Cadmium toxicity to Microcystis aeruginosa PCC 7806 and its microcystin-lacking mutant.

The adverse effects of microcystin (MC) produced by cyanobacteria have drawn considerable attention from the public. Yet it remains unclear whether MC confers any benefits to the cyanobacteria themselves. One suggested function of MC is complexation, which may influence the bioaccumulation and toxicity of trace metals. To test this hypothesis, we examined Cd toxicity to wild-type Microcystis aeruginosa PCC 7806 (WT) and its MC-lacking mutant (MT) under nutrient-enriched (+NP), phosphorus-limited (-P), and nitrogen-limited (-N) conditions. The accumulation of Cd and the biochemical parameters associated with its detoxification [total phosphorus (TP), inorganic polyphosphate (Poly-P), and glutathione (GSH) in the cells as well as intra- and extra-cellular carbohydrates] were quantified. Although the -P cyanobacteria accumulated less Cd than their +NP and -N counterparts, the different nutrient-conditioned cyanobacteria were similarly inhibited by similar free ion concentration of Cd in the medium ([Cd2+]F). Such good toxicity predictability of [Cd2+]F was ascribed to the synchronous decrease in the intracellular concentrations of Cd and TP. Nevertheless, Cd toxicity was still determined by the intracellular Cd to phosphorus ratio (Cd/P), in accordance with what has been reported in the literature. On the other hand, the concentrations of TP, Poly-P, and carbohydrates went up, but GSH concentration dropped down with the enhancement of [Cd2+]F, indicating their association with Cd detoxification. Although the inactivation of MC peptide synthetase gene had some nutrient and Cd concentration dependent effects on the parameters above, both cyanobacterial strains showed the same Cd accumulation ability and displayed similar Cd sensitivity. These results suggest that MC cannot affect metal toxicity either by regulating metal accumulation or by altering the detoxification ability of the cyanobacteria. Other possible functions of MC need to be further investigated.

Effects of nitrogen and phosphorus on arsenite accumulation, oxidation, and toxicity in Chlamydomonas reinhardtii.

We studied arsenite (iAs(III)) accumulation, oxidation, and toxicity in the freshwater green alga Chlamydomonas reinhardtii under nutrient-enriched (+NP), phosphorus-limited (-P), and nitrogen-limited (-N) conditions. The -P alga (55.1 µM) had a Michaelis constant (Kd) for uptake approximately one tenth of the +NP (419 µM) and -N (501 µM) cells, indicating iAs(III) uptake inhibition by extracellular phosphate. This conclusion was supported by the hyperbolic reduction in iAs(III) uptake rate (V) from 9.2 to 0.8 µmol/g-dw/h when the extracellular phosphate concentration went up from 0 to 250 µM. The maximal iAs(III) uptake rate (Vmax) of the -N alga (24.3 µmol/g-dw/h) was twice as much as that of the +NP (12 µmol/g-dw/h) and -P (8.1 µmol/g-dw/h) cells. It implies that more arsenic transporters were synthesized under the -N condition. Once accumulated, iAs(III) was oxidized and a higher proportion of arsenate (iAs(V)) was observed at lower [As]dis or under nutrient-limited conditions. Nevertheless, iAs(III) oxidation mainly occurred outside the cells with the extent of oxidation reciprocal to [As]dis. Based on the logistic modeling of the concentration-response curves in the +NP, -P, and -N toxicity tests, iAs(III) had an [As]dis-based EC50 of 1763, 13.1, and 1208 µM and an intracellular arsenic concentration based EC50 of 35.6, 28.8, and 195 µmol/g-dw, respectively. Higher iAs(III) toxicity to the -P cells occured because of their increased iAs(III) accumulation, whereas the underlying mechanisms why the -N alga was more tolerant need to be further revealed. Overall, both N and P had remarkable effects on the behavior and effects of iAs(III), which cannot be disregarded in the biogeochemical cycling research of arsenic.

[Characteristics and influencing factors of phosphorus adsorption on sediment in Lake Taihu and Lake Hulun].

Lake Taihu and Lake Hulun in southern and northern China were selected for the investigation of differences in the characteristics and influencing factors of phosphorus (P) adsorption on the sediments of these two lakes by laboratory simulation experiment. The results showed that: (1) The P adsorption capacity of sediment in Lake Taihu was much higher than that in Lake Hulun, and the maximum adsorption capability for the sediments in these two lakes was 1428.57 mg x kg(-1) and 56.81 mg x kg(-1), respectively; (2) Compared with the sediments in Lake Hulun, the particle diameters from sediments in Lake Taihu were much smaller, and their specific surface areas were much larger, so their P adsorption capacity were much higher; (3) The equilibrium adsorbed amounts in these two lakes were correlated with the total amounts of Al, Fe, Mn (P < 0.05) and significantly correlated to the contents of active Al, Fe, Si, Mn (P < 0.01). However, there was significant negative correlation between the equilibrium adsorbed amounts and total Si contents for the sediments in these two lakes (P < 0.01); (4) The P adsorption capacity decreased as pH values in the overlying water increased, and the increase of pH values affected the adsorption behavior of phosphorus on the sediments in Lake Taihu much more obviously. Therefore, the characteristics of sediments and the variations of pH values in the overlying water affected the adsorption behavior of P on sediments in lake water body.

Seasonal variation of phytoplankton nutrient limitation in Lake Taihu, China: a monthly study from year 2011 to 2012.

Lake Taihu is the third largest freshwater lake in China with severe eutrophication issues. However, it remains ambiguous how its phytoplankton growth is limited by various nutrients in different seasons. A series of bottle-enrichment assays in Meiliang Bay was thus performed once a month from July, 2011 to June, 2012 in the present study. The initial chlorophyll a concentration and phytoplankton cell density ranged from 4.70 to 34.6 µg/l and from 1.25×10(6) to 6.72×10(8) cells/l with three peaks in July, November, and March. Although Cyanophyta was dominant (30.9-99.2 percent) in most cases, other phyla like Chlorophyta, Bacillariophyta, and Cryptophyta could account for as much as 69.1 percent of total phytoplankton in cold seasons. The microcystin-LR content in the particulate phase followed a similar seasonal pattern as Cyanophyta. It further went up exponentially with the proportion of cyanobacteria in phytoplankton suggesting more toxigenic species and (or) upregulated microcystin synthesis when the contribution of Cyanophyta was enhanced. On the other hand, the dissolved concentrations of various nitrogen and phosphorus species reached their maxima in late spring and autumn, respectively. According to its growth response to nutrient addition, phytoplankton in Meiliang Bay was restricted by nitrogen in August, October, and November. No nutrient limitation occurred in July, September, and April, whereas phosphorus deficiency prevailed in the other months. Overall, nutrient limitation in Lake Taihu and possibly other aquatic ecosystems worldwide may be more dynamic than what we thought before, which should be considered to eliminate eutrophication.

[Analysis on the forms and release potential of nutrients in sediments from lakes in the West Jiangsu Province].

To probe the effect of nitrogen and phosphorus (P) in sediment on aqueous nutrient concentrations, forms of phosphorus, sediment water interface diffusion fluxes and phosphorus release and adsorption of 10 lakes in the west Jiangsu province were studied. Sediments were sequentially extracted by the modified Ruttenberg's method, and the result showed that inorganic phosphorus in sediments of the lakes was the major fraction of total phosphorus, but the proportion of bioavailable phosphorus was relatively low. Compared with the lakes in northern Jiangsu, the Fe-P was relatively high in sediments of lakes in southern Jiangsu. The diffusion fluxes of lakes were mainly based on NH4(+) -N diffusion in the west Jiangsu, and ions of most of lakes diffused from overlying water to sediment, NO3(-) -N diffused from sediment to overlying water in lakes except for Lake Hongze, Lake Shijiu and Lake Xuanwu. The condition leaning to acidic (pH < 4) or alkaline (pH > 10) was beneficial to phosphorus release, and except for Lake Xuanwu, the least amount of phosphorus released form sediment was higher in acidic condition than that in alkaline condition. The amount of phosphorus adsorption was increasing with temperature increase. Considering the phosphorus adsorption on native phosphorus polluted (NAP-polluted) sediments. All data of P adsorption on sediment samples collected from 7 lakes were fitted to the modified Langmuir model. And the values of the maximum adsorption capacity (Q(m)), phosphorus binding constant (k), native adsorbed exchangeable phosphorus (Q(NAP)), were subsequently obtained. The results showed that the adsorption amount of P in Lake Hongze and Lake Xuanwu was relatively low, which were liable to release P from sediments.

[Regional heterogeneity of lake eutrophication effects in China].

Although biomass of algae (Chl-a) were in variant levels between different lake regions in China under the same nutrients conditions, it demonstrated that efficiencies of TN/TP used by algae had regional differences. In order to clarify the differences, curve estimation in SPSS was used to analyze the linear relationship between Ig Chl-a and Ig TN/lg TP. The slopes of these linear equations were identified as the efficiencies of TN/TP used by algae. The slopes of linear equations from Mengxin Plateau, Yungui Plateau, Northeast Mountain-Plain, lower reach of Yangtze River Plain and North Plain were 1.002, 0.817, 0.761, 0.545, 0.250, orderly. The efficiencies of TN used by algae ranged from the highest to the lowest were lower reach of Yangtze River Plain, Yungui Plateau, North Plain, Northeast Mountain-Plain, Mengxin Plateau, and the slopes of linear equations were 1.401, 1.058, 0.447, 0.239, 0. 099, respectively. Consequently, in Northeast Mountain-Plain, Mengxin Plateau the efficiencies of TP used by algae were higher than those of TN, and in Yangtze River Plain, Yungui Plateau, North Plain, the efficiencies of TN used by algae were higher than those of TP. On the other hand, in order to describe the effects of algae on transparency in different lakes, the relationships between Chl-a and SD were analyzed. The results showed that in Yungui Plateau the effect of algae on transparency was the most obvious as the variation of SD explained by Chl-a was the highest, and Northeast Mountain-Plain, Mengxin Plateau and North Plain followed. However, in lower reach of Yangtze River Plain, the relationship between Chl-a and SD was not significant.

Accumulation kinetics of arsenic in Daphnia magna under different phosphorus and food density regimes.

In the present study, the dissolved uptake, dietary assimilation, and elimination of arsenic (initially added as arsenate) in the freshwater cladoceran Daphnia magna were examined. A biphasic correlation between the arsenic uptake rate and its ambient concentration, as well as a two-saturation-site arsenic uptake competition with phosphate was observed. The calculated uptake rate constant, as influenced by the ambient phosphorus concentration, ranged from 0.035 to 0.35 L/g/d. Food concentration substantially decreased (by 23.2-64.4%) the arsenic assimilation efficiency with the incipient limiting algal food concentration of 3.86 mg/L dry weight. Arsenic assimilation by the daphnids was independent of their own phosphorus status, but was lower when their algal diet was phosphorus-limited and thus contained a higher proportion of arsenite due to the enhanced biotransformation. Arsenic efflux rate constant ranged from 0.34 to 0.44 d(-1) with increased food concentration slightly facilitating its loss. Excretion, accounting for 51.3 to 60.6% of total loss, was the dominant pathway for arsenic elimination with a remarkable contribution from offspring production (24.7-29.8%), whereas molting (3.64-4.05%) and egestion (7.9-11.9%) had minor roles only. According to the well-established biokinetic model, dietary assimilation was predicted to be the main pathway for arsenic bioaccumulation in the daphnids, and arsenic has a great potential to be biodiminished along the food chain.

[Integration and expression of polyphosphate kinase gene in Pseudomonas putida].

In order to construct high accumulating-phosphate microorganism, the ppk gene from E. coli was inserted to the broad-host-range plasmid pBBR1MCS-2 to form plasmid pBBR1MCS-2-ppk. The complete ppk gene with promoter and terminator sequences from pBBR1MCS-2-ppk was then cloned and inserted to suicide plasmid pUTmini-Tn5 to form plasmid pUTmini-Tn5-ppk, which was transformed into Pseudomonas putida KT2440 by triparental conjugation. Finally, ppk gene was integrated into the chromosomal DNA of KT2440. The results of RT-PCR showed that the selected genetically engineered bacterium KT2440-PPK expressed ppk efficiently, while KT2440 as control expressed weakly. The results of artificial wastewater treatment showed after 1h inoculation, the concentration of poly-phosphate in KT2440-PPK came to the maximum approximately 3.05 mg/g, which was 15 times higher than that in KT2440 at the same experimental condition. And KT2440-PPK can remove more than 90% phosphate in artificial wastewater.

[Effects of phosphorus sources of different forms on phosphorus metabolism of Microcystis aeruginosa and adhesive Pseudomonas sp].

The effects of different forms of phosphorus sources on the phosphorus metabolism of Microcystis aeruginosa and attached Pseudomonas sp. were investigated after four substance of different phosphrus forms: NaH2PO4, Natrium-beta-glycerophosphate (NaGly), Ca3(PO4)2 and lecithin were added in the non-phosphorus MA culture. The growth of Microcystis aeruginosa, total phosphorus, alkaline phosphatase activity in water and total phosphorus in Microcystis aeruginosa were measured every day or every two days. Results show that attached Pseudomonas sp. could accelerate the growth of Microcystis aeruginosa, and transform some phosphrus forms which could not be assimilated very easily by Microcystis aeruginosa to some forms as phosphate to utilize for Microcystis aeruginosa. Alkaline phosphatase plays an important role in the utilization of large molecular organic phosphorus by Microcystis aeruginosa and attached Pseudomonas sp.