Modelling interactions between phytoplankton and bacteria under nutrient-regenerating conditions

The interactions of phytoplankton and bacteria in a nitrogen-limitedsteady-state system with an organic nitrogen compound or ammoniumas the sole nitrogen source were modelled. The effects of variousalgal excretion rates and two different mathematical representationsof excretion were examined. The model predicted that higherexcretion elevated the bacterial steady-state biomass, and loweredthe algal biomass. Bacterial respiration, which directly determinednitrogen regeneration, had an important effect on the system.The bacterial growth yield in the model was mainly a functionof the growth rate, and not of the nitrogen:carbon ratio ofthe substrate. In one version of the model, where the excretionof organic carbon increased with decreasing growth rate, themodel started to oscillate when the multiplication product ofmaximum specific excretion of excreted organic carbon (EOC)and the bacterial yield on EOC exceeded the dilution rate, irrespectiveof the form of nitrogen (ammonium or dissolved organic nitrogen)in the medium. The model results were compared with chemostatexperiments with the alga Emiliania huxleyi and a bacterialisolate in pure and mixed culture at two different dilutionrates. The carbon and nitrogen biomass of the bacteria was {smalltilde}1.5 times higher in mixed culture than in pure culture.In the experiments with low dilution rate, the recovery of nitrogenin the form of biomass, ammonium or amino acids was low, suggestingthe excretion by the algae of a refractory nitrogen-containingproduct which the bacteria could not use.     

Utilization of Dissolved Nitrogen by Heterotrophic Bacterioplankton: a Comparison of Three Ecosystems

The contributions of different organic and inorganic nitrogen and organic carbon sources to heterotrophic bacterioplankton in batch cultures of oceanic, estuarine, and eutrophic riverine environments were compared. The importance of the studied compounds was surprisingly similar among the three ecosystems. Dissolved combined amino acids (DCAA) were most significant, sustaining from 10 to 45% of the bacterial carbon demands and from 42 to 112% of the bacterial nitrogen demands. Dissolved free amino acids (DFAA) supplied 2 to 7% of the carbon and 6 to 24% of the nitrogen incorporated into the bacterial biomass, while dissolved DNA (D-DNA) sustained less than 5 and 12% of the carbon and nitrogen requirements, respectively. Ammonium was the second most important source of nitrogen, meeting from 13 to 45% of the bacterial demand in the oceanic and estuarine cultures and up to 270% of the demand in riverine cultures. Nitrate was taken up in the oceanic cultures (uptake equaled up to 46% of the nitrogen demand) but was released in the two others. Assimilation of DCAA, DFAA, and D-DNA combined supplied 43% of the carbon demand of the bacteria in the oceanic cultures, while approximately 25% of the carbon requirements were met by the three substrates at the two other sites. Assimilation of nitrogen from DCAA, DFAA, D-DNA, NH₄⁺, and NO₃^-, on the other hand, exceeded production of particulate organic nitrogen in one culture at 27 h and in all cultures over the entire incubation period (50 h). These results suggest that the studied nutrient sources may fully support the nitrogen needs but only partially support the carbon needs of microbial communities of geographically different ecosystems. Furthermore, a comparison of the initial concentrations of the different substrates indicated that relative pool sizes of the substrates seemed to influence which substrates were primarily being utilized by the bacteria.     

Effects of PCB on interspecific competition in natural and gnotobiotic phytoplankton communities in continuous and batch cultures

The toxicity of polychlorinated biphenyls (PCB) to the diatomThalassiosira pseudonana (formerlyCyclotella nana), grown in pure and mixed cultures, was greatest when in competition with other species. Continuous cultures were superior to batch cultures for studying competitive interactions, and PCB caused greater alteration of species composition in continuous cultures than it did in batch cultures. Natural phytoplankton communities from Vineyard Sound, maintained in continuous culture, responded to PCB stress the same as did gnotobiotic communities, withT. pseudonana showing similar responses in both communities. A PCB concentration of 0.1 μg/liter (0.1 part per billion), a level not uncommon in natural waters, did not affect algal growth in pure cultures but caused substantial disruption of continuous culture communities. The possible impact of PCB pollution on natural phytoplankton communities is discussed. Contribution No. 3181 from the Woods Hole Oceanographic Institution.     

MOLECULAR CHARACTERIZATION AND ANTIBODY DETECTION OF A NITROGEN‐REGULATED CELL‐SURFACE PROTEIN OF THE COCCOLITHOPHORE EMILIANIA HUXLEYI (PRYMNESIOPHYCEAE)1

Dissolved organic nitrogen (DON) can account for a significant portion of total nitrogen in some aquatic environments, and many species of phytoplankton are able to scavenge nitrogen from this pool especially when inorganic nitrogen is limiting. Emiliania huxleyi (Lohmann) H. W. Hay et H. Mohler is able to use various forms of DON for growth, including several amino acids, purines, and pyrimidines. A cell‐surface protein up‐regulated in the absence of inorganic nitrogen, NRP1, is hypothesized to play a role in the metabolism of one or more of these organic nitrogen forms. Here, the genomic and cDNA sequence of NRP1 is reported. Structural predictions based on the amino acid sequence suggest a pyridoxal‐5′‐phosphate‐dependent enzyme that may have a role in acquiring nitrogen from amino acids. Further evidence for the function of NRP1 is measured in spent media from nitrogen‐limited cultures, which contain NRP1 and have glutaminase and formamidase activity. Field studies using an antibody to NRP1 show that it is expressed in E. huxleyi during bloom conditions in a Norwegian fjord.     

Influence of metazooplankton on interactions of bacteria and phytoplankton in an oligotrophic lake

Commensalism based on organic carbon supplied by phytoplanktonand competition for mineral nutrients are important interactionsbetween bacteria and phytoplankton in oligotrophic clear-watersystems. Both interactions are influenced by zooplankton activity.To examine the relation ship between algae and bacteria in LakeLa Caldera, we studied: the correlations among phyto plankton,bacteria and phosphorus (P) dynamics; the ratio of organic carbonsupplied by algae to organic carbon demand by bacteria; andthe importance of P remineralized by metazooplankton for bothcommunities. Phytoplankton and bacteria had a similar seasonaldynamics, and there was a sig nificant and positive relationshipbetween bacterial abundance and algal biomass (P<0.01). However,the release of organic carbon from phytoplankton was usuallyhigher than the bacterioplankton carbon requirement. P availablevia zooplankton remineralization satisfied between 74 and 316%of the minimum P demands of algae and bacteria. To elucidatewhether zooplankton operate similarly on algae and bacterialgrowth or indirectly influence bacterial growth through phytoplanktonmetab olism, we performed zooplankton manipulation experiments.High zooplankton biomass in these experiments stimulated bothprimary and bacterial production, but release of organic carbonfrom phytoplankton declined. These results suggest a directstimulus of bacterial growth, so algae and bac teria can balancegrazing losses by compensatory growth. Further, the algal decreaseof the organic carbon supply for bacteria could, over time,lead to a change in the algae-bacteria interaction from competitionto commensalism. This reduction in organic carbon excretioncould affect the balance of the competitive interaction.     

Diatom-associated bacteria are required for aggregation of Thalassiosira weissflogii

Aggregation of algae, mainly diatoms, is an important process in marine systems leading to the settling of particulate organic carbon predominantly in the form of marine snow. Exudation products of phytoplankton form transparent exopolymer particles (TEP), which acts as the glue for particle aggregation. Heterotrophic bacteria interacting with phytoplankton may influence TEP formation and phytoplankton aggregation. This bacterial impact has not been explored in detail. We hypothesized that bacteria attaching to Thalassiosira weissflogii might interact in a yet-to-be determined manner, which could impact TEP formation and aggregate abundance. The role of individual T. weissflogii-attaching and free-living new bacterial isolates for TEP production and diatom aggregation was investigated in vitro. T. weissflogii did not aggregate in axenic culture, and striking differences in aggregation dynamics and TEP abundance were observed when diatom cultures were inoculated with either diatom-attaching or free-living bacteria. The data indicated that free-living bacteria might not influence aggregation whereas bacteria attaching to diatom cells may increase aggregate formation. Interestingly, photosynthetically inactivated T. weissflogii cells did not aggregate regardless of the presence of bacteria. Comparison of aggregate formation, TEP production, aggregate sinking velocity and solid hydrated density revealed remarkable differences. Both, photosynthetically active T. weissflogii and specific diatom-attaching bacteria were required for aggregation. It was concluded that interactions between heterotrophic bacteria and diatoms increased aggregate formation and particle sinking and thus may enhance the efficiency of the biological pump.     

The impact of associated bacteria on morphology and physiology of the dinoflagellate Alexandrium tamarense

Despite their potential impact on phytoplankton dynamics and biogeochemical cycles, biological associations between algae and bacteria are still poorly understood. The aim of the present work was to characterize the influence of bacteria on the growth and function of the dinoflagellate Alexandrium tamarense. Axenic microalgal cultures were inoculated with a microbial community and the resulting cultures were monitored over a 15-month period, in order to allow for the establishment of specific algal–bacterial associations. Algal cells maintained in these new mixed cultures first experienced a period of growth inhibition. After several months, algal growth and cell volume increased, and indicators of photosynthetic function also improved. Our results suggest that community assembly processes facilitated the development of mutualistic relationships between A. tamarense cells and bacteria. These interactions had beneficial effects on the alga that may be only partly explained by mixotrophy of A. tamarense cells. The potential role of organic exudates in the establishment of these algal–bacterial associations is discussed. The present results do not support a role for algal–bacterial interactions in dinoflagellate toxin synthesis. However, variations observed in the toxin profile of A. tamarense cells during culture experiments give new clues for the understanding of biosynthetic pathways of saxitoxin, a potent phycotoxin.     

Experimental demonstration of the roles of bacteria and bacterivorous protozoa in plankton nutrient cycles

We have used a model food chain composed of a natural bacterial assemblage, a pennate diatom and a bacterivorous microflagellate to investigate the factors controlling the relative importance of bacteria and protozoa as sources for regenerated nitrogen in plankton communities. In bacterized diatom cultures in which diatom growth was nitrogen-limited, the carbon:nitrogen (C:N) ratio of the bacterial substrate greatly affected which population was responsible for the uptake of nitrogen. When nitrogen was added as NH ₄ ⁺ and the cultures were supplemented with glucose, the bacteria competed successfully with the algae for NH ₄ ⁺ and prevented the growth of algae by rapidly assimilating all NH ₄ ⁺ in the cultures. Bacterivorous protozoa inoculated into these cultures grazed the bacterial population and remineralized NH ₄ ⁺ , thus relieving the nitrogen limitation of algal growth and allowing an increase in algal biomass. In contrast, bacteria in cultures supplemented with the amino acid glycine (C:N = 2) were major remineralizers of nitrogen, and the influence of protozoan grazing was minimal. We conclude that the relative importance of bacteria and protozoa as nutrient regenerators in the detrital food loop is dependent largely on the overall carbon:nutrient ratio of the bacterial substrate. The role of bacterivorous protozoa as remineralizers of a growth-limiting nutrient is maximal in situations where the carbon:nutrient ratio of the bacterial substrate is high.     

Biomass and production of phyto- and bacterio-plankton in eutrophic Lake Tystrup, Denmark

SUMMARY.

• 1 Phytoplankton primary production and biomass were compared with bacterial secondary production estimated by means of frequency of dividing cells (FDC) in eutrophic Lake Tystrup. Denmark.
• 2 In the upper part of the photic zone, bacterial secondary production constituted 12% of the carbon fixed by the phytoplankton. In vertical profiles, bacterial secondary production ranged from 7.6% (early spring) to 121% (during August) of the carbon fixed by the algae.
• 3 A close relationship was found between occurrence and activity of bacteria and algae, suggesting that released organic products are of primary importance to the bacteria.
• 4 The annual phytoplankton primary production was estimated as 227 g C m^-2 compared to 102 g C m^-2 assimilated by the bacteria, so 45% of the carbon fixed by the phytoplankton went through pelagic bacteria.

     

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.     

Phytoplankton response to waste-water nutrient enrichment in continuous culture

Three marine phytoplankton, Dunaliella tertiolecta Butcher, Phaeodactylum tricornutum Bohlin, and Thalassiosira pseudonana (3H) Hasle & Heindal, were grown on waste water-sea water mixtures in continuous-flow monocultures. P. tricornutum increased in biomass with increasing waste-water additions until a mixture of about 40 % waste water-60 % sea water was reached. The other species did not increase in biomass beyond a 20 % waste water-80 % sea water mixture and even showed some inhibition at higher waste water additions. The carbon/nitrogen ($\text{C}\text{N}$) ratio of the algae was consistently below 6 when nitrogen was not limiting growth, but increased with decreasing dilution rate under nitrogen-limiting conditions, depending on whether NH₄⁺-N or NO₃^?-N was the main nitrogen source.Species dominance in enriched cultures is controlled by a complex interaction of environmental factors. By altering the chemical composition ($\text{C}\text{N}$ ratio) of dominant phytoplankton such as P. tricornutum in mass culture through control of nitrogen source and concentration, it may be possible to increase the nutritional value of these organisms so that they represent a balanced diet for the growth of herbivorous shellfish.     

海水中藻菌共培养体系对碳氮磷的吸收转化

海洋环境中,细菌和微藻之间的物质交换是生源要素在自然界中迁移转化的重要方式。为进一步了解生源要素的生物地球化学循环,在实验室模拟条件下,研究了共培养体系中营养盐和有机物在细菌和微藻之间的转换。通过纯培养中肋骨条藻(Skeletonema costatum)、东海原甲藻(Prorocentrum donghaiense)、天然海水中的细菌以及藻菌混合培养,分析了营养盐和有机物随藻菌生物量的变化情况,并计算了溶解有机碳(DOC)和溶解有机氮(DON)的浓度比值[(DOC/DON)a]。结果发现,在共培养体系中,细菌对中肋骨条藻的生长有抑制作用,对东海原甲藻影响不明显;中肋骨条藻有利于细菌生长,东海原甲藻抑制细菌生长,这种不同可能与微藻的粒径有关。海洋细菌在2种藻的指数生长均期均会促进微藻吸收氨氮(NH_4-N),但在生长末期NH_4-N以释放为主。硝氮(NO_3-N)的浓度与藻的生长呈负相关,但在衰亡期NO_3-N略有增加,表明NO_3-N再生所需时间较长。细菌对硝氮的吸收量较少,但对其再生有贡献。细菌和中肋骨条藻对磷酸盐(PO_4-P)的吸收存在竞争,但与东海原甲藻的竞争关系不明显。不同培养体系中DOC浓度变化不同,在藻菌共培养体系中增加较快,纯藻培养体系中增加缓慢,在纯菌培养体系中缓慢减少。通过对DOC与DON浓度比值的分析,发现用判断颗粒有机碳(POC)来源的方法可以分析DOC的来源。     

Growth of Heterotrophic Bacteria and Algal Extracellular Products in Oligotrophic Waters

The unexpected observation of 200 to 400 coliform bacteria per 100 ml in an unpolluted pristine stream was studied within Grand Teton National Park, Wyo. The high numbers of waterborne bacteria occurred in mid- to late summer at a location where there was a coincidental bloom of an algal mat community. Periphyton samplers were used to measure the algal growth that coincided with the increase in number of bacteria. Laboratory studies followed the growth of various coliform bacteria in the supernatant obtained from a Chlorella culture isolated from the mat community. Mixed natural bacterial populations from the stream and pure cultures of water-isolated fecal and nonfecal coliforms increased by two to three orders of magnitude at 13°C when grown in the algal supernatant. Radioactive algal products were obtained by feeding an axenic Chlorella culture ¹⁴C-labeled bicarbonate under laboratory cultivation at 13°C with illumination. Radioactive organic material from the algae became incorporated into the particulate fraction of pure cultures of coliform bacteria as they reproduced and was later released as they died.     

EMILIANIA HUXLEYI (PRYMNESIOPHYCEAE): NITROGEN‐METABOLISM GENES AND THEIR EXPRESSION IN RESPONSE TO EXTERNAL NITROGEN SOURCES1

The availability and composition of dissolved nitrogen in ocean waters are factors that influence species composition in natural phytoplankton communities. The same factors affect the ratio of organic to inorganic carbon incorporation in calcifying species, such as the coccolithophore Emiliania huxleyi (Lohman) W. W. Hay et H. Mohler. E. huxleyi has been shown to thrive on various nitrogen sources, including dissolved organic nitrogen. Nevertheless, assimilation of dissolved nitrogen under nitrogen‐replete and ‐limited conditions is not well understood in this ecologically important species. In this study, the complete amino acid sequences for three functional genes involved in nitrogen metabolism in E. huxleyi were identified: a putative formamidase, a glutamine synthetase (GSII family), and assimilatory nitrate reductase. Expression patterns of the three enzymes in cells grown on inorganic as well as organic nitrogen sources indicated reduced expression levels of nitrate reductase when cells were grown on NH₄⁺ and a reduced expression level of the putative formamidase when growth was on NO₃^?. The data reported here suggest the presence of a nitrogen preference hierarchy in E. huxleyi. In addition, the gene encoding for a phosphate repressible phosphate permease was more highly expressed in cells growing on formamide than in cells growing on inorganic nitrogen sources. This finding suggests a coupling between phosphate and nitrogen metabolism, which might give this species a competitive advantage in nutrient‐depleted environments. The potential of using expression of genes investigated here as indicators of specific nitrogen‐metabolism strategies of E. huxleyi in natural populations of phytoplankton is discussed.     

Pelagic food web processes in an oligotrophic lake

Major pelagic carbon pathways, including primary production, release of extracellular products (EOC), bacterial production and zooplankton grazing were measured in oligotrophic Lake Almind (Denmark) and in enclosures (7 m³) subjected to artificial eutrophication. Simultaneous measurements at three days interval of carbon exchange rates and pools allowed the construction of carbon flow scenarios over a nineteen day experimental period.The flow of organic carbon was dominated by phytoplankton EOC release, which amounted from 44 to 58% of the net fixation of inorganic carbon. Gross bacterial production accounted for 33 to 75% of the primary production. The lower values of EOC release (44%) and bacterial production (33%) were found in the enclosures with added nutrients. The release of recently fixed photosynthetic products was the most important source of organic carbon to the bacterioplankton. Uptake of dissolved free amino acids was responsible for 52 to 62% of the gross bacterial production. Thus, amino acids constituted a significant proportion of the EOC. Zooplankton (< 50 µm) grazing on algae and bacteria accounted only for a minor proportion of the particulate production in May. Circumstantial evidence is presented that suggests the chrysophycean alga Dinobryon was the most important bacterial remover.The results clearly demonstrated EOC release and bacterial metabolism to be key processes in pelagic carbon cycling in this oligotrophic lake.     

Attached and Free-Floating Bacterioplankton in Howe Sound, British Columbia, a Coastal Marine Fjord-Embayment

Factors which influence the attachment of bacterioplankton to particles (including phytoplankton) were investigated by using (i) water samples removed from a coastal temperate fjord over an annual cycle and (ii) unialgal cultures of Prorocentrum minimum, Dunaliella tertiolecta, and Skeletonema costatum. Silt and salinity levels in this fjord seawater did not appear to influence bacterial attachment, but the percent attached bacteria was inversely related to both chlorophyll a concentrations and primary productivities. During periods of high primary productivities the percent attached bacteria was low, whereas during periods of low, increasing, and declining primary productivities the percent attached bacteria was high. A similar pattern of bacterial attachment was observed when the three phytoplankton were grown as batch cultures. The percent attached bacterial numbers increased upon the initiation of algal growth and after these cells stopped growing, but not while the algae were growing. We suggest that a major factor influencing the attachment of bacterioplankton is the physiological condition of their major nutrient source, the phytoplankton; mainly free-living bacteria are associated with growing phytoplankton, whereas a much greater proportion of the bacteria are attached among senescent phytoplankton populations.     

The diatom Thalassiosira rotula induces distinct growth responses and colonization patterns of Roseobacteraceae,Flavobacteria and Gammaproteobacteria

Diatoms as important phytoplankton components interact with and are colonized by heterotrophic bacteria. This colonization has been studied extensively in the past but a distinction between the bacterial colonization directly on diatom cells or on the aggregated organic material, exopolymeric substances (EPS), was little addressed. Here we show that the diatom Thalassiosira rotula and EPS were differently colonized by strains of Roseobacteraceae and Flavobacteriaceae in two and tree partner treatments and an enriched natural bacterial community as inoculum. In two partner treatments, the algae and EPS were generally less colonized than in the three partner treatments. Two strains benefitted greatly from the presence of another partner as the proportions of their subpopulations colonizing the diatom cell and the EPS were much enhanced relative to their two partner treatments. Highest proportions of bacteria colonizing the diatom and EPS occurred in the treatment inoculated with the enriched natural bacterial community. Dissolved organic carbon, amino acids and carbohydrates produced by T. rotula were differently used by the bacteria in the two and three partner treatments and most efficiently by the enriched natural bacterial community. Our approach is a valid model system to study physico-chemical bacteria-diatom interactions with increasing complexity.     

Host-Specificity and Dynamics in Bacterial Communities Associated with Bloom-Forming Freshwater Phytoplankton

Many freshwater phytoplankton species have the potential to form transient nuisance blooms that affect water quality and other aquatic biota. Heterotrophic bacteria can influence such blooms via nutrient regeneration but also via antagonism and other biotic interactions. We studied the composition of bacterial communities associated with three bloom-forming freshwater phytoplankton species, the diatom Aulacoseira granulata and the cyanobacteria Microcystis aeruginosa and Cylindrospermopsis raciborskii. Experimental cultures incubated with and without lake bacteria were sampled in three different growth phases and bacterial community composition was assessed by 454-Pyrosequencing of 16S rRNA gene amplicons. Betaproteobacteria were dominant in all cultures inoculated with lake bacteria, but decreased during the experiment. In contrast, Alphaproteobacteria, which made up the second most abundant class of bacteria, increased overall during the course of the experiment. Other bacterial classes responded in contrasting ways to the experimental incubations causing significantly different bacterial communities to develop in response to host phytoplankton species, growth phase and between attached and free-living fractions. Differences in bacterial community composition between cyanobacteria and diatom cultures were greater than between the two cyanobacteria. Despite the significance, major differences between phytoplankton cultures were in the proportion of the OTUs rather than in the absence or presence of specific taxa. Different phytoplankton species favoring different bacterial communities may have important consequences for the fate of organic matter in systems where these bloom forming species occur. The dynamics and development of transient blooms may also be affected as bacterial communities seem to influence phytoplankton species growth in contrasting ways.     

Growth Response and Modifications of Organic Nitrogen Compounds in Pure and Mixed Cultures of Lactic Acid Bacteria from Wine

The interactions between the proteolytic X₂L strain of Oenococcus oeni and the non-proteolytic 12p strain of Pediococcus pentosaceus were assayed. The characteristics of cell growth, protein degradation, and amino acid production of both strains were determined in pure and mixed cultures. O. oeni showed poor cell growth and greater ability in the release of amino acids to the extracellular medium, whereas P. pentosaceus showed a higher yield in cell production with a decrease in the amino acid concentration in the medium. P. pentosaceus especially consumed essential amino acids for growth, and O. oeni released several of the essential amino acids important for growth of P. pentosaceus. In the mixed culture, mutualism was observed. The higher activity of the proteolytic system of O. oeni in mixed culture produced an increase in cell growth and in the amount of essential amino acids released. These findings provide new knowledge about the metabolic interactions between lactic acid bacteria isolated from wine when proteins are degraded in mixed bacterial populations.