Studies On The Growth-inhibitory Effects On HAB Algae In East China Sea Using Biological Or Chemical Means

In the last few years, algae blooms occur frequently in East China Sea and it is imperative to seek more applicable control strategies to prevent the occurrence and mitigate the harmful effects of HABs. The present dissertation aims to study the algicidal activity of the bacterial metabolites with the advantages of low toxicity, high biodegradability and ecological acceptability, to screen algicidal bacteria and study the algicidal activity of the bacteria screened. The growth-inhibitory effects of rhamnolipid biosurfactants and phenazine pigments produced by Pseudomonas aeruginosa O-2-2 on some common HAB algae in East China Sea were studied for the first time. Some algicidal bacteria were screened from natural seawater in East China Sea during the occurrence of Prorocentrum donghaiense blooms and the algicidal activity of the bacteria screened was also studied. Moreover, the growth-inhibitory effects of some new types of synthetic surfactants on the HAB species were also investigated systematically with some common Dinoflagellates and Diatoms in East China Sea as experiment materials. A train of study results are obtained which can definitely provide valuable scientific evidence, theoretical foundation and experimental basis for the prevention and control of HABs in East China Sea by biological or chemical means. I . The growth-inhibitory effects and the lytic effects of rhamnolipids produced by P. aeruginosa O-2-2 on some common Dinoflagellates and Diatom in East China Sea were first investigated, the algicidal mechanism, the mechanism of the selective algicidal activity of rhamnolipids on the algae were also studied in detail.1. Through the thorough simulation experiments, it was first found that the growth of P. donghaiense, Pseudo-nitzschia pungens and Chaetoceros Didymus was inhibited completely at lower concentration (3.0 or 4.0mg·L^-1) of rhamnolipids produced by P. aeruginosa O-2-2. When the concentration arrived at 7.0 or 10.0 mg·L^-1, the growth of Skeletonema costatuma, Chaetoceros curvisetus, Chaetoceros Delicatula and Nitzschica closterium was also inhibited completely. In addition, the growth of Heterosigmaakashiwo was also inhibited markedly in the presence of rhamnolipids. Furthermore, it was observed that the rhamnolipids at higher concentration had a substantial lytic effect on the HAB species. Under the same experiment conditions, the inhibitory effects of the rhamnolipids on the growth of green algae were negligible or could not persist. Accordingly, the rhamnolipids have a species-selective growth-inhibiting effect on some HAB species.2. Considering the changes in the cell shape and especially ultrastructure induced by rhamnolipid treatment, the inhibiting-effect on the growth and especially cell lysis may be mainly attributable to the amphiphilic surfacitivity of the rhamnolipids. The rhamnolipid molecules can cohere on the surface of phospholipids bimolecular layer of the algae due to their lipophilic moiety, thus enhancing the cell permeability so that rhamnolipid molecules can easily intrude into the cell. With increasing amounts of rhamnolipid inside the cell, the ultrastructure (including nuclear envelope, mitochondrial membrane, Golgi apparatus and endoplasmic reticulum) is destroyed, with the result that the cell disintegrates totally.3. The analysis of the correlation between the inhibitory effect of the rhamnolipids on the algae and the fatty acid composition of the algae tested indicates that the differences in the fatty acid composition, especially Hie proportion of polyunsaturated fatty acids (PUFAs) of the biological membranes of the algae are responsible for the species-specific responses to rhamnolrpids. Namely, the growth-inhibitory effect of rhamnolipids on the algae increases with the decreasing proportion of PUFAs of the algae.II. The P. aeruginosa 0-2-2 was cultured in the seawater PB medium and the pigments produced by the bacterium were isolated. The inhibitory effect of the pigments on the growth of some HAB species was first investigated with the P. donghaiense and Gymnodinium sp. as the main aimed objects, and the probable structures of the pigments were identified by means of UV and GC-MS.1. It was found that the growth of P. donghaiense was inhibited completely at lower concentration (4.0mg-L"1) of yellow pigment produced by P. aeruginos a 0-2-2. The blue pigment produced simultaneously by the bacterium showed lower algicidal capability, compared with the yellow pigment. It had remarkable inhibitory effect on the growth ofP. donghaiense when its concentration reached 10.0 mg-L"1, however, it exerted a lesser effect on the Gymnodinium sp. The yellow pigment also showed a drastic inhibitory effect on the growth of H. akashiwo while the blue pigment exerted almost no influence on the growth of the alga. Under the same experiment conditions, neither of the pigments exerted inhibitory effect on the growth of the green algae. Although the blue pigment showed a tendency of inhibitory effect on the growth of Dicrateria zhanjiangensis in its early stages, the alga showed signs of recovery. The results indicate that the pigments also possess a species-selective growth-inhibiting effect on some HAB species. 2. The pigments produced by P. aeruginosa 0-2-2 were indentified as phenazine compounds by means of UV and GC-MS. The main composition of the yellow pigment was 1 -Hydroxyphenazine and the main composition of the blue pigment was pyocyanine.III. The inhibitory effects of marine bacteria, screened from East China Sea, on the growth ofthe P. donghaiense, the high-biomass bloom-forming species in East China Sea were studied in present dissertation. The results would provide valuable evidence for preventing and controlling P. donghaiense blooms by biological means.1. Among the thirty stains of marine bacteria which were screened from the seawater collected from East China Sea when the P. donghaiense blooms occurred, thirteen of them showed strong inhibitory effects on the growth of P. donghaiense with the inhibition rate of 65 90%.The density of the bacteria was one of the important factors affecting their algicidal activity. Culture assays in the laboratory indicated that the growth of P. donghaiense was inhibited obviously by the bacterial strains at the concentration of 1.12 1.52 x lO'cell-mL"1, while at low concentration (1.12 -1.52 *10 6, 1.12 1.52x10 7 cell-mL"1) the bacteria had no effects on the growth of the algae.2. Under the same experiment conditions, the 13 bacterial stains exerted a lesser effect on the S. costatuma, another kind of common HAB species in East China Sea and essentially no effect on green algae and D. zhanjiangensis. These results strongly suggest that the algicidal effects of the bacteria are P. donghaiense-specific.IV. The inhibitory effects of synthetic surfactants including APGs and some new types of quarternary ammonium compounds (QACs) on the growth of some common HAB species in East China Sea had been first investigated systematically and an emphasis wasmade on the algicidal efficiency, the range of algicidal effect, algicidal mechanism and the mechanism of the selective algicidal action of the surfactants against HAB species. These results provide theoretical foundation and experimental basis for prevention and control of HAB species using new types of synthetic surfactants.1. The results indicate that the APGs can selectively inhibite the growth of P. donghaiense or even lyse the alga. Moreover, the APGs composed of long-chain alkyl are distinctly more efficient. The new types of QACs possess stronger algicidal activity than APGs and the cost of the QACs as algicide is much lower than that of the APGs. All of the QACs show remarkable inhibitory effects on the growth of P. donghaiense, Alexandrium tamarense and S. costatuma, some common HAB species in East China Sea at lower concentrations. The surfactants also show strong inhibitory effects on the growth of H. akashiwo. Under the same experiment conditions, the inhibitory effects of the surfactants on the growth of Platymonas helgolanidica and Platymonas subcordiforus are negligible or can not persist.2. Although the effects of different synthetic surfactants on the growth of the algae are different, the inhibitory effects of the different surfactants decrease at the same trend: Pyrrophyta , Xanthophyta < Bacillariophyta < Chlorophyta. Accordingly, the synthetic surfactants selectively inhibit the growth of some HAB species.3. The algicidal activity, including the inhibiting and especially the lytic effect, is also initiated by impairment of the biological membrane by the surfactant according to the results of microscope and TEM observations and a comparison of the correlation between the contents of PUFAs of the microalgae tested and the inhibitory effects of the synthetic surfactants on the algae implies that the species-specific responses to surfactants are also mainly attributable to the difference in the proportion of PUFAs of the algae.