Simultaneous increases of filter-feeding fish and bivalves are key for controlling cyanobacterial blooms in a shallow eutrophic lake.

Eutrophication and cyanobacterial blooms have severely affected many freshwater ecosystems. We studied the effects of filter-feeding fish and bivalves on algal populations using a mesocosm experiment and long-term monitoring data from Lake Taihu (China). The mesocosm study, comprised of a two-way factorial design with the clam Corbicula fluminea and the fish Aristichthys nobilis at three biomass levels, resulted in lower chlorophyll a (Chl a) in high fish treatments, but no significant differences in the low and medium fish treatments. Chl a also decreased with an increase in clam biomass in the high fish treatments. Moreover, filter-feeding fish resulted in a decrease in algal sizes (e.g., the colony size of Microcystis aeruginosa was inversely related to fish biomass) which likely increased the filter-feeding efficiency of bivalves. Biomass of filter-feeding fish was found to be a key factor driving the synergistic effects of filter-feeding fish and bivalves in waters dominated by Microcystis colonies. Long-term monitoring revealed increasing trends in Chl a concentration, total fish catch per unit effort (TF-CPUE), and filter-feeding fish (FF-CPUE), and slightly decreasing trends in bivalve biomass and nitrogen to phosphorus ratios (N:P) from 2006 to 2016. Bivalve biomass and N:P were negatively correlated with Chl a, while FF-CPUE was not significantly related to Chl a. The current filter-feeding fish biomass in Lake Taihu is estimated to be too low to drive synergistic algal control effects together with bivalves. Furthermore, the lack of filter feeders in Lake Taihu may lead to top-down control by predators that cannot counteract the bottom-up effects of nutrients on phytoplankton. Collectively, these long-term monitoring and experimental data support the combined use filter-feeding fish and bivalves for managing cyanobacteria blooms in Lake Taihu.

Top-down effects of filter-feeding fish and bivalves moderate bottom-up effects of nutrients on phytoplankton in subtropical shallow lakes: An outdoor mesocosm study.

Biomanipulation has been widely used in the ecological restoration of eutrophic lakes for decades. However, biomanipulation is prone to failure if external nutrient loads are not reduced. In order to explore the importance of filter-feeding fish and bivalves on algal control, an outdoor mesocosm experiment was conducted using different nutrient concentrations. Four treatments simulating daily loads of nutrients in Lake Taihu were studied: current, two times, and three times average daily loads of nutrients with both fish (Aristichthys nobilis) and Asian clam (Corbicula fluminea) and as a control current daily loads without fish or bivalves. Results showed that stocking of filter-feeding fish and bivalves (80 g m-3 bighead carp; 200 g cm-2 clams) at two times daily nutrient loads could effectively control water column Chl a concentrations and phytoplankton biomass. At higher nutrient concentrations (TN ≥ 260 µg L-1 d-1; TP ≥ 10 µg L-1 d-1), top-down control of filter-feeding fish and bivalves was less effective and bottom-up effects resulted in significant increases of Chl a concentration. Thus, as phytoplankton biomass in freshwater ecosystems is determined by both the top-down effects of predators and the bottom-up effects of nutrients, external loadings should be controlled when filter-feeding fish and bivalves are used for algal control to ensure the efficacy of biomanipulation.

Piscivore stocking significantly suppresses small fish but does not facilitate a clear-water state in subtropical shallow mesocosms: A biomanipulation experiment.

Biomanipulation by piscivore stocking has been widely used to combat eutrophication in north temperate lakes, but its applicability in warm lakes has not yet been well elucidated. Here, we used experimental mesocosms to test the effects of a native benthi-piscivore (snakehead, Channa argus Cantor) on water clarity under subtropical conditions where small omni-benthivorous fish like crucian carp (Carassius carassius L.) prevail. Our results showed that, despite of a great reduction of crucian carp biomass, snakehead stocking did not create a strong trophic cascade as neither (herbivorous) zooplankton biomass nor their grazing pressure, indicated by biomass ratio of (herbivorous) zooplankton to phytoplankton, changed significantly. Moreover, snakehead stocking significantly increased water non-algal turbidity as well as nutrient and chlorophyll-a concentrations, suggesting that these benthi-piscivores also disturbed sediments like crucian carp did. Our study showed that biomanipulation by stocking of snakehead does not facilitate clear-water state in warm shallow lakes, even on the short-term.

Reducing nutrient increases diatom biomass in a subtropical eutrophic lake, China-Do the ammonium concentration and nitrate to ammonium ratio play a role?

Response of aquatic organisms to eutrophication have been well reported, while less studies are available for the recovery of eutrophic lakes following a reduction in the external loading, especially for systems where nitrogen is reduced but the phosphorus concentration is maintained high due to internal loading. Diatoms are nitrate (NO3-N) opportunists but can also use ammonium (NH4-N). They may, therefore, be more sensitive to nitrogen reduction than other algae that typically prefer NH4-N. We document the variations of nutrients and diatoms in subtropical, eutrophic Lake Taihu over 28 yr during which a reduction of the external loading resulted from lake management. According to the results of change point analysis, data on environmental variables were divided into two periods (P1: 1992-2006; P2: 2007-2019) with two different seasons (WS: Winter-Spring; SA: Summer-Autumn), respectively. Compared with P1-WS, the concentration of NH4-N decreased significantly whereas NO3-N showed no significant change in P2-WS. In contrast, NH4-N concentrations were low and showed no significant changes in P1-SA and P2-SA and NO3-N decreased significantly in the latter period. Accordingly, NO3-N: NH4-N mass ratios in P1-SA and P2-WS were all significantly higher than those in P2-SA and P1-WS, respectively. The biomass of WS diatom increased significantly and the timing of the peak biomass shifted from P1-SA to P2-WS since 2007. The SEM analysis showed that NO3-N was retained as a statistically significant predictor for diatom biomass in P1-SA and significant effects of windspeed, zooplankton and NH4-N on diatom biomass in P2-WS. Windspeed and zooplankton have further changed the biomass of diatoms in the case of declining inorganic nitrogen. We conclude that the magnitude of vernal suppression or stimulation of diatom assemblages has increased, concomitant with the variations of NH4-N and NO3-N: NH4-N mass ratios. Diatoms response to NH4-N or NO3-N is apparently changing in response to water temperature in this eutrophic shallow lake. Thus, parallel reductions in external nitrogen loading, along with variations in dominant inorganic nitrogen, will stimulate the growth of diatom and therefore increase the total biomass of phytoplankton in still high internal phosphorus loading, which is should be regarded as a good sign of restoration measures.

Impacts of nutrient reduction on temporal ß-diversity of rotifers: A 19-year limnology case study on Lake Wuli, China.

There have been many studies on the effects of eutrophication on beta diversity (ß-diversity) of species assemblages. However, few studies have focused on the effects of nutrient reduction on ß-diversity and community structure, and long-time series analyses are particularly scarce. We conducted a 19-year case study on the impacts of management intervention on the temporal ß-diversity of aquatic grazers in a lake at the Yangtze River Basin. In our study, we compared the changes in temporal ß-diversity as well as its two components, nestedness and turnover, and the synchrony of the rotifer community after management intervention. Our results showed that while the abundance of some sensitive species increased, there was no trend in species richness. Moreover, both the seasonality and interannual stabilities of rotifer assemblages increased. The species synchrony decreased in both spring and summer after management intervention. We also found that management intervention significantly reduced nutrient concentrations but not water clarity and phytoplankton abundance. The total nitrogen (TN): total phosphorous (TP) ratio was reduced after management intervention, causing an increase in the abundance of cyanobacteria that may contribute to the increase of rotifer synchrony in autumn. Our results imply that stable environmental fluctuations after management intervention may increase temporal ß-diversity and stability of herbivorous assemblages. However, imbalanced changes in TN and TP after management intervention may weaken the top-down control of zooplankton on phytoplankton and slow down water clarity improvement.

Co-occurrence of multiple cyanotoxins and taste-and-odor compounds in the large eutrophic Lake Taihu, China: Dynamics, driving factors, and challenges for risk assessment.

Cyanobacterial blooms producing toxic metabolites occur frequently in freshwater, yet the environmental behaviors of complex cyanobacterial metabolites remain largely unknown. In this study, the seasonal and spatial variations of several classes of cyanotoxins (microcystins, cylindrospermopsins, saxitoxins) and taste-and-odor (T&O) compounds (ß-cyclocitral, ß-ionone, geosmin, 2-methylisoborneol) in Lake Taihu were simultaneously investigated for the first time. The total cyanotoxins were dominated by microcystins with concentrations highest in November (mean 2209 ng/L) and lowest in February (mean 48.7 ng/L). Cylindrospermopsins were abundant in May with the highest content of 622.8 ng/L. Saxitoxins only occurred in May (mean 19.2 ng/L) and November (mean 198.5 ng/L). Extracellular T&O compounds were most concentrated in August, the highest being extracellular ß-cyclocitral (mean 240.6 ng/L) followed by 2-methylisoborneol (mean 146.6 ng/L). Environment variables play conflicting roles in modulating the dynamics of different groups of cyanotoxins and T&O compounds. Total phosphorus (TP), total nitrogen (TN), chlorophyll-a and cyanobacteria density were important factors affecting the variation of total microcystins, ß-cyclocitral and ß-ionone concentrations. In contrast, total cylindrospermopsins, 2-methylisoborneol and geosmin concentrations were significantly influenced by water temperature and TP. There was a significant and linear relationship between microcystins and ß-cyclocitral/ß-ionone, while cylindrospermopsins were positively correlated with 2-methylisoborneol and geosmin. The perceptible odors may be good indicators for the existence of cyanotoxins. Hazard quotients revealed that potential human health risks from microcystins were high in August and November. Meanwhile, the risks from cylindrospermopsins were at moderate levels. Cylindrospermopsins and saxitoxins were first identified in this lake, suggesting that diverse cyanotoxins might co-occur more commonly than previously thought. Hence, the risks from other cyanotoxins beyond microcystins shouldn't be ignored. This study also highlights that the necessity for further assessing the combination effects of these complex metabolites.

Combining bivalve (Corbicula fluminea) and filter-feeding fish (Aristichthys nobilis) enhances the bioremediation effect of algae: An outdoor mesocosm study.

In outdoor mesocosms, we experimentally studied the effect of combining two native filter feeders, bighead carp (Aristichthys nobilis) and Asian clam (Corbicula fluminea), to control nuisance cyanobacterial blooms. Four treatments - clam-only, fish-only, combined and controls - were used. In combination, bighead carp (80 g m-3) and Asian clam (260 g m-2), had a remarkable controlling effect on phytoplankton biomass and improved water quality. By the end of the experiment, the chlorophyll a (Chl a) concentration of the combined group was below 5 µg L-1 compared with 36 µg L-1 in the clam-only treatments and 31 µg L-1 in the controls, whereas Chl a in the fish-only group had increased to 211 µg L-1. Large-sized algae (Microcystis sp.) dominated in the control group and the clam-only group, whereas small-sized algae (Actinastrum hantzschii and Clamydomonas globose) dominated in the fish-only group. The concentrations of total nitrogen (TN) and total phosphorus (TP) in the water column were significantly reduced by clam stocking, regardless of the presence of fish. A 24-h laboratory experiment was conducted to verify whether the excrements produced by clams filtering small-sized algae could be eaten by bighead carp. At the end of the experiment, the density of Scenedesmus obliquus had decreased significantly in the clam-present mesocosms, and the gut fullness index of bighead carp was significantly higher in the clam-present mesocosms than when clams were absent. Our results show that, under the selected experimental conditions and densities, water clarity improved when bighead carp and Asian clam occurred together, whereas Chl a concentrations and algae biomass increased in the fish-only mesocosms; in the clam-only mesocosms no significant effects were observed. We suggest that the combination of filter-feeding fish and clams may enhance water clarity and it may therefore potentially be a useful restoration tool.

Effects of juvenile crucian carp (Carassius carassius) removal on submerged macrophyte growth-implications for subtropical shallow lake restoration.

Rapid expansion of juvenile fish after biomanipulation can delay the successful restoration of submerged macrophytes, leading to a turbid water status in subtropical shallow lakes. Aimed to test the effects of direct removal of juvenile crucian carp (Carassius carassius) on water quality and growth of two submerged macrophytes Vallisneria natans and Hydrilla verticillate, a short-term outdoor mesocosm experiment was conducted in the Taihu Laboratory for Lake Ecosystem Research (TLLER). The results indicated that the concentrations of TN, TP, suspended solids, and chlorophyll a decreased significantly with increasing removal density of juvenile crucian carp, thus resulting in a clear status of the water. Additionally, the mean relative growth rate of V. natans and H. verticillata in the low- and high-density removal treatments were higher than that in the controls. Moreover, the removal of juvenile crucian carp also significantly increased the stem length of V. natans, but no significant effect on that of H. verticillata. Meanwhile, the total number of V. natans and H. verticillata in the low- and high-density removal treatments was higher than that in the controls, but all of H. verticillata were lower than the initials. Our results indicated that removing juvenile crucian carp could improve the water quality, increasing relative growth rate, height, and reproduction of V. natans, and improving the survival rate of H. verticillata. The promotion of fish removal on the V. natans growth was greater than H. verticillata. The results also implied that it was necessary to continuously remove the juvenile benthivorous fish several times for restoring the submerged macrophytes in shallow lakes.

[Seasonal Difference in Water Quality Between Lake and Inflow/Outflow Rivers of Lake Taihu, China].

The seasonal and spatial variation of external nutrient loading from rivers and their impact on lake water quality were analyzed in Lake Taihu, China, using the monthly monitoring data from 16 major inflow/outflow rivers and 32 observation sites in the lake. The results showed:① The average monthly values of total nitrogen (TN), dissolved total nitrogen (DTN), total phosphorus (TP), and dissolved total phosphorus (DTP) in rivers were all higher than the corresponding areas in the lake. Significant positive correlations were found between nutrient concentrations in the inflow rivers and the corresponding areas in the lake, indicating the pronounced impact of external loading on lake water. ② Remarkable seasonal variations of nutrient concentration were found both in the rivers and in the lake. The highest TN and TP concentrations in inflow rivers were 4.82 mg·L-1 (March) and 0.218 mg·L-1 (December), while the highest TN and TP concentrations in the lake were 4.13 mg·L-1 and 0.255 mg·L-1 in July. ③ Extreme rainfall events could decrease the nutrient concentration in the rivers in the short-term, but finally would increase the external loading of nutrients, and indicated disadvantages for the restoration of Lake Taihu. Our study concluded that inflow pollution showed an obvious "shaping effect" on the seasonal and spatial distribution of water quality indicators in large and shallow lakes. Additionally, the self-purification ability of lakes, wind-induced accumulation and migration of algae, as well as the sediment resuspension under the prevailing winds in different seasons, all have vital effects on nutrient concentrations and their spatial-temporal variations.

[Insertion efficiency of Tgf2 transposon in the genome of Megalo-brama amblycephala].

To study insertion efficiency of goldfish Tgf2 transposon in the genome of Megalobrama amblycephala, we built Tgf2-Mlyz2-RFP donor plasmid with goldfish Tgf2 transposon left and right arms, and then co-injected with goldfish Tgf2 transposase mRNA into the 1-2 cell stage fertilized eggs of M. amblycephala. In 30 d- and 180 d-stage larval, RFP fluorescence can be observed in back and side muscle of the fish. The rate of RFP fluorescence expression was 48.1%. In adult fish, PCR results demonstrated that integration efficiency of goldfish Tgf2 transposition system was 31.5% in M. am-blycephala genome. RT-PCR analysis showed that RFP RNAs were highly transcribed among all the 12 tissues in three transgenic fishes, while it could be highly detected only in muscle, skin, and kidney in another two individuals. Our results suggested that RFP expression in tissues vaied among different M. amblycephala. By means of the inverse PCR, the copy numbers of Tgf2 transposon were at least 2 in transgenic M. amblycephala. The average copy number of each fish was about 5. Over 50% of flanking sequences at the insertion site have homologous sequence in other vertebrate species. Our data suggest that goldfish Tgf2 transposon can efficiently mediate gene insertion in M. amblycephala, which could been used in transgene and gene trap in M. amblycephala.

Goldfish transposase Tgf2 presumably from recent horizontal transfer is active.

Hobo/Activator/Tam3 (hAT) superfamily transposons occur in plants and animals and play a role in genomic evolution. Certain hAT transposons are active and have been developed as incisive genetic tools. Active vertebrate elements are rarely discovered; however, Tgf2 transposon was recently discovered in goldfish (Carassius auratus). Here, we found that the endogenous Tgf2 element can transpose in goldfish genome. Seven different goldfish mRNA transcripts, encoding three lengths of Tgf2 transposase, were identified. Tgf2 transposase mRNA was detected in goldfish embryos, mainly in epithelial cells; levels were high in ovaries and mature eggs and in all adult tissues tested. Endogenous Tgf2 transposase mRNA is active in mature eggs and can mediate high rates of transposition (>30%) when injected with donor plasmids harboring a Tgf2 cis-element. When donor plasmid was coinjected with capped Tgf2 transposase mRNA, the insertion rate reached >90% at 1 yr. Nonautonomous copies of the Tgf2 transposon with large-fragment deletions and low levels of point mutations were also detected in common goldfish. Phylogenetic analysis indicates the taxonomic distribution of Tgf2 in goldfish is not due to vertical inheritance. We propose that the goldfish Tgf2 transposon originated by recent horizontal transfer and maintains a highly native activity.

HIF-1alpha and -2alpha genes in a hypoxia-sensitive teleost species Megalobrama amblycephala: cDNA cloning, expression and different responses to hypoxia.

Transcriptional responses to hypoxia are primarily mediated by hypoxia-inducible factor (HIF) alpha subunits, i.e. HIF-1alpha, -2alpha and -3alpha. In fish, the molecular constructions, expression characteristics and hypoxic regulation of HIF-alpha subunits are still not well known. In this study, we identified the HIF-1alpha and HIF-2alpha full-length cDNAs in a hypoxia-sensitive fish species Wuchang bream, Megalobrama amblycephala. The whole length of HIF-1alpha cDNA was 3,815bp, consisting of an open reading frame (ORF) encoding 774 amino acid (aa) residues. The HIF-2alpha cDNA totaled 3,121-bp including an 835-aa ORF. The Wuchang bream HIF-1alpha and HIF-2alpha subunits were structurally similar in the DNA-binding and dimerization domains, but differed in the transactivation domain. In adult fish, both HIF-1alpha and HIF-2alpha mRNAs were detected in different tissues under normoxic conditions. HIF-1alpha mRNA was highly expressed in the liver, gill and testis, whereas HIF-2alpha mRNA was abundantly expressed in most of the Wuchang bream tissues. Both HIF-1alpha and HIF-2alpha mRNAs were detected in all stages of embryogenesis and expressed in a ubiquitous pattern. In contrast to HIF-1alpha, the mRNA levels of HIF-2alpha fluctuated in different stages, with higher expression in the zygote, 8-, 28-, 48- and 52-hr post fertilization (hpf) embryos. During hypoxic treatment, the mRNA levels of HIF-2alpha were significantly (p<0.01) up-regulated to 910% in the liver and 320% in the kidney, whereas no significant changes of HIF-1alpha mRNA were observed in the corresponding tissues. These results suggest that the Wuchang bream HIF-1alpha and -2alpha would be involved in different physiological functions under normoxia and hypoxia situations.