relationships between them could occur due to production and consumption of extracellular organic carbon by phytoplankton and bacteria. In our model, phytoplankton are limited in their growth by light and phosphorus, and bacteria are limited in their growth by phosphorus and carbon released from phytoplankton. We adopted permanence as a criterion of the coexistence in mathematical analysis, and led necessary conditions of permanence in the model. Under these coexistence conditions, we estimated the strength of total effects of interactions between phytoplankton and bacteria at the steady state by press perturbation method. The results of this estimation indicated the mutualistic interactions between phytoplankton and bacteria. This suggests that mutualistic situation could occur due to the introduction of carbon flow from phytoplankton to bacteria, even if phytoplankton and bacteria compete with each other through common resource, inorganic phosphorus.     

Dissolved organic carbon released by zooplankton grazing activity-a high-quality substrate pool for bacteria

Experiments were designed to investigate whether processes relatedto zooplankton feeding have a positive effect on bacterial growth.Bacterial abundance and [³H]thymidine incorporation rates werefollowed in grazer-free batch cultures originally containingeither Scenedesmus quadricauda or Rhodomonas lacustris as foodsources, and Daphnia cucullata or Eudiaptomus graciloides asgrazers. Compared with controls lacking either animals or algae,a significantly higher bacterial abundance and productivityoccurred in cultures which contained both phyto- and zoo-plankton.The same experimental methodology was tested during the declineof a diatom spring bloom in a eutrophic, temperate lake. A significantincrease in bacterial biomass was observed due to the grazingactivity of in situ mesozooplankters during the clear-waterphase. Our results demonstrated that the dissolved carbon pathwayfrom mesozooplankton to bacteria averaged 57% (26–78%)of the algal carbon filtered from suspension.     

Improving estimates of maximal organic carbon stabilization by fine soil particles

Organic carbon (C) associated with fine soil particles (<20 μm) is relatively stable and accounts for a large proportion of total soil organic C (SOC). The soil C saturation concept proposes a maximal amount of SOC that can be stabilized in the fine soil fraction, and the soil C saturation deficit (i.e., the difference between current SOC and the maximal amount) is presumed to affect the capacity, magnitude, and rate of SOC storage. In this study, we argue that predictions using current models underestimate maximal organic C stabilization of fine soil particles due to fundamental limitations of using least-squares linear regression. The objective was to improve predictions of maximal organic C stabilization by using two alternative approaches; one mechanistic, based on organic C loadings, and one statistical, based on boundary line analysis. We collected 342 data points on the organic C content of fine soil particles, fine particle mass proportions in bulk soil, dominant soil mineral types, and land use types from 32 studies. Predictions of maximal organic C stabilization using linear regression models are questionable because of the use of data from soils that may not be saturated in SOC and because of the nature of regression itself, resulting in a high proportion of presumed over-saturated samples. Predictions of maximal organic C stabilization using the organic C loading approach fit the data for soils dominated by 2:1 minerals well, but not soils dominated by 1:1 minerals; suggesting that the use of a single value for specific surface area, and therefore a single organic C loading, to represent a large dataset is problematic. In boundary line analysis, only data representing soils having reached the maximal amount (upper tenth percentile) were used. The boundary line analysis estimate of maximal organic C stabilization (78 ± 4 g C kg^?1 fraction) was more than double the estimate by the linear regression approach (33 ± 1 g C kg^?1 fraction). These results show that linear regression models do not adequately predict maximal organic C stabilization. Soil properties associated with soil mineralogy, such as specific surface area and organic C loading, should be incorporated to generate more mechanistic models for predicting soil C saturation, but in their absence, statistical models should represent the upper envelope rather than the average value.     

RNA-directed DNA polymerase from particles released by normal goose cells.

Cells from a goose embryo were shown to release particle-associated RNA-directed DNA polymerase and RNase H activities that required the presence of Nonidet P-40 for detection. The particles were not infectious and did not have endogenous DNA synthesis. The goose particle DNA polymerase was related to the DNA polymerase of spleen necrosis virus with respect to size and was inhibited by immunoglobulin G to spleen necrosis virus DNA polymerase. However, goose cells producing DNA polymerase-containing particles did not contain reticuloendotheliosis virus-related nucleotide sequences in their DNA.     

Plankton photosynthesis, extracellular release and bacterial utilization of released dissolved organic carbon (RDOC) in lakes of different trophy

The major objective of these studies was to assess the extent of the flow of released dissolved organic carbon (RDOC) from phytoplankton to heterotrophic bacteria. The second aim was to test Nalewajko and Schindler's (1976) hypothesis that the percentage of extracellular release (PER) of RDOC by phytoplankton is higher in oligotrophic than eutrophic waters. The studies on the relationship between the bacterial assimilation of RDOC and productivity of phytoplankton in the lakes of different trophy have shown a direct correlation. It was observed that higher utilization rate of phytoplankton RDOC occurred in lakes with higher primary production. The inverse relationship between productivity of lakes, i.e. trophy of waters, and amount of RDOC or PER is closely dependent on the heterotrophic activity of bacteria in waters.     

The use of antibiotics to reduce bacterioplankton uptake of phytoplankton extracellular organic carbon (EOC) in the Potomac River estuary

Phytoplankton production and accumulation of extracellular organic carbon (EOC) was tracked during diel intervals in microcosms by inhibiting bacterioplankton assimilation of EOC with streptomycin and kanamycin. Bacterioplankton production (³H-thymidine incorporation) and metabolism (¹⁴C-glucose incorporation) were monitored in samples collected from the Potomac River estuary to determine the effect of the antibiotics. Particulate (i.e., raw water) primary production and EOC (i.e., water passing through 1.0 μm glass fiber filter) production rates were monitored to determine the impact of antibiotics on phytoplankton. In preliminary experiments, neither streptomycin nor kanamycin alone significantly inhibited bacterioplankton activity compared to controls, but when both were present secondary production and metabolism were reduced up to 90%, and remained as such for 45 h. During field evaluations using a streptomycin and kanamycin mixture (50 μM each) particulate primary production and EOC production were not statistically different in control and antibiotic treated samples indicating that the antibiotics did not negatively influence phytoplankton production rates. In the presence of antibiotics dissolved free amino acids (DFAA) and, to a lesser extent, monosaccharides were significantly elevated compared to controls. This study demonstrates that streptomycin and kanamycin are capable of inhibiting bacterioplankton metabolism and uptake of dissolved organic carbon (DOC) in the samples tested so that the contribution of EOC to the DOC pool and to bacterioplankton metabolism could be measured and assessed.     

Colloidal organic carbon and phytoplankton speciation during a coastal bloom

During the 1995 spring bloom in Bedford Basin, Nova Scotia,dissolved organic carbon (DOC) in the photic zone was separatedinto low-molecular-weight and colloidal size fractions by cross-flowultrafiltration. DOC, colloidal organic carbon (COC) and low-molecular-weightorganic carbon (LOC) were then analyzed by high-temperaturecatalytic oxidation. COC (associated with the production ofphytoplankton exudates) did not follow the concentration ofbulk chlorophyll a (chi a) or the total number of phytoplanktoncells. Instead, surface-active COC (that adhered to the ultrafiltrationsystem) was produced early during the bloom when the diatomSkeletonema costatum was at a maximum. Later on, as the bloombegan to decline, less surface-active COC (that remained largelyfree of the ultrafiltration surfaces) was produced and was associatedmore with variations in cell number of Chaetoceros socialis,the predominant diatom for most of the bloom. These resultssuggest that chl a or the total number of phytoplankton cellsmay not be reliable indices of the production of COC. On thecontrary, the results suggest that relatively high COC concentrationswere associated with specific diatom species. In addition, themaximum amount of COC was associated during the early stagesof the bloom with a diatom (S.costatum) that was a small fraction(<5.6%) of total phytoplankton cell number. This finding,that relatively large amounts of colloids were associated withexopolymer production during the onset and development of abloom, does not agree with reports suggesting that the productionof exopolymers by diatoms is primarily an end-of-bloom occurrence.     

Assessment of oxidative DNA damage formation by organic complex mixtures from airborne particles PM(10)

The free radical generating activity of airborne particulate matter (PM₁₀) has been proposed as a primary mechanism in biological activity of ambient air pollution. In an effort to determine the impact of the complex mixtures of extractable organic matter (EOM) from airborne particles on oxidative damage to DNA, the level of 8-oxo-2′-deoxyguanosine (8-oxodG), the most prevalent and stable oxidative lesion, was measured in the human metabolically competent cell line Hep G2. Cultured cells were exposed to equivalent EOM concentrations (5–150 μg/ml) and oxidative DNA damage was analyzed using a modified single cell gel electrophoresis (SCGE), which involves the incubation of whole cell DNA with repair specific DNA endonuclease, which cleaves oxidized DNA at the sites of 8-oxodG. EOMs were extracted from PM₁₀ collected daily (24 h intervals) in three European cities: Prague (Czech Republic, two monitoring sites, Libuš and Smíchov), Košice (Slovak Republic) and Sofia (Bulgaria) during 3-month sampling periods in the winter and summer seasons. No substantial time- and dose-dependent increase of oxidative DNA lesions was detected in EOM-treated cells with the exception of the EOM collected at the monitoring site Košice, summer sampling. In this case, 2 h cell exposure to EOM resulted in a slight but significant increase of oxidative DNA damage at three from total of six concentrations. The mean 8-oxodG values at these concentrations ranged from 15.3 to 26.1 per 10⁶ nucleotides with a value 3.5 per 10⁶ nucleotides in untreated cells. B[a]P, the positive control, induced a variable but insignificant increase of oxidative DNA damage in Hep G2 cell (approximately 1.6-fold increase over control value).

Based on these data we believe that EOM samples extracted from airborne particle PM₁₀ play probably only a marginal role in oxidative stress generation and oxidative lesion formation to DNA. However, adsorbed organic compounds can undergo various interactions (additive or synergistic) with other PM components or physical factors (UV-A radiation) and in this way they might enhance/multiply the adverse health effects of air pollution.