|Other Abstract||Cyanobacterial blooms are a major worldwide environmental problem. Among the bloom-forming cyanobacteria, Anabaena is one of the common planktonic genera in freshwater bodies. Like many other cyanobacteria, a number of Anabaena species can produce toxins or release odors into drinking and recreational waters. Thus, the surface blooms of Anabaena pose potential risks to aquatic environments and human health. In China, Anabaena is one of the most prevalent planktonic cyanobacterial genera, and it is frequently found in lakes Dianchi and Erhai. But few researches on blooming Anabaena and its toxicity in China has been reported. In the present study, the species composition, ecophysiology and toxicology of Anabaena spp. in lakes Dianchi and Erhai were investigated. The main objective of this study was to elucidate the distribution of Anabaena species and the mechanism of phytoplankton succession, to detect the toxin and to evaluate the toxic effects of Anabaena on mammal. The main results are summarized as follows:
1) Composition of phytoplankton and distribution of physicochemical factors in Lake Dianchi in years 2002 and 2003 were investigated to present the existence of Anabaena and analyze the correlation of phytoplankton species with the environmental factors. Results showed that Lake Dianchi has been in a hypereutrophic status; Microcystis is the dominant species in most months of the year, and its population dominance held more than 90%; Anabaena occurred heavily in winter and spring, and its population dominance can account for about 30％; SD was higher and Chla was lower during the occurrence of Anabaena or Aphanizomenon bloom in the cooling season.
2) Field studies were conducted in Lake Erhai from June to October of the year 2006. The investigation on phytoplankton composition showed that the cyanobacterial blooms occurred in the whole investigated period, phytoplankton abundance reached to 107 cells/L and more, and dominant species of cyanobacterial blooms displayed an obvious succession from Anabaena to Microcystis. Analysis of water quality showed that Lake Erhai remained at a mesotrophic status presently, but it was experiencing a transition from the mesotrophic to eutrophic condition; the trophic level was higher in August with lower phytoplankton diversity, and the biomass of phytoplankton (Chl a) was the most in this month. In addition, it was suggested that TP and T were the important factors to drive the succession between Anabaena bloom and Microcystis bloom by using CCA analysis.
3) Annual variations of phytoplankton composition and water quality in Lake Erhai were investigated. Results showed that the trophic status of Lake Erhai was probably transferring from mesotrophic to eutrophic, and the trophic levels were higher in the summer and autumn than in the winter and spring. TN in higher level was the primary pollution. Obvious process of phytoplankton succession was, with the dominant species one after the other as the followings: Diatom from January to March; Aphanizomenon in May and June; Anabaena in July and August; Microcystis from September to November. Correlative analysis showed that TN, TP and T were the important factors in changing the phytoplankton abundance, and TP and T were more important for phytoplankton succession.
4) Morphological characteristics and phylogenetic relationships of seven Anabaena strains isolated and purified from the two plateau lakes Dianchi and Erhai were investigated. Phylogenetic relationships were determined with the 16S rRNA gene tree which was calculated and constructed by using the neighbour joining algorithm. The seven strains were reasonably divided into three groups with the morphology of akinetes and the width of trichomes. The 16S rRNA gene tree also resulted in three groups of the seven strains, and the phylogenetic relationships of the studied strains were in agreement with the taxonomic classification. Strains EH-2, EH-3 and 4 were classified as Anabaena circinalis or Anabaena crassa. Strains DC-1, DC-2 and EH-1 were identified as Anabaena flos-aquae. Strain DC-3 was different from the other test strains and determined as Anabaena cylindrica.
5) In the study, all strains tested grew well, but the growth was different among strains. The obvious correction was found between growth and photosynthetic pigments. In addition, Yield, rETRmax and Alpha were all high to hold effective photosynthesis. This study provided some basic information and indicated significant ecological and physiological meanings for further researches.
6) In this study, the dominant species from the bloom of Lake Erhai, Anabaena circinalis, was isolated and purified. The growth and physiological characteristics of an Anabaena circinalis strain under different environmental conditions were studied to evaluate the influence of environmental factors on Anabaena bloom formation. Results showed that the Anabaena circinalis strain had best growth at temperature of 25℃, light intensity of 30 μE/ m﹒s, pH of 8.0-10.0, nitrogen 1.5mmol/L and phosphorus 12 µmol/L. The activities of nitrate reductase (NR) and alkaline phophatase (APA), which indicate metabolism of nitrogen and phosphorus, were measured with different concentrations of nitrogen and phosphorus in the medium. NR activity of the Anabaena circinalis strain increased with the raising concentration of nitrogen when nitrogen supply level was low (0 mmol/L-0.36 mmol/L). The NR activity was related to the growth under rich nitrogen supply (0.36 mmol/L-6 mmol/L), i.e. the better, the strain’s growth and the higher, the NR activity. APA activity of the Anabaena circinalis strain largely depended on the phosphorus concentration. APA activity was lower with more phosphorus supply. And Nitrogen did not play an important role in the activity change of APA when phosphorus supply was rich. In addition, it was observed that the Anabaena circinalis strain can grow with low concentration of nitrogen or phosphorus. This may be related to special regulatory mechanism of nitrogen fixing and phosphorus utilizing of Anabaena.
7) The toxicological effects induced by non-centrifuged extracts and centrifuged extracts from the cultures of the isolated strains were assessed with mouse by intraperitoneal (i.p.) injection. Results showed that the seven strains isolated from lakes Dianchi and Erhai all presented toxic effects on mouse, and LPS was proved to increase the toxic effects on mouse for cellular toxins.
8) In the present study, the mouse toxicity of an Anabaena flos-aquae strain isolated from Lake Dianchi was further investigated. The lesions of ultrastructur in liver, kidney, lung, spleen, intestines and myocardiocyte were characterized. Significant alterations were found in the serum biochemical parameters indicating changes in the hepatic functions, renal functions, and immune functions. Histopathological observations with haematoxilin and eosin (HE) stain also showed that severe lesions were in the livers, kidneys and lungs of the mice. The alterations of biochemical parameters and the severities of histological lesions were in a dose-dependent manner. Based on the biochemical and histological studies, and the above mentioned biological toxicity testing, this research firstly showed the presence of toxin producing Anabaena species in Lake Dianchi and the toxic effects of its crude extracts on mammals.|