Determining critical nutrient thresholds needed to control harmful cyanobacterial blooms in eutrophic Lake Taihu, China.

Nutrient overenrichment has led to dramatic increases in harmful cyanobacterial blooms, creating serious threats to drinking water supplies, ecological and economic sustainability of freshwater ecosystems. Nutrient-cyanobacterial bloom interactions were examined in eutrophic Lake Taihu, China. In situ microcosm nutrient dilution bioassays and mesocosm nutrient addition experiments were conducted to determine nitrogen (N) and phosphorus (P) concentration and load thresholds needed to control cyanobacterial bloom formation. Blooms were dominated by toxic, non N2 fixing Microcystis spp, from May to December. Dilution bioassays showed seasonality in nutrient limitation, with P-availability controlling prebloom spring conditions and N-availability controlling summer-fall blooms. Nutrient dilution and enrichment bioassays indicated that total nitrogen (TN) and total phosphorus (TP) concentration thresholds should be targeted at below 0.80 mg L(­1) and 0.05 mg L(­1), respectively, to limit intrinsic growth rates of Microcystis dominated blooms. Based on estimates of nutrient loading and observed stoichiometry of phytoplankton biomass, 61­71% TN and 20­46% TP reduction are necessary to bring Taihu's phytoplankton biomass to "acceptable" sub-bloom conditions of less than 20 µg L(­1) chlorophyll a.

Spatiotemporal patterns and ecophysiology of toxigenic microcystis blooms in Lake Taihu, China: implications for water quality management.

Whole lake monitoring of hypertrophic Lake Taihu, China, was conducted during the summers of 2009-2010, with the intent of identifying environmental factors influencing Microcystis bloom formation and promoting the growth of toxigenic strains (mcyE possessing). Low N:P ratios (replete N & P)appeared to select for toxigenic populations of Microcystis spp., whereas nontoxic Microcystis spp. strains were dominant in more nutrient limited regions of the lake. Chlorophyll a (Adj. R(2) = 0.83, p < 0.0001) was equally predicative of microcystin variance across the lake as fluorescence based real-time quantitative PCR (qPCR) measurements of microcystin synthetase E (mcyE) gene equivalents (Adj. R(2) = 0.85, p < 0.0001). Interestingly, chlorophyll a was identified as a more robust and useful metric for predicting microcystin concentrations than qPCR measurements enumerating the total Microcystis population based on c-phycocyanin (α subunit; cpcA) gene equivalents (Adj. R(2) = 0.61, p < 0.0001). Overall, the lakewide composition of Microcystis spp. was highly variable over time and space, and on average the population consisted of 36 ± 12% potentially toxic cells. On the basis of this study's findings, a framework for the design and implementation of a water safety plan for Taihu water quality managers and public health officials is proposed.

Perspective: Advancing the research agenda for improving understanding of cyanobacteria in a future of global change.

Harmful cyanobacterial blooms (=cyanoHABs) are an increasing feature of many waterbodies throughout the world. Many bloom-forming species produce toxins, making them of particular concern for drinking water supplies, recreation and fisheries in waterbodies along the freshwater to marine continuum. Global changes resulting from human impacts, such as climate change, over-enrichment and hydrological alterations of waterways, are major drivers of cyanoHAB proliferation and persistence. This review advocates that to better predict and manage cyanoHABs in a changing world, researchers need to leverage studies undertaken to date, but adopt a more complex and definitive suite of experiments, observations, and models which can effectively capture the temporal scales of processes driven by eutrophication and a changing climate. Better integration of laboratory culture and field experiments, as well as whole system and multiple-system studies are needed to improve confidence in models predicting impacts of climate change and anthropogenic over-enrichment and hydrological modifications. Recent studies examining adaptation of species and strains to long-term perturbations, e.g. temperature and carbon dioxide (CO2) levels, as well as incorporating multi-species and multi-stressor approaches emphasize the limitations of approaches focused on single stressors and individual species. There are also emerging species of concern, such as toxic benthic cyanobacteria, for which the effects of global change are less well understood, and require more detailed study. This review provides approaches and examples of studies tackling the challenging issue of understanding how global changes will affect cyanoHABs, and identifies critical information needs for effective prediction and management.

Detritus in lake tahoe: structural modification by attached microflora.

Readily identifiable groups of microorganisms present on nonliving particulate organic matter (detritus) in the upper waters of Lake Tahoe are attached in specific ways and appear responsible for detrital aggregation. This microflora is associated with active heterotrophic metabolism, but deeper waters possess little detrital microflora and little heterotrophic activity.

Direct measurement of o2-depleted microzones in marine oscillatoria: relation to n2 fixation.

Among the nitrogen (N(2))-fixing Cyanobacteria, the filamentous, nonheterocystous marine Oscillatoria spp. (Trichodesmium) appears enigmatic; it exhibits N(2) fixation in the presence of oxygenic photosynthesis without structural protection of the N(2-)fixing apparatus (nitrogenase) from potential inhibition by molecular oxygen (O(2)). Characteristically, N(2) fixation is largely confined to aggregates (bundles) of filaments. Previous work has suggested that spatial partitioning of photosynthesis and of N(2) fixation occurs in the bundles as a means of allowing both processes to occur contemporaneously. The probing of freshly sampled bundles with O(2) microelectrodes directly confirmed such partitioning by showing the presence of O(2-)depleted (reduced) microzones in photosynthetically active, N(2-)fixing bundles. Bundle size was directly related to both the development of internal reduced microzones and cellular N(2) fixation rates. By enhancing microzone formation, bundles optimize N(2) fixation as a means of supporting Oscillatoria spp. blooms in surficial, nitrogen-depleted tropical and subtropical waters.

Nitrogen-fixing anabaena: physiological adaptations instrumental in maintaining surface blooms.

Both laboratory and in situ studies indicate that the nitrogen-fixing blue-green nuisance algae Anabaena spp. have developed adaptive means of dominating surface lake waters. During the dramatic diurnal shifts in surface light intensity and oxygen saturation accompanying blooms, Anabaena can overcome oxygen toxicity by sequential optimization of carbon dioxide and nitrogen fixation and by pigment alteration. These mechanisms allow optimal utilization of the radiant energy while minimizing competition for photoreductant between two main energy-demanding processes.

Role of chemotaxis in establishing a specific nitrogen-fixing cyanobacterial-bacterial association.

A specific association with the cyanobacterium Anabaena oscillarioides was established by positive bacterial (pseudomonad) chemotaxis to Anabaena oscillarioides heterocysts. This association enhanced nitrogen fixation in A. oscillarioides, and positive chemotaxis was particularly strong during periods of active nitrogen fixation. Addition of compounds known to elicit positive chemotaxis in pseudomonads interfered with the establishment of the association, while removal of these compounds led to reestablishment of the association. Anabaena oscillarioides excretion products, some of which are exuded from heterocyst-vegetative cell junctions, are likely to be responsible for positive chemotactic responses. Chemotaxis-controlled associations such as this one explain in part why aquatic bacterial-algal and bacterial-particle associations occur sporadically and are heterogeneously distributed in time and space.

Perennial Antarctic lake ice: an oasis for life in a polar desert.

The permanent ice covers of Antarctic lakes in the McMurdo Dry Valleys develop liquid water inclusions in response to solar heating of internal aeolian-derived sediments. The ice sediment particles serve as nutrient (inorganic and organic)-enriched microzones for the establishment of a physiologically and ecologically complex microbial consortium capable of contemporaneous photosynthesis, nitrogen fixation, and decomposition. The consortium is capable of physically and chemically establishing and modifying a relatively nutrient- and organic matter-enriched microbial "oasis" embedded in the lake ice cover.

Structure and function of anthropogenically altered microbial communities in coastal waters.

Human-based (anthropogenic) nutrient and other pollutant enrichment of the world's coastal waters is causing unprecedented changes in microbial community structure and function. Symptoms of these changes include accelerating eutrophication, the proliferation of harmful microalgal blooms, excessive oxygen consumption (hypoxia, anoxia), increasing toxicity, altered routes and fluxes of organic and inorganic matter cycling, and disruption of food webs. Biogeochemical and trophic consequences are expanding on local, regional and global scales.

The role of nutrient loading and eutrophication in estuarine ecology.

Eutrophication is a process that can be defined as an increase in the rate of supply of organic matter (OM) to an ecosystem. We provide a general overview of the major features driving estuarine eutrophication and outline some of the consequences of that process. The main chemical constituent of OM is carbon (C), and therefore rates of eutrophication are expressed in units of C per area per unit time. OM occurs in both particulate and dissolved forms. Allochthonous OM originates outside the estuary, whereas autochthonous OM is generated within the system, mostly by primary producers or by benthic regeneration of OM. The supply rates of limiting nutrients regulate phytoplankton productivity that contributes to inputs of autochthonous OM. The trophic status of an estuary is often based on eutrophication rates and can be categorized as oligotrophic (<100 g C m(-2) y(-1), mesotrophic (100-300 g C m(-2) y(-1), eutrophic (300-500 g C m(-2) y(-1), or hypertrophic (>500 g C m(-2) y(-1). Ecosystem responses to eutrophication depend on both export rates (flushing, microbially mediated losses through respiration, and denitrification) and recycling/regeneration rates within the estuary. The mitigation of the effects of eutrophication involves the regulation of inorganic nutrient (primarily N and P) inputs into receiving waters. Appropriately scaled and parameterized nutrient and hydrologic controls are the only realistic options for controlling phytoplankton blooms, algal toxicity, and other symptoms of eutrophication in estuarine ecosystems.

Microscale physiological and ecological studies of aquatic cyanobacteria: macroscale implications.

Cyanobacteria have had a profound and unparalleled biogeochemical impact on the earth's biosphere. As the first oxygenic phototrophs, cyanobacteria were responsible for the transition from anaerobic to aerobic life. Ironically, molecular oxygen (O2) is inhibitory to critical components of cyanobacterial metabolism, including photosynthesis and nitrogen fixation. Cyanobacteria have developed a great variety of biochemical, structural, and biotic adaptations ensuring optimal growth and proliferation in diverse oxic environments to counter this difficult situation. Structurally, cyanobacteria reveal remarkable diversity, including the formation of highly differentiated, O2-deplete cells (heterocysts), multicellularity as trichomes, and aggregates, that, among N2-fixing genera, facilitate division of labor between aerobic and anaerobic processes. Cyanobacteria enjoy unique consortial and symbiotic associations with other microorganisms, higher plants, and animals, in which O2 consumption is closely coupled in time and space to its production. Because as prokaryotes they are devoid of O2-consuming organelles (e.g., mitochondria), cyanobacteria have developed alternative strategies for locally protecting O2-sensitive processes, including consortial relationships with other microorganisms. Specific organic compounds released by cyanobacteria are capable of chemotactically attracting bacterial consorts, which in turn attach to the host cyanobacteria, consume O2, and recycle inorganic nutrients within the cyanobacterial "phycosphere." Multicellularity and aggregation lead to localized O2 gradients and hypoxic/anoxic microzones in which O2-sensitive processes can coexist. Microscale partitioning of O2-producing and O2-inhibited processes promotes contiguous and effective metabolite and nutrient exchange between these processes in oxygenated waters, representing a bulk of the world's oceanic and freshwater ecosystems.

Microscale characterization of dissolved organic matter production and uptake in marine microbial mat communities.

Intertidal marine microbial mats exhibited biologically mediated uptake of low molecular weight dissolved organic matter (DOM), including D-glucose, acetate, and an L-amino acid mixture at trace concentrations. Uptake of all compounds occurred in darkness, but was frequently enhanced under natural illumination. The photosystem 2 inhibitor, 3-(3,4-dichlorophenyl)-1,1-dimethyl urea (DCMU) generally failed to inhibit light-stimulated DOM uptake. Occasionally, light plus DCMU-amended treatments led to uptake rates higher than light-incubated samples, possibly due to phototrophic bacteria present in subsurface anoxic layers. Uptake was similar with either 3H- or 14C-labeled substrates, indicating that recycling of labeled CO2 via photosynthetic fixation was not interfering with measurements of light-stimulated DOM uptake. Microautoradiographs showed a variety of pigmented and nonpigmented bacteria and, to a lesser extent, cyanobacteria and eucaryotic microalgae involved in light-mediated DOM uptake. Light-stimulated DOM uptake was often observed in bacteria associated with sheaths and mucilage surrounding filamentous cyanobacteria, revealing a close association of organisms taking up DOM with photoautotrophic members of the mat community. The capacity for dark- and light-mediated heterotrophy, coupled to efficient retention of fixed carbon in the mat community, may help optimize net production and accretion of mats, even in oligotrophic waters.

Distribution of Nitrogen-Fixing Microorganisms along the Neuse River Estuary, North Carolina.

Nitrogen fixation genes (nifH) were amplified and sequenced from DNA extracted from surface water samples collected from six stations along the length of the Neuse River Estuary, North Carolina, in order to determine the distribution of nitrogen-fixing organisms in the transition from fresh- to saltwater. Nitrogenase genes were detected in all samples by a nested polymerase chain reaction method, and the amplification products from the upriver, midriver, and downriver stations were cloned, sequenced, and used for phylogenetic analysis. The composition of nifH clone libraries from upriver, midriver, and downriver stations (each composed of 14 randomly selected clones) were very diverse (samples from upriver and midriver stations were composed of 14 unique sequences, downriver station composed of 7 unique sequences) and differed among the stations. Some phylotypes were found at more than one station, but were usually found in the upriver and midriver stations or in the midriver and downriver stations, indicating that the phylotypes were probably transported along the river. Cyanobacterial nifH were not found at the most upriver site, but were a large fraction of sequences (50%) recovered from the downriver station, where nitrate concentration was an order of magnitude lower and salinity was higher. In contrast, g proteobacteria nifH sequences were much more common at the midriver and upriver sites (58% and 64%, respectively), compared to the downriver site (14%). Results indicate that substantially different nitrogen-fixing assemblages are present along the river, reflecting differential watershed hydrological inputs, sedimentation, and environmental selection pressures, along the salinity gradient.

Diazotrophy in Modern Marine Bahamian Stromatolites.

N2 fixation (nitrogenase activity), primary production, and diazotrophic community composition of stromatolite mats from Highborne Cay, Exuma, Bahamas, were examined over a 2-year period (1997-1998). The purpose of the study was to characterize the ecophysiology of N2 fixation in modern marine stromatolites. Microbial mats are an integral surface component of these stromatolites and are hypothesized to have a major role in stromatolite formation and growth. The stromatolite mats contained active photosynthetic and diazotrophic assemblages that exhibited temporal separation of nitrogenase activity (NA) and photosynthesis. Maximal NA was detected at night. Seasonal differences in NA and net O2 production were observed. Photosynthetic activity and the availability of reduced organic carbon appear to be the key determinants of NA. Additions of the de novo protein synthesis inhibitor chloramphenicol did not inhibit NA in March 1998, but greatly inhibited NA in August 1998. Partial sequence analysis of the nifH gene indicates that a broad diversity of diazotrophs may be responsible for NA in the stromatolites.

Effects of the blue-green alga Microcystis aeruginosa on zooplankton competitive relations.

Field distribution patterns and laboratory feeding experiments have suggested that blooms of colonial blue-green algae strongly inhibit relatively large-bodied daphnid cladocerans. We conducted laboratory experiments to test the hypothesis that blooms of the colonial blue-green alga Microcystis aeruginosa would shift competitive dominance away from large-bodied daphnid cladocerans toward smaller-bodied cladocerans, copepods, and rotifers. In laboratory competition experiments, increasing the proportion of M. aeruginosa in the algal food supply resulted in a shift from dominance by the relatively largebodied cladoceran Daphnia ambigua to dominace by the copepod Diaptomus reighardi. The small-bodied cladoceran Bosmina longirostris was always numerically heavily dominant over D. ambigua, but its estimated population biomasses were only slightly higher than those of D. ambigua. Daphnia ambigua consistently outcompeted the rotifer Brachionus calyciflorus. Our results demonstrate that blooms of M. aeruginosa can alter zooplankton competitive relations in laboratory experiments, favoring small-bodied cladocerans and copepods at the expense of large-bodied cladocerans. However, contrary to predictions, blooms of M. aeruginosa did not improve the competitive ability of rotifers.

Spatiotemporal variability of wet atmospheric nitrogen deposition to the Neuse River Estuary, North Carolina.

Excessive nitrogen (N) loading to N-sensitive waters such as the Neuse River estuary (North Carolina) has been shown to promote changes in microbial and algal community composition and function (harmful algal blooms), hypoxia and anoxia, and fish kills. Previous studies have estimated that wet atmospheric deposition of nitrogen (WAD-N), as deposition of dissolved inorganic nitrogen (DIN: NO3-, NH3/NH4+) and dissolved organic nitrogen, may contribute at least 15% of the total externally supplied or "new" N flux to the coastal waters of North Carolina. In a 3-yr study from June 1996 to June 1999, we calculated the weekly wet deposition of inorganic and organic N at eleven sites on a northwest-southeast transect in the watershed. The annual mean total (wet DIN + wet organics) WAD-N flux for the Neuse River watershed was calculated to be 956 mg N/m2/yr (15026 Mg N/yr). Seasonally, the spring (March-May) and summer (June-August) months contain the highest total weekly N deposition; this pattern appears to be driven by N concentration in precipitation. There is also spatial variability in WAD-N deposition; in general, the upper portion of the watershed receives the lowest annual deposition and the middle portion of the watershed receives the highest deposition. Based on a range of watershed N retention and in-stream riverine processing values, we estimate that this flux contributes approximately 24% of the total "new" N flux to the estuary.

Etiologies, observations and reporting of estuarine finfish lesions.

Lesions in estuarine finfish are associated with a variety of organisms including parasites and bacterial, viral, and fungal infectious agents. In addition, trauma, suboptimal water quality, and other abiotic stress factors may result in the loss of homeostasis. We have observed solitary ulcerative lesions on menhaden sampled from the Chesapeake Bay, Maryland, the Pimlico River, North Carolina, and the St. Johns River, Florida. Histologically, the lesions demonstrated a marked chronic inflammatory infiltrate and granulomas in response to fungal hyphae throughout large areas of exposed necrotic muscle. Gram-negative rod-shaped bacteria were also observed in the lesions, a common finding in ulcers of aquatic organisms. Similar observations in menhaden and other species have been described previously in the literature as ulcerative mycosis, mycotic granulomatosis, red spot disease, and epizootic ulcerative syndrome. Despite the many different known causes of fish lesions, the popular press and the scientific literature have recently emphasized Pfiesteria piscicida and other Pfiesteria-like dinoflagellates (and their bioactive compounds) as the primary causative agent for finfish lesions, particularly mycotic granulomatous ulcers in Atlantic menhaden. While some laboratory data suggest that Pfiesteria may play a role in field-observed lesions, much more cause-and-effect evidence is needed to determine the importance of other risk factors, both alone or and in combination with Pfiesteria. In order to better understand the etiology of lesion initiation and progression in estuarine finfish, accurate assessments of environmental conditions collected on appropriate temporal and spatial scales, and fish morphological indicators consistent with gross and histological pathologic terminology, should be used for reporting fish lesion observations and kills. Further, this outlook will help to avoid bias and may foster a broader perspective for examining the health of estuarine systems in general.

Harmful freshwater algal blooms, with an emphasis on cyanobacteria.

Suspended algae, or phytoplankton, are the prime source of organic matter supporting food webs in freshwater ecosystems. Phytoplankton productivity is reliant on adequate nutrient supplies; however, increasing rates of nutrient supply, much of it manmade, fuels accelerating primary production or eutrophication. An obvious and problematic symptom of eutrophication is rapid growth and accumulations of phytoplankton, leading to discoloration of affected waters. These events are termed blooms. Blooms are a prime agent of water quality deterioration, including foul odors and tastes, deoxygenation of bottom waters (hypoxia and anoxia), toxicity, fish kills, and food web alterations. Toxins produced by blooms can adversely affect animal (including human) health in waters used for recreational and drinking purposes. Numerous freshwater genera within the diverse phyla comprising the phytoplankton are capable of forming blooms; however, the blue-green algae (or cyanobacteria) are the most notorious bloom formers. This is especially true for harmful toxic, surface-dwelling, scum-forming genera (e.g., Anabaena, Aphanizomenon, Nodularia, Microcystis) and some subsurface bloom-formers (Cylindrospermopsis, Oscillatoria) that are adept at exploiting nutrient-enriched conditions. They thrive in highly productive waters by being able to rapidly migrate between radiance-rich surface waters and nutrient-rich bottom waters. Furthermore, many harmful species are tolerant of extreme environmental conditions, including very high light levels, high temperatures, various degrees of desiccation, and periodic nutrient deprivation. Some of the most noxious cyanobacterial bloom genera (e.g., Anabaena, Aphanizomenon, Cylindrospermopsis, Nodularia) are capable of fixing atmospheric nitrogen (N2), enabling them to periodically dominate under nitrogen-limited conditions. Cyanobacteria produce a range of organic compounds, including those that are toxic to higher-ranked consumers, from zooplankton to further up the food chain. Both N2- and non-N2-fixing genera participate in mutualistic and symbiotic associations with microorganisms, higher plants, and animals. These associations appear to be of great benefit to their survival and periodic dominance. In this review, we address the ecological impacts and environmental controls of harmful blooms, with an emphasis on the ecology, physiology, and management of cyanobacterial bloom taxa. Combinations of physical, chemical, and biotic features of natural waters function in a synergistic fashion to determine the sensitivity of water bodies. In waters susceptible to blooms, human activities in water- and airsheds have been linked to the extent and magnitudes of blooms. Control and management of cyanobacterial and other phytoplankton blooms invariably includes nutrient input constraints, most often focused on nitrogen (N) and/or phosphorus (P). The types and amount of nutrient input constraints depend on hydrologic, climatic, geographic, and geologic factors, which interact with anthropogenic and natural nutrient input regimes. While single nutrient input constraints may be effective in some water bodies, dual N and P input reductions are usually required for effective long-term control and management of harmful blooms. In some systems where hydrologic manipulations (i.e., plentiful water supplies) are possible, reducing the water residence time by enhanced flushing and artificial mixing (in conjunction with nutrient input constraints) can be particularly effective alternatives. Implications of various management strategies, based on combined ecophysiological and environmental considerations, are discussed.

Nitrogenase activity and nifH expression in a marine intertidal microbial mat.

N(2) fixation, diazotrophic community composition, and organisms actively expressing genes for N(2) fixation were examined over at 3-year period (1997-1999) for intertidal microbial mats on a sand flat located in the Rachel Carson National Estuarine Research Reserve (RCNERR) (Beaufort, NC, USA). Specifically, diel variations of N(2) fixation in the mats from the RCNERR were examined. Three distinct diel patterns of nitrogenase activity (NA) were observed. NA responses to short-term inhibitions of photosynthesis corresponded to one of the three patterns. High rates of NA were observed during peak O(2) production periods for diel experiments during summer months. Different types of NA diel variations correspond to different stages of mat development. Chloramphenicol treatments indicated that the mechanism of protein synthesis supporting NA changed throughout the day. Analysis of mat DNA and RNA gave further evidence suggesting that in addition to cyanobacteria, other functional groups were responsible for the NA observed in the RCNERR mats. The role of microbial diversity in the N(2) fixation dynamics of these mats is discussed.

Role of heterotrophic bacteria in promoting N2 fixation byAnabaena in aquatic habitats.

Anabaena species are commonly colonized by bacteria, especially during N2-fixing blooms. Generally these associations do not represent bacterial attack on algal hosts. Instead, the algal N2-fixing capabilities are increased in the presence of the bacteria. Possible mechanisms promoting the mutual growth of algae and attached bacteria were investigated by observing specific sites of bacterial attachment, by noting reduced microzones created by the bacteria, and by locating sites of bacterial uptake of organics representative of algal excretion products.Attached bacteria show preference for typical algal excretion products and their growth is enhanced by such products. In return, enhancement of algal nitrogenase activity occurs when bacteria create O2-consuming microzones around the nitrogenase-bearing heterocysts.