Selective suppression of harmful cyanobacteria in an entire lake with hydrogen peroxide

Although harmful cyanobacteria form a major threat to water quality, few methods exist for the rapid suppression of cyanobacterial blooms. Since laboratory studies indicated that cyanobacteria are more sensitive to hydrogen peroxide (H₂O₂) than eukaryotic phytoplankton, we tested the application of H₂O₂ in natural waters. First, we exposed water samples from a recreational lake dominated by the toxic cyanobacterium Planktothrix agardhii to dilute H₂O₂. This reduced the photosynthetic vitality by more than 70% within a few hours. Next, we installed experimental enclosures in the lake, which revealed that H₂O₂ selectively killed the cyanobacteria without major impacts on eukaryotic phytoplankton, zooplankton, or macrofauna. Based on these tests, we introduced 2 mg L⁻¹ (60 μM) of H₂O₂ homogeneously into the entire water volume of the lake with a special dispersal device, called the water harrow. The cyanobacterial population as well as the microcystin concentration collapsed by 99% within a few days. Eukaryotic phytoplankton (including green algae, cryptophytes, chrysophytes and diatoms), zooplankton and macrofauna remained largely unaffected. Following the treatment, cyanobacterial abundances remained low for 7 weeks. Based on these results, we propose the use of dilute H₂O₂ for the selective elimination of harmful cyanobacteria from recreational lakes and drinking water reservoirs, especially when immediate action is urgent and/or cyanobacterial control by reduction of eutrophication is currently not feasible. A key advantage of this method is that the added H₂O₂ degrades to water and oxygen within a few days, and thus leaves no long-term chemical traces in the environment.     

Seasonal occurrence and toxicity of Microcystis in impoundments of the Huron River, Michigan, USA

Occurrence and toxicity of Microcystis spp. were measured by ELISA in three impoundments of the Huron River in southeast Michigan, USA. Assays were conducted weekly from June through October 2005. Additional samples were collected to assess microcystin concentrations throughout the drinking water treatment process at the Ann Arbor Water Treatment Facility. Water column stability, nutrient concentrations, and N:P ratios were examined as potential predictors of phycocyanin and microcystin. Microcystin was found in two of the impoundments at seasonally varying concentrations. Microcystis presence was associated with N:P ratios between 40 and 80 by moles, and toxin levels typically peaked one to two weeks after a peak in phycocyanin. The toxin was also detected at low levels at all stages of the drinking water treatment process. Freezing and thawing water samples prior to analysis yielded maximum microcystin assay concentrations. Experiments indicated that the competitive ELISA method is susceptible to false negative reporting. This is the first report of algal toxins in this catchment, and results demonstrate that sensitive and rapid analytical methods offer the chance to link the dynamics of toxin production with environmental conditions.     

Establishment of an Alert Level Framework for cyanobacteria in drinking water resources by using the Algae Online Analyser for monitoring cyanobacterial chlorophyll a

The Algae Online Analyser (AOA) fluorometer simultaneously distinguishes four different phytoplankton groups by their specific fluorescence spectra and thus allows for real-time in-situ chlorophyll a measurements per algal group. This AOA was used for monitoring cyanobacterial chlorophyll a in the drinking water at the Bronis?awow Bay abstraction point in Sulejow Reservoir (Poland). The main goal of this research was to develop an early warning method for the detection of cyanobacterial biovolume in the source water, in order to establish an Alert Level Framework for the drinking water abstraction point in Sulejow Reservoir. A positive correlation between cyanobacterial biovolume, as determined by conventional methods, and cyanobacterial chlorophyll a, as measured by the AOA, was found (p < 0.05). The results of this study were used to determine threshold values for the Alert Level Framework, based on cyanobacterial chlorophyll a concentrations in the source water of Sulejow Reservoir. The presented threshold values are determined specifically for this abstraction point, but the principles can be applied to other locations.     

Simultaneous determination of microcystin contaminations in various vertebrates (fish, turtle, duck and water bird) from a large eutrophic Chinese lake, Lake Taihu, with toxic Microcystis blooms

This is the first to conduct simultaneous determination of microcystin (MC) contaminations in multi-groups of vertebrates (fish, turtle, duck and water bird) from Lake Taihu with Microcystis blooms. MCs (-RR, -YR, -LR) in Microcystis scum was 328 μg g^− 1 DW. MCs reached 235 μg g^− 1 DW in intestinal contents of phytoplanktivorous silver carp, but never exceeded 0.1 μg g^− 1 DW in intestinal contents of other animals. The highest MC content in liver of fish was in Carassius auratus (150 ng g^− 1 DW), followed by silver carp and Culter ilishaeformis, whereas the lowest was in common carp (3 ng g^− 1 DW). In livers of turtle, duck and water bird, MC content ranged from 18 to 30 ng g^− 1 DW. High MC level was found in the gonad, egg yolk and egg white of Nycticorax nycticorax and Anas platyrhynchos, suggesting the potential effect of MCs on water bird and duck embryos. High MC contents were identified for the first time in the spleens of N. nycticorax and A. platyrhynchos (6.850 and 9.462 ng g^− 1 DW, respectively), indicating a different organotropism of MCs in birds. Lakes with deaths of turtles or water birds in the literatures had a considerably higher MC content in both cyanobacteria and wildlife than Lake Taihu, indicating that toxicity of cyanobacteria may determine accumulation level of MCs and consequently fates of aquatic wildlife.     

Elucidation and short-term forecasting of microcystin concentrations in Lake Suwa (Japan) by means of artificial neural networks and evolutionary algorithms

Non-supervised artificial neural networks (ANN) and hybrid evolutionary algorithms (EA) were applied to analyse and model 12 years of limnological time-series data of the shallow hypertrophic Lake Suwa in Japan. The results have improved understanding of relationships between changing microcystin concentrations, Microcystis species abundances and annual rainfall intensity. The data analysis by non-supervised ANN revealed that total Microcystis abundance and extra-cellular microcystin concentrations in typical dry years are much higher than those in typical wet years. It also showed that high microcystin concentrations in dry years coincided with the dominance of the toxic Microcystis viridis whilst in typical wet years non-toxic Microcystis ichthyoblabe were dominant. Hybrid EA were used to discover rule sets to explain and forecast the occurrence of high microcystin concentrations in relation to water quality and climate conditions. The results facilitated early warning by 3-days-ahead forecasting of microcystin concentrations based on limnological and meteorological input data, achieving an r(2)=0.74 for testing.     

Differences in microcystin production and genotype composition among Microcystis colonies of different sizes in Lake Taihu

The cyanobacterium Microcystis, which occurs as colonies of different sizes under natural conditions, can produce toxic microcystins (MCs). To monitor the toxicity and assess the risk of Microcystis blooms in Lake Taihu, it is important to investigate the relationship between MC production and Microcystis colony size. In this study, we classified Microcystis collected from Zhushan Bay of Lake Taihu during blooms into four classes with size of <50 μm, 50–100 μm, 100–270 μm and >270 μm and studied their differences in MC production and genetic structure. The results showed that colonies with size of <50, 50–100, 100–270 and >270 μm produced 12.2 ± 11.2%, 19.5 ± 7.9%, 61.3 ± 12.6%, and 7.0 ± 9.6% of total MC, respectively. The proportion of cell density of colonies with size of 50–100, 100–270 and >270 μm was positively correlated with MC concentration during blooms, while that of colonies with size of <50 μm was negatively correlated. The MC cell quota tended to be higher during blooms in colonies with larger size except that of colonies with size of 100–270 μm was higher than that of colonies with size of >270 μm from June 11 to September 16. Colonies with size of <50 μm showed the highest proportion of the less toxic MC congener MC-RR, and colonies with size of >100 μm showed higher proportion of the most toxic MC congener MC-LR than colonies with size of <100 μm. Real-time PCR indicated that larger colonies had higher proportion of potential toxic genotype. Principal component analysis of PCR-denaturing gradient gel electrophoresis profile showed that cpcBA and mcyJ genotype compositions were different between colonies with size of <50 μm and colonies with size of >50 μm, and cpcBA genotype composition was also different among colonies with size of 50–100 μm, 100–270 μm and >270 μm. These results indicated that MC cell quota and congener composition were different in Microcystis colonies with different sizes in Lake Taihu during blooms, and the differences in MC production in colonies with different size resulted chiefly from the difference in their genotype composition. Therefore, the authorities of water quality monitoring and drinking water supply service in Lake Taihu should be alert that the toxicity of Microcystis colony with different size was different during blooms, and the high abundance of colonies larger than 50 μm could be an indicator of relatively high bloom toxicity.     

The fate of cyanobacterial blooms in vegetated and unvegetated sediments of a shallow eutrophic lake: A stable isotope tracer study

An experiment using nitrogen stable isotope tracer (¹⁵N) was conducted to track the fate of nitrogen derived from cyanobacterial blooms and the effectiveness with which the seasonal blooms are retained by vegetated and unvegetated sediment in a large shallow eutrophic lake (Lake Taihu, China). ¹⁵N enriched Microcystis was injected into both unvegetated sediment and sediment occupied by common reed (Phragmites australis) in the littoral zone. Nutrient retention by the vegetated sediment was greater than by the unvegetated sediment, resulting in higher δ¹⁵N in the sediment nitrogen pool. The labeled Microcystis material was also distributed deeper into the vegetated sediment than the unvegetated sediment. A portion of the Microcystis-derived nitrogen was quickly assimilated, appearing first in the belowground biomass and subsequently in the aboveground biomass of the reed plants. The labeled nitrogen was found to support new growth as evidenced by ¹⁵N enrichment of new leaves. This study indicates that common reed beds in the littoral zone may play an important role in retention of sedimented planktonic materials.     

Benthic cyanobacteria (Oscillatoriaceae) that produce microcystin-LR, isolated from four reservoirs in southern California

Cyanobacteria that produce the toxin microcystin have been isolated from many parts of the world. Most of these organisms are planktonic; however, we report on several microcystin-producing benthic filamentous cyanobacterial isolates from four drinking-water reservoirs in southern California (USA): Lake Mathews, Lake Skinner, Diamond Valley Lake (DVL), and Lake Perris. Some samples of benthic material from these reservoirs tested positive for microcystin by an ELISA tube assay, and all the positive samples had in common a green filamentous cyanobacterium 10-15microm in diameter. Seventeen unialgal strains of the organism were isolated and tested positive by ELISA, and 11 cultures of these strains were found to contain high concentrations of microcystin-LR (90-432microgL(-1)). The cultures were analyzed by protein phosphatase inhibition assay (PPIA) and HPLC with photodiode array detector (PDA) or liquid chromatography/mass spectrometry (LC/MS). Microcystin per unit carbon was determined for six cultures and ranged from 1.15 to 4.15microgmg(-1) C. Phylogenetic analysis of four cultures from Lake Skinner and DVL using cyanobacterial-specific PCR and sequencing of the partial 16S rRNA gene suggested the highest similarity to an unidentified cyanobacterium in the oscillatoriales, and to a Phormidium sp. Morphologically, some of the isolates were similar to Oscillatoria, and others resembled Lyngbya. The significance of these organisms lies in the relative scarcity of known toxin producers among freshwater benthic cyanobacteria, and also as a source of cell-bound microcystin in these reservoirs.     

Impact of toxic cyanobacteria on gastropods and microcystin accumulation in a eutrophic lake (Grand-Lieu, France) with special reference to Physa (= Physella) acuta

Hepatotoxic microcystins (MCs) produced by cyanobacteria are known to accumulate in gastropods following grazing of toxic cyanobacteria and/or absorption of MCs dissolved in water, with adverse effects on life history traits demonstrated in the laboratory. In the field, such effects may vary depending on species, according to their relative sensitivity and ecology. The aims of this study were to i) establish how various intensities of MC-producing cyanobacteria proliferations alter the structure of gastropod community and ii) compare MC tissue concentration in gastropods in the field with those obtained in our previous laboratory experiments on the prosobranch Potamopyrgus antipodarum and the pulmonate Lymnaea stagnalis. We explored these questions through a one-year field study at three stations at Grand-Lieu Lake (France) affected by different intensities of cyanobacteria proliferations. A survey of the community structure and MC content of both cyanobacteria and gastropods was associated with a caging experiment involving P. antipodarum and L. stagnalis. In total, 2592 gastropods belonging to 7 prosobranch and 16 pulmonate species were collected. However, distribution among the stations was unequal with 62% vs 2% of gastropods sampled respectively at the stations with the lowest vs highest concentrations of MC. Irrespective of the station, pulmonates were always more diverse, more abundant and occurred at higher frequencies than prosobranchs. Only the pulmonate Physa acuta occurred at all stations, with abundance and MC tissue concentration (≤ 4.32 µg g DW^− 1) depending on the degrees of MC-producing cyanobacteria proliferations in the stations; therefore, P. acuta is proposed as a potential sentinel species. The caging experiment demonstrated a higher MC accumulation in L. stagnalis (≤ 0.36 µg g DW^− 1 for 71% of individuals) than in P. antipodarum (≤ 0.02 µg g DW^− 1 for 12%), corroborating previous laboratory observations. Results are discussed in terms of differential gastropod sensitivity and MC transfer through the food web.     

Citizen monitoring: Testing hypotheses about the interactive influences of eutrophication and mussel invasion on a cyanobacterial toxin in lakes

An existing volunteer monitoring network in the state of Michigan was exploited to conduct a statewide survey of the cyanobacterial toxin, microcystin, and to test hypotheses about the interactive influences of eutrophication and dreissenid mussel invasion. A total of 77 lakes were sampled by citizen volunteers for microcystin, total phosphorus (TP) and chlorophyll a. Microcystin was measured in depth-integrated samples collected from the euphotic zone as well as in surface-water samples collected along the shoreline. Average microcystin in samples collected by volunteers was not different from samples collected side-by-side by professionals. Euphotic-zone microcystin was positively related to TP in lakes without dreissenids (uninvaded) but not in lakes with dreissenids (invaded). Regression-tree analysis indicated that euphotic-zone microcystin was eight times higher in the presence of dreissenids for lakes with TP between 5 and 10 μg L⁻¹. In contrast, euphotic-zone microcystin was almost identical in invaded and uninvaded lakes with TP between 10 and 26 μg L⁻¹. Across all lakes, microcystin concentrations at the surface were on average more than double, and in some cases an order-of-magnitude greater than, concentrations in the euphotic-zone. Given these results, it seems prudent to include dreissenid invasion status in forecasting models for microcystin, and to include shoreline sampling in monitoring programs aimed at assessing recreational exposure to cyanobacterial toxins.     

Analysis of dissolved microcystins in surface water samples from Kovada Lake, Turkey

Dissolved (extracellular) microcystin (MC) concentrations were determined at 3 sampling stations on Lake Kovada, Turkey. The dominant species of cyanobacteria found in August and September of 2006 were Microcystis aeruginosa, Synechococcus sp., Phormidium limosum, Phormidium formosa and Planktothrix limnetica. MC concentrations in water were measured by ELISA and MC variants were examined by HPLC-PDA. Quantitative analysis by HPLC indicated that five MC variants (MC-LR, -RR, -LA, -LW, -LF) were identified in water samples from Kovada Lake. The maximum concentration of dissolved MC-LW was 98.9 µg l^− 1 in October. MC-LR was only detected in May at a concentration of 0.5 µg l^− 1. The cross reactivity of the antibody (MC10E7) to variants such as MC-LA MC-LW & MC-LF was low. Hence the results determined by ELISA were lower than those determined by HPLC in September and October samples due to differences in the specificity of the antibody to MC variants. Total extraceullar MCs was quantified by ELISA and ranged from 0.73 to 48.5 µg MC-LR equivalents l^− 1, which in some cases exceeded the WHO provisional Guideline Value for MC-LR in drinking water. This study confirms that the lakes of Turkey should be monitored for toxic cyanobacteria and for MCs to avoid or reduce the potential exposure of people to these health hazards.     

Occurrence of dissolved and particle-bound taste and odor compounds in Swiss lake waters

The occurrence of algal taste and odor (T&O) compounds was investigated in three Swiss lakes which exhibit different nutrient levels from eutrophic to oligotrophic (Lake Greifensee, Lake Zurich and Lake Lucerne). Apart from dissolved T&O compounds, the study also encompassed particle-bound compounds, i.e., compounds that can be released from damaged algal cells during drinking water treatment. A combined instrumental (SPME-GC-MS) and sensory method was applied that allowed to detect and quantify T&O compounds in natural waters in the sub ppt to low ppt-range.In addition to the prominent T&O compounds geosmin and 2-methyl-isoborneol (MIB), four other T&O compounds could be detected in the lake waters, though all at relatively low concentrations (maximum concentrations of geosmin 19 ng L⁻¹, MIB 3 ng L⁻¹, β-ionone 27 ng L⁻¹, β-cyclocitral 7 ng L⁻¹, 2-isobutyl-3-methoxypyrazine 2 ng L⁻¹, 2-isopropyl-3-methoxypyrazine 16 ng L⁻¹). The concentration peaks typically occurred in the epilimnion during summer concurrent with a high phytoplankton biomass. Consistently, the concentration levels for most of the compounds varied substantially between the three lakes and generally decreased in the order eutrophic Lake Greifensee > mesotrophic Lake Zurich > oligotrophic Lake Lucerne. Furthermore, our data revealed that the occurrence of β-ionone was largely influenced by Planktothrix rubescens. This is the first time that a correlation between β-ionone and this cyanobacterium has been reported for natural waters.     

Comparison of the efficacy of an existing versus a locally developed metabolic fingerprint database to identify non-point sources of faecal contamination in a coastal lake

A comparison of the efficacy of an existing large metabolic fingerprint database of enterococci and Escherichia coli with a locally developed database was undertaken to identify the sources of faecal contamination in a coastal lake, in southeast Qld., Australia. The local database comprised of 776 enterococci and 780 E. coli isolates from six host groups. In all, 189 enterococci and 245 E. coli biochemical phenotypes (BPTs) were found, of which 118 and 137 BPTs were unique (UQ) to host groups. The existing database comprised of 295 enterococci UQ-BPTs and 273 E. coli UQ-BPTs from 10 host groups. The representativeness and the stability of the existing database were assessed by comparing with isolates that were external to the database. In all, 197 enterococci BPTs and 179 E. coli BPTs were found in water samples. The existing database was able to identify 62.4% of enterococci BPTs and 64.8% of E. coli BPTs as human and animal sources. The results indicated that a representative database developed from a catchment can be used to predict the sources of faecal contamination in another catchment with similar landuse features within the same geographical area. However, the representativeness and the stability of the database should be evaluated prior to its application in such investigation.     

Various physico-chemical stress factors cause prophage induction in Nitrosospira multiformis 25196- an ammonia oxidizing bacteria

Bacteriophages are virsues that infect bacteria and contribute significant changes in the overall bacterial community. Prophages are formed when temperate bacteriophages integrate their DNA into the bacterial chromosome during the lysogenic cycle of the phage infection to bacteria. The prophage (phage DNA integrated into bacterial genome) on the bacterial genome remains dormant, but can cause cell lysis under certain environmental conditions. This research examined the effect of various environmental stress factors on the ammonia oxidation and prophage induction in a model ammonia oxidizing bacteria Nitrosospira multiformis ATCC 25196. The factors included in the study were pH, temperature, organic carbon (COD), the presence of heavy metal in the form of chromium (VI) and the toxicity as potassium cyanide (KCN). The selected environmental factors are commonly encountered in wastewater treatment processes, where ammonia oxidizing bacteria play a pivotal role of converting ammonia into nitrite. All the factors could induce prophage from N. multiformis demonstrating that cell lysis due to prophage induction could be an important mechanism contributing to the frequent upset in ammonia oxidation efficiency in full scale treatment plants. Among the stress factors considered, pH in the acidic range was the most detrimental to the nitrification efficiency by N. multiformis. The number of virus like particles (VLPs) increased by 2.3E+10 at pH 5 in 5 h under acidic pH conditions. The corresponding increases in VLPs at pH values of 7 and 8 were 9.67E+9 and 1.57E+10 in 5 h respectively. Cell lysis due to stress resulting in phage induction seemed the primary reason for deteriorated ammonia oxidation by N. multiformis at lower concentrations of Cr (VI) and potassium cyanide. However, direct killing of N. multiformis due to the binding of Cr (VI) and potassium cyanide with cell protein as demonstrated in the literature at higher concentrations of these toxic compounds was the primary mechanism of cell lysis of N. multiformis. Organics represented by the chemical oxygen demand (COD) did not have any effect on the phage induction in N. multiformis. This AOB remained dormant at low temperature (4 °C) without any phage induction. Significant decrease in the number of live N. multiformis cells with a corresponding increase in the number of VLPs was recorded when the temperature was increased to 35 °C. Death of N. multiformis at 45 °C was attributed to the destruction of cell wall rather than to the phage induction.     

Detection of virulence genes in Escherichia coli of an existing metabolic fingerprint database to predict the sources of pathogenic E. coli in surface waters

A collection of 366 Escherichia coli strains from 10 host groups and surface waters were tested for the presence of 15 virulence genes associated with strains causing intestinal and extra-intestinal infections. The virulence genes included eaeA, VT1, 2 and 2e, LT1, ST1 and 2, Einv gene, EAgg gene, CNF1 and 2, papC, O111 and O157 side chain LPS. Of the 262 strains obtained from nine different hosts, 39 (15%) carried one or more of these virulence genes. These included six strains from humans, two from horses, eight from dogs, two from ducks, five from cattle, seven from chickens, four from pigs, two from sheep and three from deer. Of the remaining 104 strains obtained from water samples, 10 (10%) also carried one or more of the tested virulence genes. Of these, six had identical biochemical phenotypes (BPTs) to strains isolated from humans (two strains), dogs (two strains), chickens (one strain) and sheep (one strain) with 4 BPTs also carrying same virulence genes. Our results indicate that the sources of clinically important E. coli strains found in surface waters due to faecal contamination can be predicted by using a combination of biochemical fingerprinting method and the detection of virulence genes. From the public health point of view this information will be of great importance for evaluating the risk associated with public use of the catchment.     

Use of static Quantitative Microbial Risk Assessment to determine pathogen risks in an unconfined carbonate aquifer used for Managed Aquifer Recharge

Managed Aquifer Recharge (MAR) is becoming a mechanism used for recycling treated wastewater and captured urban stormwater and is being used as a treatment barrier to remove contaminants such as pathogens from the recharged water. There is still a need, however, to demonstrate the effectiveness of MAR to reduce any residual risk of pathogens in the recovered water. A MAR research site recharging secondary treated wastewater in an unconfined carbonate aquifer was used in conjunction with a static Quantitative Microbial Risk Assessment (QMRA) to assess the microbial pathogen risk in the recovered water following infiltration and aquifer passage. The research involved undertaking a detailed hydrogeological assessment of the aquifer at the MAR site and determining the decay rates of reference pathogens from an in-situ decay study. These variables along with literature data were then used in the static QMRA which demonstrated that the recovered water at this site did not meet the Australian Guidelines for recycled water when used for differing private green space irrigation scenarios. The results also confirmed the importance of obtaining local hydrogeological data as local heterogeneity can influence of residence time in the aquifer which, in turn, influences the outcomes. The research demonstrated that a static QMRA can be used to determine the residual risk from pathogens in recovered water and showed that it can be a valuable tool in the preliminary design and operation of MAR systems and the incorporation of complementary engineered treatment processes to ensure that there is acceptable health risk from the recovered water.