Variation in allelopathy of extracellular compounds produced by Cylindrotheca closterium against the harmful-algal-bloom dinoflagellate Prorocentrum donghaiense.

Allelopathy between algae is an ecological strategy that can facilitate or inhibit the occurrence of algal blooms. The role of allelopathic effects of marine microalgae Cylindrotheca closterium in other phytoplankton population dynamics are still limited. In the current study, the effects of cell-free filtrates of diatom Cylindrotheca closterium on two common dinoflagellates (Prorocentrum donghaiense and Prorocentrum cordatum), a chrysophyceae (Isochrysis galbana) and a diatom (Chaetoceros curvisetus) were investigated within controlled laboratory experiments. It was observed that the growth of P. donghaiense was significantly suppressed and approximately 80% cells disappeared after 8-d exposure, while the other three algae was less sensitive. P. donghaiense was very sensitive to the exudates of C. closterium from the stationary phase by comparing various percentage (10, 30, 50, 70 and 100%) of filtrates. In addition, the allelopathic effects of extracellular compounds of C. closterium extracted by three different organic solvents (ethyl acetate, chloroform and petroleum ether) on P. donghaiense were explored by determining cell density, chlorophyll content and maximum photosystem II (PSII) quantum yield (Fv/Fm). It was found that the compounds extracted by ethyl acetate and chloroform appeared to exhibit less toxicity on P. donghaiense than that of petroleum ether. The present results indicated that the allelochemicals released by C. closterium might be concentrated effectively in the petroleum ether extraction phase, which provided a new perspective for controlling the red tides of P. donghaiense in the East China Sea by means of the ecological inhibitors extracted.

The stresses of allelochemicals isolated from culture solution of diatom Phaeodactylum tricornutum Bohlin on growth and physiology of two marine algae.

The allelopathic effects of extracts isolated from the culture filtrate of diatom Phaeodactylum triconutum Bohlin on typical marine microalgae Prorocentrum donghaiense Lu and Dunaliella salina Teodoresco were investigated by determining different physiological and biochemical parameters, such as growth rate, membrane systems and esterase activity under controlled laboratory conditions. The growth of P. donghaiense was significantly inhibited immediately after exposure to the allelochemicals, while the algae density of D. salina was less sensitive. Chlorophyll-a content, membrane systems, as well as esterase activity were simultaneously investigated by flow cytometry with particular fluorescent markers and exhibited changeable sensitivities. The results demonstrated that the membrane systems of P. donghaiense were suppressed by the allelochemicals directly, causing loss of integrity and membrane penetration. Esterase activity was the most sensitive indicator as that of P. donghaiense cells significantly increased in short time and was inhibited subsequently. However, the membrane of D. salina remained intact still after exposure to the extracts and the esterase activity was only inhibited on last day during experiment period. Membrane potential and chlorophyll-a content of the two marine algae also showed somewhat different changes, as that of P. donghaiense cells were impaired after 5 day exposure to all volume conditions while these two characteristics of D. salina was only suppressed by exposure to high volume of the allelochemicals on day 6. The present results indicated that the inhibition of culture filtrate of P. triconutum on P. donghaiense was algicidal whereas the effect on D. salina appeared to be algistatic.

Effects of an allelochemical in Phaeodactylum tricornutum filtrate on Heterosigma akashiwo: Morphological, physiological and growth effects.

The effects of an allelochemical extracted from the culture filtrate of diatom Phaeodactylum tricornutum on the raphidophyte Heterosigma akashiwo were investigated using a series of morphological, physiological and biochemical characters. Growth experiments showed that H. akashiwo was significantly inhibited immediately after exposure to the allelochemical, with many cells rapidly dying and lysing based on microscopic observation. The effects of the allelochemical on the surviving cells were explored using Scanning Electron Microscopy (SEM) and Flow cytometry (FCM), the latter by examination of a suite of physiological parameters (membrane integrity, esterase activity, chlorophyll-a content, membrane potential). The results demonstrate that the membrane of H. akashiwo was attacked by the allelochemical directly, causing cell membrane breakage and loss of integrity. Esterase activity was the most sensitive indicator of the impacts of the allelochemical. Membrane potential and chlorophyll-a content both showed significant decreases following exposure of the Heterosigma cells to high concentrations of the allelochemical for 5 and 6 days. Both were affected, but the membrane potential response was more gradual compared to other effects. The cell size of H. akashiwo did not change compared with the control group. The surviving cells were able to continue to grow and in a few days, re-establish a successful culture, even in the presence of residual allelochemical, suggesting either development of cellular resistance, or the degradation of the chemical.

Preliminary analysis of allelochemicals produced by the diatom Phaeodactylum tricornutum.

Marine diatom Phaeodactylum tricornutum is known to exude allelochemicals with negative effects on Heterosigma akashiwo according to our previous study, while the information about the allelochemical compounds remains unknown. The present study dealt with isolation and analysis of the active substances released by P. tricornutum into the culture medium. Filtrate of P. tricornutum was extracted using ethyl acetate and chloroform respectively. The anti-algal fractions were isolated using high performance liquid chromatography (HPLC) and screened using activity-guided fraction methods. Results demonstrated that fraction Ⅱ and Ⅵ showed significant allelopathic effect on H. akashiwo growth. Then the anti-algal activity fractions were analyzed preliminary using gas chromatography-mass spectrometry (GC-MS) and high performance liquid chromatography-electrospray time-of-flight mass spectrometry (HPLC-ESI-TOF-MS). An active compound was derived from fraction Ⅵ with the molecular weight of 578 and possible molecular formula of C30H38N6O6, which was speculated to be TYR-PRO-PHE-PRO-GLY-NH2. a kind of glycinamides.