Microalgae Samples Research Articles

Drying of Algae: Analysis of Oedogonium Dehydration Kinetics

Algae cultivation has multiple potential applications including biofuel production, wastewater treatment, and carbon capture. However, dewatering and dehydration remain a challenge for commercialization. Research has focused mainly on microalgae, but filamentous algae have recently gained attention due to their larger size and thus relative ease of harvesting. The drying mechanics of filamentous algae has yet to be fully understood and optimized. This study investigated the drying behavior of the filamentous algae Oedogonium at 50℃, 60℃ and 70℃. The moisture content of triplicate samples of microalgae and Oedogonium (intact filaments, fractured filaments, and aggregated forms) was measured during drying. Intact Oedogonium filaments were fractured using a food blender. Intact Oedogonium had an average shorter falling rate phase and higher diffusivity then microalgae at all temperatures, resulting in a 15%, 8.3%, and 11% faster total average drying time at 50℃, 60℃ and 70℃ respectively. Fractured Oedogonium further had a shorter falling rate phase than intact Oedogonium at 50℃ and 60℃ where its average total drying time was respectively 18% and 9% faster, but the effect of the fracturing treatment reduced with increasing temperature and became negligible at 70℃. Thus, Oedogonium appears to dry more effectively than microalgae, and fracturing Oedogonium results in faster dehydration at lower temperatures. These findings could help optimize industrial drying and assist in the commercialization of algae cultivation.

The effects of microalgae (Spirulina platensis and Chlorella vulgaris) extracts on the quality of vacuum packaged sardine during chilled storage

The influence of water extracts of Spirulina platensis and Chlorella vulgaris (1%) on sensory, chemical and microbiological changes was investigated in vacuum packaged sardine fillets at 4 ± 1 °C for 15 days. Mineral content and bioactive chemical components of microalgae samples were also detected by ICP-MS and GC–MS, respectively. The results showed that S. platensis and C. vulgaris were rich in mineral. Main components in the extracts of C. vulgaris and S. platensis were 3,5-dichloro-6- nitrocholestane (31.74%) and dioctylamine (67.64%), respectively. The extracts prolonged sensorial shelf life of sardine for 3 days although S. platensis group was more preffered than C. vulgaris. These microalgae extracts delayed lipid oxidation and inhibited bacterial growth apart from Gram-positive lactic acid bacteria in sardine meat. The study result revelaed that both extracts can be used as antioxidant and antimicrobial additives to maintain sensory characteristic of sardine fish.

Использование автоматического счетчика частиц Luna-II для оценки численности клеток морских микроводорослей и их размеров в культурах

Marine microalgae isolated in algologically pure cultures are widely used for basic and applied research. All studies related to microalgae are accompanied by an assessment of their abundance and cell size. The main method for determining these parameters is light microscopy. However, this method is quite time-consuming, therefore, at present, various kinds of automatic particle counters are increasingly being used. This significantly reduces the processing time of microalgae samples and increases the accuracy of measurements. The purpose of this work is to evaluate the possibility of using an automatic particle counter Luna-II to find out the abundance of cells of certain types of marine microalgae and determine their linear sizes. The performed work allowed us to show the possibility of using this counter to quickly record the abundance of culture cells of marine microalgae and determine their linear sizes with high accuracy. Reliable results can be obtained for dense cell suspensions represented by single cells with a low percentage of doublets and triplets (not more than 3%) and a high sphericity index (above 0,95). For algal cultures, the cells of which have an average and low sphericity index, reliable results can only be obtained by the abundance of cells, provided that there is a slight presence of sticking cells (doublets and triplets) in the suspension. The presented automatic counter can be used when conducting ecological and physiological studies on the cultures of certain types of marine planktonic algae, as well as in monitoring activities related to assessing the quality of the aquatic environment using microalgae as bioassays.

Biomass and Bio-butanol Production from Borodinellopsis texensis CCALA 892 in Synthetic Wastewater: Determination of Biochemical Composition

Microalgae can generally maintain the high amounts of biomass in the wastewater and they can be converted from biomass to bio-butanol. Bio-butanol is a liquid biofuel and it has significant physical and chemical properties. In this study, we carried out bio-butanol production from Borodinellopsis texensis CCALA 892 grown in various concentrations of the municipal wastewater. Also, we determined biochemical composition ratios of microalgae samples and studied the some antioxidant enzymes such as catalase, superoxide dismutase and ascorbate peroxidase. In the current study, bio-butanol was produced by the acetone-butanol-ethanol (ABE) fermentation method. The microalgae sample grown in 25% of wastewater had the highest biomass productivity among five wastewater samples with 0.114 ± 0.002 g L-1d-1. The carbohydrate and protein concentrations of control group increased day by day and their values reached stationary phases at seven days. The sample grown in 25% of wastewater had the highest carbohydrate concentration with 0.30 g L-1 and protein concentration with 0.35 g L-1 at the stationary phase. The maximum enzyme activities for catalase, superoxide dismutase and ascorbate peroxidase were 15.33 ± 0.88, 8.67 ± 0.67 and 33 ± 1.53 µmole/mg, respectively at 25% of wastewater. In addition, bio-butanol content of B. texensis CCALA 892 was 3.63 ± 0.21 g L-1 and its bio-butanol yield was found as 0.18 ± 0.011 g/g sugar. In the next study, we can examine large scale butanol production.

Single Chlamydomonas reinhardtii cell separation from bacterial cells and auto‐fluorescence tracking with a nanosieve device

AbstractA planar, transparent, and adaptable nanosieve device is developed for efficient microalgae/bacteria separation. In the proposed method, a sacrificial layer is applied with dual photolithography patterning to achieve a 1D channel with a very low aspect ratio (1:10 000). A microalgae/bacteria mixture is then introduced into the deformable PDMS nanochannel. The hydrodynamic deformation of the nanochannel is regulated to allow the bacteria cells to pass through while leaving the microalgae cells trapped in the device. At a flow rate of 4 μL/min, the supernatant collected from the device is indistinguishable from a control solution, indicating that nearly all the microalgae cells are trapped in the device. Additionally, this device is capable of single cell auto‐fluorescence tracking. These microalgae cells demonstrate minimal photobleaching over 250 s laser exposure and could be used to monitor hazardous compounds in the sample with a continuous flow. This method will be valuable to purify microalgae samples containing contaminations and study single‐cell heterogeneity.

Microalgae a Superior Source of Folates: Quantification of Folates in Halophile Microalgae by Stable Isotope Dilution Assay.

A multitude of human nutritional supplements based on Chlorella vulgaris biomass has recently been introduced to the specialty food market. In this study, an analysis of total folate contents in Chlorella sp. and a series of marine microalgae was conducted to evaluate folate content in alternative algae-based food production strains. For the first time, total folate content and vitamer distribution in microalgae were analyzed by stable isotope dilution assay (SIDA) using LC-MS/MS, which has demonstrated its superiority with respect to folate quantification. Consistently, high folate contents were detected in all examined microalgae samples. High folate concentrations of 3,460 ± 134 μg/100 g dry biomass were detected in freshly cultivated Chlorella vulgaris, notably also in other well-researched microalgae strains. To that end, the highest folate content currently documented for any algae sample was measured in the marine microalgae Picochlorum sp. isolate with values of 6,470 ± 167 μg/100 g dry biomass. This calls for alternative products based on other algae biomass. Our data indicate that freshwater and marine microalgae provide extremely high concentrations of folates, which warrant further studies on the regulation of pteroylpolyglutamates in algae as well as on bioaccessibility, absorption, and retention in humans.

Chlorella vulgaris sp. microalgae as a feedstock for biofuel

As need of energy is increasing, day by day reserves of fossil fuels are exhausting and use of non-renewable fuels causes setting pollution, there's have to boost the utilization of alternation inexhaustible source of energy. Among renewable resources biodiesel is one of significant biofuel. Microalgae are well known as a promising tool for biofuel production like a renewable energy source that can be converted to biofuel. In the current study, possibility of green algae like a possible feedstock for biodiesel output continues to be evaluated. Entire 113 isolates from four samples of microalgae collected from various localities of Ramanathapuram, Tamilnadu, were screened for biodiesel production. Biomass fatty acid, yield, lipid comfortable composition as well as optimum problems for lipid production for microalgae are generic/specie unique. Chlorella vulgaris is examined as ideal applicants for biodiesel production on the foundation of biomass yield, lipid written content in addition to quality of lipids. There's should lengthen the work for mass cultivation of these microalgae on cost substrates that are low therefore price of biomass output may be lowered.

The Community Structure of Microalgae and Exploring its Potentially to Biofuel Producing in Ranu Grati, East Java Indonesia

Microalgae are microscopic autotrophic organisms which act as one of the biofuel-producing organisms. Ranu Grati is one of the volcanic lakes located in Pasuruan, East Java. Human activities around the waters are responsible for eutrophication of the water quality. The aim of this study was (1) described the profile of water quality and microalgae community structure, and (2) investigated microalgae potential as a producer of biofuels based on lipid content. Explorative research was carried out by taking a sample of water at a depth of 50 cm at 5 stations. Water quality parameters measured were DO, pH, BOD, TP, and TKN. Microalgae samples were taken by filtering 3 L of water using Plankton Nets. Plankton samples were identified and calculated using Haemocytometer. Furthermore, microalgae abundance, Important Value Index (IVI), diversity index, richness taxa, Evenness, and Richness index were determined. Species with the two highest IVI was determined the lipid levels use literature. The results showed that the waters of Ranu Grati had been contaminated with high organic matter (BOD levels 8.30-8.72 mg/L) with trophic status classified as mesotrophic to hypertrophic (TP 0.05-0.08 mg/L and TKN 0.23-0.32 mg/L). DO levels was high and have a range value 7.47-9.87 mg / L, while range of pH value 8.73-9.07. There were found 18 species of microalgae with varied 10-18 species in each station. Pollution status of Ranu Grati based on Shannon Wiener diversity index categorized as moderately polluted with range values 1.95-2.34. Value of microalgae evenness 0.77-0.86 indicated high uniformity with equally even distribution of species. Chlorella sp. and Scenedesmus sp. have the highest IVI and have a potential to produce biofuel with lipid levels 28 -32% (dry weight) and 12-14% (dry weight) respectively.

Coolia guanchica sp. nov. (Dinophyceae) a new epibenthic dinoflagellate from the Canary Islands (NE Atlantic Ocean)

ABSTRACT A new warm-water benthic dinoflagellate species, Coolia guanchica sp. nov., is described from the Canary Islands (NE Atlantic Ocean), a subtropical archipelago with a rich benthic biodiversity. Samples of benthic microalgae were collected at Tenerife, and three strains of a previously unreported ribotype of the genus Coolia were found. Identification was carried out by combining morphological (light microscope, LM; and scanning electron microscope, SEM) and molecular (large subunit ribosomal gene, LSU rRNA; and internal transcribed spacer region, ITS1-5.8S-ITS2 rRNA) data from clonal cultures. Phylogenetic analyses revealed that the new strains formed a well-supported clade, sister to Coolia canariensis. The genetic distance of more than 0.20 for LSU and 0.30 for the ITS region (uncorrected p values) from the closest members of the genus (C. canariensis and C. cf. canariensis) showed species-level divergence to justify the description of a new species, Coolia guanchica sp. nov. A bioassay with Artemia salina was used to check the toxicity of the species, with negative results. Coolia guanchica has rounded cells, with plate 4′ as the largest plate of the epitheca and plate 6′′ about twice as wide as long, similar to C. canariensis and C. areolata. It differs morphologically from the other two ribotypes in the C. canariensis complex in having a lighter hypothecal ornamentation, larger thecal plate pores scattered randomly throughout the cell and higher pore density per thecal plate.

Comparison of Nile Red and Cell Size Analysis for High‐Throughput Lipid Estimation Within Oleaginous Yeast

AbstractWith growing interest in oleaginous yeast as producers of future fuels and bulk chemicals, a robust, high‐throughput method for estimating lipid production is required. Although the lipophilic dye Nile red is frequently used to assay large samples of yeast and microalgae, inconsistent stain permeability between species and strains limits its effectiveness for some microorganisms. In this study, the oleaginous yeast Metschnikowia pulcherrima is used to develop a fluorescence‐free, cell‐size‐based image analysis method for estimating lipid production, which is then compared with an optimized Nile red method across several experimental scenarios. Cell size analysis (CSA) outperforms Nile red in all scenarios, correlating well with lipid extraction data when screening multiple strains, screening a subset of strains grown in different conditions, and tracking the lipid accumulation of a culture over time. Stain permeability is shown to vary significantly among the strains trialled, with lipid droplet size and cell wall thickness having a deleterious effect in the permeability of high‐lipid‐accumulating cells. CSA can also allow culture population dynamics to be monitored, providing key process information of cell size distribution in response to changing media compositions.Practical Applications: Nile red is currently the go‐to method for high‐ throughput lipid screening; however, staining inconsistencies in some organisms caused by varying cell morphology makes it challenging to optimize a robust protocol. Although fluorescence‐free methods exist (Raman spectroscopy, Fourier‐transform infrared spectroscopy, GCMS), the need for extensive sample preparation and specialist equipment restricts their widespread adoption. The CSA method presented here offers an accurate, robust, and cheap alternative for the study of microorganisms where fluorescence‐based avenues are not feasible. Furthermore, the population dynamics collected during CSA can easily be applied to bioreactor style processing, where tracking size distributions can provide real time information of culture status. This additional information is valuable even if fluorescence screening is a possibility.

An “omic” approach to Pyrocystis lunula: New insights related with this bioluminescent dinoflagellate

Pyrocystis lunula (Schutt) is a photoautotrophic dinoflagellate without armored form, frequently found in marine environments. Today, there are several biotechnological applications derived from the bioluminescent system of this species. From a post-genomic perspective, in order to have a starting point for studying the proteome of P. lunula, an “omics” approach (transcriptomics-proteomics) was assessed using fresh microalgae samples. A total of 80,874,825 raw reads were generated (11,292,087,505 bp; 55.82% GC) by mRNA sequencing. Very high-quality sequences were assembled into 414,295 contigs (219,203,407 bp; 55.38% GC) using Trinity software, generating a comprehensive reference transcriptome for this species. Then, a P. lunula proteome was inferred and further employed for its analysis on this species. A total of 17,461 peptides were identified, yielding 3182 protein identification hits, including 175 novel proteins. The identified proteins were further categorized according to functional description and gene ontology classification. SignificanceThe major contribution of the present work is making available a reference transcriptome and proteome of P. lunula, that is now accessible for the research community, and a functional description of the 3182 proteins inferred from the transcriptome, including 175 novel proteins, which have already been deposited in the ProteomeXchange and NCBI SRA databases, respectively. In addition to this, a series of important factors related to the bioluminescent system and the regulation of gene expression, were identified and described.

Effects of acids pre-treatment on the microbial fermentation process for bioethanol production from microalgae

BackgroundMicroalgae are one of the promising feedstock that consists of high carbohydrate content which can be converted into bioethanol. Pre-treatment is one of the critical steps required to release fermentable sugars to be used in the microbial fermentation process. In this study, the reducing sugar concentration of Chlorella species was investigated by pre-treating the biomass with dilute sulfuric acid and acetic acid at different concentrations 1%, 3%, 5%, 7%, and 9% (v/v).Results3,5-Dinitrosalicylic acid (DNS) method, FTIR, and GC-FID were employed to evaluate the reducing sugar concentration, functional groups of alcohol bonds and concentration of bioethanol, respectively. The two-way ANOVA results (p < 0.05) indicated that there was a significant difference in the concentration and type of acids towards bioethanol production. The highest bioethanol yield obtained was 0.28 g ethanol/g microalgae which was found in microalgae sample pre-treated with 5% (v/v) sulfuric acid while 0.23 g ethanol/g microalgal biomass was presented in microalgae sample pre-treated with 5% (v/v) acetic acid.ConclusionThe application of acid pre-treatment on microalgae for bioethanol production will contribute to higher effectiveness and lower energy consumption compared to other pre-treatment methods. The findings from this study are essential for the commercial production of bioethanol from microalgae.

Enrichment procedure based on graphitized carbon black and liquid chromatography-high resolution mass spectrometry for elucidating sulfolipids composition of microalgae

Microalgae have recently become a popular functional food due to their health benefits. Sulfolipids, a class of substances abundant in this matrix, have been reported to have interesting bioactivities, such as anti-carcinogenic activity. However, despite the potential interest in sulfolipids, a dedicated analytical method for their characterization is currently lacking but would significantly increase the coverage of sulfolipids with respect to the direct lipidomic analysis. To achieve this goal, in this work a procedure, based on graphitized carbon black solid phase extraction, was developed for clean-up and enrichment of sulfolipids (sulfoquinovosyldiacylglycerols and sulfoquinovosylmonoacylglycerols) and it was applied to spirulina (Arthrospira platensis) microalgae. A careful study of the solid phase extraction conditions was performed, first to maximize the recovery of reference standards, then to increase the total number of identified sulfolipids from the spirulina lipid extract. All samples were analysed by ultra-high performance liquid chromatography coupled to high resolution mass spectrometry and lipids were tentatively identified by Lipostar, for a reliable lipid structure assignment. The developed method was compared to the direct lipidomic analysis without enrichment, to establish the enrichment efficiency in terms of number of identifications. From the comparison, the enrichment procedure proved better and allowed the tentative identification of 199 sulfolipids, which is the largest number reported so far for the Arthrospira platensis species. The described method was validated in terms of precision, accuracy, recovery, limit of quantitation and detection for two sulfolipids. Finally, a relative lipid quantitation based on peak area was carried out on the microalgae sample, which indicated nine abundant sulfolipids as representing ca. 60% of sulfolipids in spirulina microalgae.

Fatty Acid Compositions of Mixed Microalgae from Tilapia Fish Ponds

Microalgae has been getting broad attention of researchers and investors lately, especially when discussing on healthy food and energy sources for the future. In this study, twelve samples of mixed microalgae from outdoor ponds were analyzed for their fatty acid compositions. The potential of microalgae to solve variety of world’s problems was not realized because of bottleneck in microalgal supplies at reasonable cost. Therefore, the objective of this study is to determine fatty acid profiles of mixed microalgae from tilapia fish ponds. The study was conducted in Tapak Ternakan Ikan, Taman Pertanian Universiti and Department of Biology, Faculty of Science, Universiti Putra Malaysia. Mixed microalgae were extracted for their lipids with methanol: chloroform mixture and after transesterification, the fatty acid methyl ester were analyzed using gas chromatography equipped with flame ionization detector. Results showed that saturated was the major constituent fatty acids. The average percentages of saturated fatty acids, monounsaturated fatty acids, and polyunsaturated fatty acids obtained were 45.62 ± 1.37%, 20.05 ± 1.14%, and 34.33 ± 3.17% respectively. The most dominant fatty acid profiles were C18:3n3 (α-linolenic acid) and C16:0 (palmitic acid), with the overall percentages of 19.97% and 19.40% respectively. The fatty acid profiles of mixed microalgae was good with a decent balance of saturated, monounsaturated and polyunsaturated fatty acids.

Insight on a comprehensive profile of volatile compounds of Chlorella vulgaris extracted by two "green" methods.

Some green extraction methods were selected and tested for the extraction of volatile compounds from different samples of the microalga Chlorella vulgaris: ultrasound‐assisted liquid–liquid extraction using environment‐friendly solvents (LLE) and solid‐phase microextraction (SPME). The obtained profiles of volatile chemical compounds were different. Only one molecule was found in common to both extractions. Using the SPME method, the main chemical classes of identified volatile compounds were sulfuric compounds, aldehydes, and alcohols. Using the LLE method, the volatile profile was more balanced with alkanes, fatty acids, terpenes, alcohols, and aldehydes. Multivariate data analyses permitted discrimination among samples. Additionally, the relationship between the physicochemical properties of identified volatile compounds and the methods of extraction was studied. The results showed that the LLE extraction allowed the extraction of volatile compounds having a high boiling point (>160°C) and a high log P (>3). The SPME method was more effective to extract volatile compounds with a low boiling point (<160°C) and a low log P (<3). It is thus necessary to combine several extraction methods to obtain a complete view of the volatile profile for microalgae samples.

Morphological and genetic variability of Skeletonema dohrnii and Skeletonema japonicum (Bacillariophyta) from the northwestern Sea of Japan

ABSTRACTSpecies of the genus Skeletonema are one of the most common marine planktonic diatoms of the temperate zone of the World Ocean. Despite the worldwide distribution of the genus, there are only fragmentary data on its ecology and quantitative characteristics of habitats, as well as limited information on the species diversity in the northeastern Pacific Ocean. In the present study, 13 strains of Skeletonema were obtained from planktonic and benthic samples of microalgae collected during the period from 2009 to 2017 from the northwestern Sea of Japan. We identified Skeletonema dohrnii and S. japonicum based on light and electron microscopic observations and molecular genetic analysis of 28S rDNA sequences. A detailed description, as well as new data on water temperature, salinity and species distribution, is provided for each species and their morphological features are discussed. The presence of several rimoportula processes on the terminal valve of Skeletonema species is shown for the first time. Our results support the usage of the topological criterion for the delimitation of the S. dohrnii–marinoi, as opposed to the distance-based criterion.

Effect of NaNO3 and NaCl concentration on Nannochloropsis oculata cell biomass and FAME composition for biodiesel production

In the present study, the growth of microalgal strain Nannochloropsis oculata was studied in different concentration of NaNO3 and NaCl as nutrient factors in culture medium to investigate its effect on the cell biomass and fatty acid methyl esters (FAME) composition. The microalgae were exposed to nitrogen and salt stress in two different experiment set-ups. The nitrogen source, NaNO3 was supplied at different concentration which are 25, 50, 75, and 100 mg/L while NaCl concentration was fixed at 25 g/L whereas NaCl concentration were tested at 0, 10, 25 and 35 g/L while NaNO3 was fixed at 75 mg/L. The results observed indicated that the growth of microalgae sample cultivated in NaNO3 concentration of 100 mg/L and NaCl concentration of 25 g/L is higher compared to that cultivated in other concentration of NaNO3 and NaCl, respectively after being analysed by the cell optical density. The specific growth rate shows similar results as treatment of 100 mg/L NaNO3 and 25 g/L NaCl have highest growth rate. The FAMEs content of different cultivation condition produces different composition of saturated fatty acids (SFAs), mono-unsaturated fatty acids (MUFAs) and poly-unsaturated fatty acids (PUFAs).

Valorisation of the microalgae Nannochloropsis gaditana biomass by proteomic approach in the context of circular economy

Nannochloropsis gaditana is a non-flagellated microalgae that has been widely used for different purposes, mostly related with the industrial production of biofuels or aquiculture. However, in order to increase the economic viability of the obtained microalgae biomass from a production plant coupled to a coal power plant, a proteomic approach was initiated by using fresh and atomized microalgae samples, as the main used commercial forms. Above 51,000 high quality spectra were obtained per sample in the MS/MS analysis of whole proteome of N. gaditana, yielding above 7,500 peptides, leading the identification of 1,950 proteins, from the N. gaditana protein database, where 655 proteins were presented in all the replicates. The identified proteins were categorized according to gene ontology classification by molecular function and biological process. In this study, it has been described the first proteomic analysis of the microalgae N. gaditana under industrial conditions containing an important number of identified proteins. A significative presence of proteins with a potential role in different agri-food and biomedical applications was detected and studied being the core of future N. gaditana research to expand the current biotechnological applications of this microalga. Significance of the studyThree quarters of the planet earth correspond to seas and oceans, however its potential biotechnological use is still unknown. We described the first proteomic description of the microalgae N. gaditana under industrial conditions. Following the spirit of the EU initiatives of blue growth and the statements of circular economy, CO2 waste from a coal plant power has been transformed in a resource for microalgae biomass production, common product presentations were evaluated by proteomic, and its potential use of identified proteins in Agri-food and Biomedicine has been revealed.

Microalgae degradation follow up by voltammetric electronic tongue, impedance spectroscopy and NMR spectroscopy

Microalgae play a fundamental role in aquatic primary production and the food chain. They are a recognized source of fatty acids and fatty acid-based lipids of potential interest in the preparation of functional health products, biofuels and renewable chemicals. The exploitation of this bioresource requires a fine monitoring of each production stage. The aim of this work is the microalgae degradation follow up after the concentration stage at the end of the production process by voltammetric electronic tongue, impedance spectroscopy and 1H NMR spectroscopy. Microalgae samples were allowed to progress along time (from 1 to 23 days). At scheduled selected times, voltammetry, impedance spectroscopy and 1H NMR measurements were performed. Multivariate analysis was carried out on these data by PLSR. A model calculated in a training set was then applied to a set of validation to predict the time of evolution. For the three techniques good results in prediction for the validation set were obtained (R2/RMSEP of 0.961/1.51, 0.956/1.67 and 0.969/1.25 respectively for impedance, voltammetry and NMR spectroscopy). The three techniques were sensitive to the evolution of the microalgae samples. The detection of metabolical changes in the 1H NMR spectra is also included. This proof of concept could be the basis for future development of rapid and robust strategies for quality control on microalgae production plants.

Visualisation of the obligate hydrocarbonoclastic bacteria Polycyclovorans algicola and Algiphilus aromaticivorans in co-cultures with micro-algae by CARD-FISH

Some studies have described the isolation and 16S rRNA gene sequence-based identification of hydrocarbon-degrading bacteria living associated with marine eukaryotic phytoplankton, and thus far the direct visual observation of these bacteria on micro-algal cell surfaces (‘phycosphere’) has not yet been reported. Here, we developed two new 16S rRNA-targeted oligonucleotide probes, PCY223 and ALGAR209, to respectively detect and enumerate the obligate hydrocarbonoclastic bacteria Polycyclovorans algicola and Algiphilus aromaticivorans by Catalyzed Reporter Deposition Fluorescence in situ Hybridization (CARD-FISH). To enhance the hybridization specificity with the ALGAR209 probe, a competitor probe was developed. These probes were tested and optimized using pure cultures, and then used in enrichment experiments with laboratory cultures of micro-algae exposed to phenanthrene, and with coastal water enriched with crude oil. Microscopic analysis revealed these bacteria are found in culture with the micro-algal cells, some of which were found attached to algal cells, and whose abundance increased after phenanthrene or crude oil enrichment. These new probes are a valuable tool for identifying and studying the ecology of P. algicola and A. aromaticivorans in laboratory and field samples of micro-algae, as well as opening new fields of research that could harness their ability to enhance the bioremediation of contaminated sites.