Microalgae synthesize and store large volumes of lipids (potentially over 25% of dry weight) which could provide a renewable source of biodiesel. Traditional extraction techniques often produce poor lipid yields particularly from microalgae with robust cell walls. This project investigated the role of power ultrasound as a cell disruption step in lipid extraction from four microalgal species. Nile Red staining was used to assess the time when ultrasound induced increased membrane permeability in each species and lipids were extracted using an ultrasound assisted Bligh and Dyer extraction method. A 20 kHz probe system (40% amplitude, 0.086 W/cm3) caused increased lipid recovery from dry biomass in all cases; D. salina (no cell wall) from 15 to 22.5% of dry biomass after 1 minute (26% when stressed with 35 g/L NaCl). C. concordia (thin cell wall) from 7.5 to 10.5% of dry biomass after 2 minutes (27% with 25% nitrogen reduction in growth media). N. oculata (thick cell wall) from 6.5 to 10% of dry biomass after 16 minutes (31.5% when stressed with 30 g/L NaCl). The stressed cultures yield could be improved to 35% when ultrasound was combined with S070 beating beads. Chlorella sp. (thick cell wall) from 6.3 to 8.7% of dry biomass, after 16 minutes (44% was achieved when harvested at day 9 instead of 15). A Dual Frequency Reactor (16 and 20 kHz, 0.01 W/cm3) flow system with S070 beads demonstrated that high lipid extraction yields could be achieved on a larger level with N. oculata. After 4:48 minutes sonication 24% lipid recovery was achieved. This system could theoretically increase daily microalgal oil production from 3.96 to 5.76 L per day when compared to conventional techniques, at an extra production cost of only 2.9 p/litre (1.5% increase). D. salina, N. oculata and C. concordia resumed normal growth following sonication at 20 kHz after 1-20 days (8 minutes treatment for D. salina, 60 minutes treatment for N. oculata and 16 minutes treatment for C. concordia). It was found that the supernatant of sonicated D. salina and C. concordia when added to established cultures were able to boost their growth.

The effect of power ultrasound on algae blooms (Microcystis aeruginosa) over a 30 minute period was assessed using 200 and 400 mL suspensions of optical density of 2.0 at 680 nm. The frequencies employed were 20, 40, 580 (40%, 80%, and maximum intensity), 864 (40%, 80% and maximum intensity) and 1146 kHz (40%, 80% and maximum intensity). Ultrasound can induce two different effects on algal cells; inactivation at high power (≥ 0.0022 Wcm-3) and de-agglomeration at low power (≤ 0.0042 Wcm-3). Ultrasonic effects were observed using haemocytometer, optical density, UV-visible spectrometer, fluorospectrometer and flow cytometry. Using a 40 kHz bath (0.0214 Wcm-3) led to de-agglomeration resulting in an overall increase in algae of -0.28% by haemocytometer and -4.20% by optical density. The highest inactivation achieved was 91.54% (haemocytometer) and 44.63% (optical density) using 1146 kHz (maximum intensity, 0.0248 Wcm-3) and 200 mL suspension. In terms of efficiency to achieve inactivation (i.e. inactivation % / power) the best result was observed at 864 kHz (40% power setting, 0.0042 Wcm-3) with 200 mL suspension giving 8226.19 by haemocytometer and 5011.90 by optical density. This initial part of the study allowed a comparison to be made of the ultrasonic parameters that would lead to optimum algae removal in terms of acoustic energy input. The haemocytometer results for cells number were generally higher than those indicated by optical density which is probably due to the fact that the former records only cell numbers remaining whereas the latter is an overall measure of algae concentration (ruptured cells will still register, because their contents remain in suspension). Studies on de-agglomeration and inactivation were also undertaken using small or medium-scale ultrasonic equipment that were models for industrial scale systems. The following volumes of algae suspension and equipment were employed: Sonolator (Sonic Corporation, 5L flow), 16 kHz and 20 kHz Dual Frequency Reactor (DFR, Advanced Sonics LLC, 1L static and 3.5 L flow), 20 kHz Vibrating Tray (Advanced Sonics LLC, 1.5L static) and 20 kHz ultrasonic probe (made at Southeast University, 4L static). The most effective inactivation effects were obtained with the DFR reactor in static mode and 60% power setting for 10 minutes which achieved reductions calculated at 79.25% using haemocytometry and 60.44% by optical density. The third part of this study was to gain a greater understanding of the basic mechanisms of the action of ultrasound on algae and to interpret this in terms of its potential for algal cell removal and control. Algal cell activity was assessed by three methods: using a UV-visible spectrometer (Shimazu, 2450PC), a fluorometer (Shimazu, RF5301) and a flow cytometer (BD FACS Calibur). Ultrasonic damage to Chlorophyll A was revealed through observation of the loss in UV-Vis spectrophotometer peaks around 600 nm together with the decrease in fluorometer results for peaks around 500 and 680 nm. Flow cytometer results were able to identify the number of both intact cells and damaged/ruptured cells thus giving greater insight into the mechanism of ultrasonic inactivation. The direct rupture of cells by power ultrasound was prevalent at low frequencies ≤ 40 kHz due to the mechanical effects of cavitation collapse and inactivation of algal cells by free radicals occurred at high frequencies ≥ 100 kHz and medium powers where mechanical effects are much reduced. In conclusion, this work has shown that power ultrasound can provide a suitable method to control algal growth in small and medium laboratory scales. Scale-up beyond this point is the subject of further research but the results herein clearly demonstrate the importance of choosing the correct ultrasonic parameters in terms of frequency, power and exposure time.

Vampirovibrio chlorellavorus (Gromov et Mamkaeva, 1980) is a member of the phylum cyanobacteria that has been described as an obligate pathogen of several of the green microalga, Chlorella. It utilizes as yet unknown functions to access the contents of individual Chlorella sp. host cells, which results in cell death. Its presence in a cultivated Chlorella sorokiniana culture was first discovered using polymerase chain reaction (PCR) to amplify the 16S ribosomal DNA gene, followed by DNA sequencing. Its continued routine detection throughout much of the cultivation season suggested it was an endemic member of the phycosphere community in this open cultivation system, located in Tucson, Arizona. Ultimately, its presence resulted in rapid death of C. sorokiniana in open pond systems and reduced biomass harvest. PCR analysis of total DNA isolated from sand and soil layers removed from a nearby riverbed indicated that V. chlorellavorus resides naturally in the riverbed. The ability to manage this bacterial pathogen in cultivated Chlorella host species is hindered by the limited information available in the literature regarding the biological and genomic characteristics of V. chlorellavorus. The objective of this study was to identify environmental factors that trigger the apparent increased growth rate of V. chlorellavorus and rapid algal death during the cultivation cycle. In laboratory experiments, V. chlorellavorus was shown to cause death of C. sorokiniana when the temperature exceeded 28°C, whereas, algal death was not observed when the temperature was 24°C or lower, among the temperatures tested. Also, the bacterium was more pathogenic to C. sorokiniana, grown in open cultivation systems during the summer months, compared to the cooler season months. Futhermore, when C. sorokiniana and V. chlorellavorus were co-cultivated in the presence of sodium chloride ranging from 0-10 g per liter, the growth of the bacterium was not impeded to any extent that might suggest C. sorokiniana was rendered less susceptibility to pathogen attack. Future work involves examining more triggers and ways to inhibit V. chlorellavorus growth.

Eutrophication caused by nutrient loads from human activities is considered as one of the most serious environmental threats to the Baltic Sea. Due to climate change, cyanobacterial blooms are expected to increase in the future. This could affect people’s utility of beach recreation negatively in countries surrounding the Baltic. Based on a web survey carried out in south-eastern Sweden, public’s reactions and attitudes to cyanobacterial blooms are analysed. Possible economic impact on Gotland of more widespread blooms are estimated, and public demand for better information is evaluated. The result shows that 30% of the respondents from south-eastern Sweden would consider cancelling their plans of travelling to Gotland with knowledge about forthcoming algal blooms around the island. Determinants of tourists’ tendency to cancel their travel arrangements are earlier negative experiences of algal blooms and concerns regarding their pets’ bathing. The annual local economic loss for Gotland’s tourism industry is estimated to between 15 and 221 million SEK. The median willingness to pay for a mobile application which provides one-day forecasts of algal blooms is 25 SEK on Gotland and 20 SEK in southeastern Sweden. Boat owners, people who visit beaches often and those who travel to Gotland frequently, are more likely to pay for the mobile application. People who think algal blooms are natural show less demand for information.

The impact of grazers on the primary production of marine ecosystems has largely been explored in tropical environments. A number of studies support theories on the functional importance of grazers in the community structure of coral reefs. However, large-bodied grazers, like juvenile green turtles, co-occur with herbivorous fishes in subtropical and tropical regions throughout the world and we know little about their combined impact on macroalgal communities and whether they compete for macroalgal resources. My dissertation research was composed of four studies that were conducted simultaneously to further our understanding of plant/herbivore interactions in marine ecosystems. Studies were conducted at the Trident Basin, a non-public military facility within the Port Canaveral Inlet at Cape Canaveral, Florida, USA. The macroalgal study (Chapter 1), determined the spatial and temporal distribution of the macroalgal community. The foraging habits of juvenile green turtles were compared with the macroalgal abundance within the Basin and over time (Chapter 2). Selection ‘for’ specific macroalgal species (based on their availability in the macroalgae study) was used to determine the level of overlap and/or partitioning of resources among herbivorous fishes and juvenile green turtles (Chapter 3). The final empirical study (Chapter 4) measured the impact on thallus height, diameter and/or branching of macroalgae as well as the macroalgal community composition from caging experiments that excluded herbivorous fishes and juvenile green turtles. The algal community was predominantly composed of nine red and green macroalgal species that were persistent year-round. Grazer-resistant macroalgae were rarely observed. Green turtles foraged on many of these same macroalgae but also opportunistically foraged on flotsam, including anthropogenic debris (e.g., plastic). The gut content of the major herbivorous fishes in the community (Abudefduf saxatilis, Archosargus probatocephalus, Diplodus holbrooki, and Lagodon rhomboides) foraged as omnivores depending on where they were captured within the Basin area or their size. All herbivores showed selection for less abundant green algae (i.e., Ulva spp.). Results of the exclusion of juvenile green turtles and large herbivorous fishes in caging experiments suggest that grazing by these large-bodied herbivores had no impact on the composition of the macroalgal community and little impact on the morphological structure of the macroalgal species that were examined. Collectively these four studies contribute to a better understanding of how multiple grazers have evolved to forage in macroalgal communities without detrimental effects on their food resources. / Includes bibliography. / Dissertation (Ph.D.)--Florida Atlantic University, 2014. / FAU Electronic Theses and Dissertations Collection

Crustose coralline algae (CCA) are important reef stabilizers and their susceptibility to anthropogenic climate change and ocean acidification (OA) is of concern. Ocean acidification effects on benthic algal communities were determined by the response of CCA, fleshy macroalgae and microalgae to the interaction of pCO2 and light. I examined if elevated pCO2 and light influences CCA dominance by assessing their growth, recruitment and calcification. Elevated pCO2 under natural reef diurnal CO2 cycles did not significantly affect CCA percent cover, calcification rates or survival of adult CCA lobes. No significant community pCO2 effects were observed, rather light controlled dominance. The percent cover of microalgae increased in highlight, while CCA increased in the shade. My results indicate that algal response to irradiance is a more significant driver of reef benthic algal change than pCO2 levels predicted for 2100; however, this conclusion should be corroborated in longer-term and in field experiments. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2015. / FAU Electronic Theses and Dissertations Collection

Field data from CO2 vents, a current model of future ocean acidification conditions, show a positive correlation between elevated seawater pCO2 and fleshy macroalgal abundance, as well as a negative correlation between elevated seawater pCO2 and calcareous macroalgal abundance on coral reefs. One underlying physiological mechanism for increases of fleshy macroalgae species in response to greater pCO2 could be an increase in their photosynthesis. Furthermore, inorganic carbon use mechanisms, irradiance and depth may influence species-specific responses to ocean acidification. Therefore, this thesis aimed to discern carbon use strategies and photosynthetic responses to elevated pCO2 of dominant tropical fleshy and calcareous macroalgae. All species studied were able to utilize HCO3 - for photosynthesis. 33% of calcifying macroalgae and 80% of fleshy macroalgae had increased photosynthetic rates in response to lower pH. Thus, future conditions of OA may perpetuate or exacerbate the abundance of fleshy seaweeds at the expense of calcareous species. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2017. / FAU Electronic Theses and Dissertations Collection

The small dinoflagellate Azadinium spinosum produces azaspiracid (AZA) toxins which can contaminate filter feeding shellfish to dangerous levels. Toxin-contaminated shellfish flesh, when consumed by humans, can cause acute intense illness and chronic health issues. Shellfish biotoxins are monitored in Scottish shellfish by Food Standards Scotland (FSS), and the concurrent monitoring of harmful phytoplankton in the water column acts as an important early warning system of future shellfish toxin contaminations. Since A. spinosum is very small (12-16 μm long) it is difficult to identify using a light microscope, therefore molecular techniques have been developed to detect species-specific environmental DNA from phytoplankton samples. In this thesis the application and verification of quantitative real time polymerase chain reaction (qPCR) is discussed in detail and documents its first use in Scottish waters to survey A. spinosum abundance and seasonality. The limit of detection of the method was found to be 2000 ±5600 cells L-1, however it is unclear whether this is adequate for regulatory monitoring because it is not yet understood how cell density in the water column relates to AZA shellfish toxicity. The qPCR probe and primer sequences were also found to be too specific to detect all strains of the A. spinosum species, as new strains have been isolated since their development. This is a significant hindrance to the application of the tool for monitoring which will need to be addressed in the future through the isolation of local A. spinosum strains. Over a year long sampling period, A. spinosum was detected only twice (maximum cell density of 2545 ±5600 cells L-1, August 2014) off the Shetland Islands. The seasonality of the species in Scottish waters could not be assessed with so little data, however other observed harmful species of importance to shellfish regulatory monitoring are discussed; of particular note an unusual bloom of Dinophysis acuta as its association with a temperature front at the mouth of Loch Fyne. This thesis critiques the use of this qPCR technique for A. spinosum detection at high-throughput. The issues which have been highlighted do not prevent its future use by FSS, but highlight specific areas of development which need addressed before national monitoring can occur.

This study was carried out on algal biofilms grown using rotating algal biofilm reactors (RABRs) with the aim of: i) characterizing their growth in terms of photosynthetic activity and morphology ii) evaluating their performance as a wastewater treatment option and a feedstock for biofuels production, and iii) examining the algal-bacteria interactions. A review of algal biofilm technologies currently employed in wastewater treatment processes was made to compare nutrient removal efficiencies, factors that influenced algal biofilm growth, and the different bioproducts generated from algal biomass. Consequently, research efforts were directed towards addressing pertinent issues identified in literature in order to optimize these systems for wastewater treatment and bioproducts production. Successful growth of algal biofilms in municipal wastewater and subsequent removal of nutrients from the wastewater was demonstrated. Photosynthetic and respiration rates observed with depth of the biofilm were influenced by the biofilm composition (single vs. mixed species), culturing conditions (laboratory vs. outdoor), orientation to the light, nitrogen availability (N-replete vs. N-deplete), and dissolved inorganic carbon availability (presence or absence of bicarbonate). Slight enhancement in lipid production was also observed as a result of nitrogen stress and bicarbonate addition. However, the accumulated lipids were not as much as expected or as reported in suspended cultures. Presence of bacteria positively influenced microalgae growth in the mixed cultures but the reverse was not true. In conclusion, photosynthetic activity and biofilm structure were characterized with methods developed for the algal biofilms in this study. For now, productivity of the algal biofilms needs to be maximized in order to fully utilize its potential as a biofuel feedstock and nutrient removal option. Further research on algae-bacteria interactions using species native to the wastewater grown algal biofilms is recommended.

The rates of algal production and Sphaerotilus assimilation in the lower Logan River benthos were investigated in 1966 and 1967 . The rate of annual gross primary production (3,416 Kcal/m2/yr) was estimated from the relation of pigments to the photosynthetic rate of benthic communities in a submersible, metabolism chamber. The photosybthetic rate was predicted with high precision when a measure of the accessory pigments (D480/D665 and/or chlorophyll-c ) was considered with the chlorophyll-a . The pigment density estimates were obtained from the community present on paraffin-coated, concrete hemispheres after immersion in the river for periods ranging from 11 to 20 weeks. The daily rate of energy utilization by Sphaerotilus (1.3 c a l/m2/day) was estimated from the observed generation time of this bacterium on glass slides suspended in the river at various locations and metabolic coefficients obtained from other sources. The magnitude of microbial activity in the river water Has estimated to be 448 cal/m2/day. The accumulation rate of Sphaerotilus biomas s on glass slides was 0.3 mg? (net weight)/m-/48 hours , and could be predicted from the temperature, nitrate (plus nitrate) content , and the dissolved organic carbon content of the river water. The generation time of Sphaerotilus (average was 20 hours) could be predicted f rom temperature, nitrate (plus nitrite) content, and velocity of t he water. The daily P/B coefficient for this bacterium was 1.20.