| Fluorescence techniques have been effectively used for monitoring biomass, community composition, productivity and physiological status of phytoplankton, with the advantage of high precision and low noise. In this thesis, three kinds of fluorescence-based techniques including water-PAM, flow cytometry and fluorescence spectroscopy were applied to detect changes of arctic microalgae with two parameters including temperature and irradiance, for the purpose of establishing a new dynamic methodology.Four kinds of arctic microalgae including Heterocapsa arctica, Rhodomonas sp., Chlamydomonas sp.and Micromonas sp.were cultured under three different temperature (Heterocapsa arctica and Rhodomonas sp. were in -2°C, 2°C and 6°C; Chlamydomonas sp.and Micromonas sp. were in -2°C, 6°C and 10°C) with irradiance of 5000 lux, and three different irradiance (5000 lux, 15000 lux, 25000 lux) at 6°C. The main results are as follows:( 1 ) Photosynthetic physiological characteristics including Fv/Fm and RLC parameters were studied. The statistical analysis results showed the four arctic microalgae species (Heterocapsa arctica, Rhodomonas sp., Chlamydomonas sp.and Micromonas sp.) exhibited the best photosynthetic capacities at 6°C, 2°C, 6°C and -2°C, respectively. Besides, the four arctic microalgae presented similar photosynthetic physiological variations with detected irradiances. In the conditions of two higher irradiances, the largest photosynthetic electron-transport rate (rETRmax) did not increase significantly, and the ability to absorb irradiance was reduced. Under the irradiance of 25000 lux, arctic microalgae without Chlamydomonas sp. were stressed physiologically, while the tolerance to irradiance was increased.(2) Flow cytometry was used to detect cellular fluorescence of three kinds of strains, including Rhodomonas sp., Chlamydomonas sp. and Micromonas sp. whose diameters were smaller than 20μm. Data analysis results indicate that the cell size of the two arctic nanophytoplankton (Rhodomonas sp.and Chlamydomonas sp.) had significant response to temperature and growth periods (P﹤0.05). All kinds of microalgae existed the highest cellular content of Chl a at -2°C. Furthermore, the strain of Rhodomonas sp. also had the highest content of cellular phycobilin, whose pigments played positive effects on maintaining photosynthesis by improve the ability to absorb irradiance. Two strains of Chlamydomonas sp.and Micromonas sp. had significantly higher content of carotenoids at the two lower temperatures (P﹤0.05), which showed protective effect. These phtopigments exhibited different reactions among three different irradiances. Celluar content of carotenoids in Chlamydomonas sp. decreased significantly with increased irradiances (P﹤0.05). Besides, the content of Chl a decreased and the one of carotenoids increased in Micromonas sp. significantly.(3) Fluorescence spectral changes with different pigments were studied. The observed spectral variation showed that photopigments exhibited different reactions on environmental conditions. Ratios of fluorescence intensity at the three temperature conditions indicates: The ratio of carotenoids (490 nm) (the orange light in FCM) to Chl c (460 nm) had the highest value at -2°C, suggesting that these two kinds of photopigments had positive effect on maintaining higher photosynthesis ratio at -2°C, as well as Chl a (438nm) was the most effective pigment at 6°C. In addition, Chl b and lutein of Chlamydomonas sp. and Micromonas sp. which have a maximum absorption near 470 nm had the highest ratio of Chl b to lutein at 10°C. These photopigments also had significant response to the variation of irradiance. Pycobilin in Rhodomonas sp. play a positive role in maintaining community alive at 5000 lux. Chl c had the highest ratio at 15000 lux for Rhodomonas sp..(4) Changes of community abundance and bulk phytoplankton biomass were studied.Temperature and irradiance had significant effect on the arctic phytoplankton abundance and biomass. On the status of healthy photosynthetic physiology, arctic microalgae could not grow fast at lower temperature. At higher temperature, the lag phase was shorter and the community abundance was relatively high. Meanwhile, the high irradiance accelerated celluar divisions. The arctic microalgae including Heterocapsa arctica, Rhodomonas sp., Chlamydomonas sp. and Micromonas sp. had the highest bulk Chl a fluorescence intensity at -2°C, 2°C, 10°C and 2°C respectively. Within three irradiance conditions, Heterocapsa arctica and Rhodomonas sp. showed the highest bulk Chl a fluorescence intensity at 15,000 lux, while Chlamydomonas sp. and Micromonas sp. presented the highest bulk Chl a fluorescence intensity at 5000 lux. Consequently, the Chlamydomonas sp. relatively adapted to higher temperature and might be the dominant species in the future. The Heterocapsa arctica and Micromonas sp.were relatively prone to live in the ice core. Wheareas the Rhodomonas sp. relatively adapted to the morden arctic water.Generally, the optical parameters of FCM and the fluorescence spectroscopy provide mechanism to the dynamic fluorescence, while dynamic fluorecence is the synthetical response of the steady fluorescence. The fluorescence of FCM is more convenient but less precise,whereas the fluorescence spectroscopy were more precise, especially with the application of the wavelet. Consequently, photosynthetic physiological, single-celled optical properties and fluorescence spectra of four arctic microalgae at different temperature and irradiance conditions were studied. The synthetical application of the three kinds of fluorescence techniques was applied. We suggest an effective method for monitoring arctic phytoplankton in the rapidly changing environmental conditions with combination utilization of water-PAM, flow cytometry and fluorescence spectroscopy. |
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