Eicosapentaenoic acid and docosahexaenoic acid production potential of microalgae and their heterotrophic growth

Twenty microalgal strains were investigated in photoautotrophic flask cultures for their potential for eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) production. The highest EPA proportion (% of total fatty acids) was produced by Monodus subterraneus UTEX 151 (34.2%), followed by Chlorella minutissima UTEX 2341 (31.3%) and Phaeodactylum tricornutum UTEX 642 (21.4%). The highest DHA proportion (% of total fatty acids) was obtained in Crypthecodinium cohnii UTEX L1649 (19.9%), followed by Amphidinium carterae UTEX LB 1002 (17.0%) and Thraustochytrium aureum ATCC 28211 (16.1%). Among the 20 strains screened, the EPA yield was high in M. subterraneus UTEX 151 (96.3 mg/L), P. tricornutum UTEX 642 (43.4 mg/L), Chl. minutissima UTEX 2341 (36.7 mg/L), and Por. cruentum UTEX 161 (17.9 mg/L) owing to their relatively high biomass concentrations. The DHA yield was high in C. cohnii UTEX L1649 (19.5 mg/L) and A. carterae UTEX LB 1002 (8.6 mg/L). Heterotrophic growth of these 20 microalgae was also tested on two different carbon sources, acetate and glucose. All microalgae except Nannochloropsis oculata UTEX LB 2164 showed growth on glucose (5 g/L) under heterotrophic conditions. Twelve of them could grow heterotrophically when acetate (1 g/L) was used as their sole carbon and energy source.     

Three Novel Bacteria Associated with Two Centric Diatom Species from the Mediterranean Sea,Thalassiosira rotula and Skeletonema marinoi

Diatoms are a successful group of microalgae at the base of the marine food web. For hundreds of millions of years, they have shared common habitats with bacteria, which favored the onset of interactions at different levels, potentially driving the synthesis of biologically active molecules. To unveil their presence, we sequenced the genomes of bacteria associated with the centric diatom Thalassiosira rotula from the Gulf of Naples. Annotation of the metagenome and its analysis allowed the reconstruction of three bacterial genomes that belong to currently undescribed species. Their investigation showed the existence of novel gene clusters coding for new polyketide molecules, antibiotics, antibiotic-resistance genes and an ectoine production pathway. Real-time PCR was used to investigate the association of these bacteria with three different diatom clones and revealed their preference for T. rotula FE80 and Skeletonema marinoi FE7, but not S. marinoi FE60 from the North Adriatic Sea. Additionally, we demonstrate that although all three bacteria could be detected in the culture supernatant (free-living), their number is up to 45 times higher in the cell associated fraction, suggesting a close association between these bacteria and their host. We demonstrate that axenic cultures of T. rotula are unable to grow in medium with low salinity (<28 ppt NaCl) whereas xenic cultures can tolerate up to 40 ppt NaCl with concomitant ectoine production, likely by the associated bacteria.     

Antibiotic Algae by Chemical Surface Engineering

Chemical cell‐surface engineering is a tool for modifying and altering cellular functions. Herein, we report the introduction of an antibiotic phenotype to the green alga Chlamydomonas reinhardtii by chemically modifying its cell surface. Flow cytometry and confocal microscopy studies demonstrated that a hybrid of the antibiotic vancomycin and a 4‐hydroxyproline oligomer binds reversibly to the cell wall without affecting the viability or motility of the cells. The modified cells were used to inhibit bacterial growth of Gram‐positive Bacillus subtilis cultures. Delivery of the antibiotic from the microalgae to the bacterial cells was verified by microscopy. Our studies provide compelling evidence that 1) chemical surface engineering constitutes a useful tool for the introduction of new, previously unknown functionality, and 2) living microalgae can serve as new platforms for drug delivery.     

Sustainable Hydrogen Photoproduction by Phosphorus-Deprived Marine Green Microalgae Chlorella sp.

Previously it has been shown that green microalga Chlamydomonas reinhardtii is capable of prolonged H₂ photoproduction when deprived of sulfur. In addition to sulfur deprivation (-S), sustained H₂ photoproduction in C. reinhardtii cultures can be achieved under phosphorus-deprived (-P) conditions. Similar to sulfur deprivation, phosphorus deprivation limits O₂ evolving activity in algal cells and causes other metabolic changes that are favorable for H₂ photoproduction. Although significant advances in H₂ photoproduction have recently been realized in fresh water microalgae, relatively few studies have focused on H₂ production in marine green microalgae. In the present study phosphorus deprivation was applied for hydrogen production in marine green microalgae Chlorella sp., where sulfur deprivation is impossible due to a high concentration of sulfates in the sea water. Since resources of fresh water on earth are limited, the possibility of hydrogen production in seawater is more attractive. In order to achieve H₂ photoproduction in P-deprived marine green microalgae Chlorella sp., the dilution approach was applied. Cultures diluted to about 0.5–1.8 mg Chl·L⁻¹ in the beginning of P-deprivation were able to establish anaerobiosis, after the initial growth period, where cells utilize intracellular phosphorus, with subsequent transition to H₂ photoproduction stage. It appears that marine microalgae during P-deprivation passed the same stages of adaptation as fresh water microalgae. The presence of inorganic carbon was essential for starch accumulation and subsequent hydrogen production by microalgae. The H₂ accumulation was up to 40 mL H₂ gas per 1iter of the culture, which is comparable to that obtained in P-deprived C. reinhardtii culture.     

Boronic Acid Moieties Stabilize Adhesion of Microalgal Biofilms on Glassy Substrates: A Chemical Tool for Environmental Applications

Photosynthetic organisms such as diatoms microalgae provide innovative routes to eco-friendly technologies for environmental pollution bioremediation. Living diatoms are capable to incorporate in vivo a wide variety of chemical species dispersed in seawater, thus being promising candidates for eco-friendly removal of toxic contaminants. However, their exploitation requires immobilization methods that allow to confine microalgae during water treatment. Here we demonstrate that a biofilm of Phaeodactylum tricornutum diatom cells grown on the surface of a glassy substrate bearing boronic acid protruding moieties is stably anchored to the substrate resisting mechanical stress and it is suitable for removal of up to 80 % metal ions (As, Cr, Cu, Zn, Sn, Pb, Sb) in a model polluted water sample. Control experiments also suggest that stabilization of the biofilm adhesion occurs by interaction of boronic acid surface groups of the substrate with the hydroxyl groups of diatoms extracellular polysaccharides.     

Long-Term Heat Selection of the Coral Endosymbiont Cladocopium C1acro (Symbiodiniaceae) Stabilizes Associated Bacterial Communities

Heat-tolerant strains of the coral endosymbiont, Cladocopium C1^acro (Symbiodiniaceae), have previously been developed via experimental evolution. Here, we examine physiological responses and bacterial community composition (using 16S rRNA gene metabarcoding) in cultures of 10 heat-evolved (SS) and 9 wild-type (WT) strains, which had been exposed for 6 years to 31 °C and 27 °C, respectively. We also examine whether the associated bacterial communities were affected by a three-week reciprocal transplantation to both temperatures. The SS strains had bacterial communities with lower diversities that showed more stability and lower variability when exposed to elevated temperatures compared with the WT strains. Amplicon sequence variants (ASVs) of the bacterial genera Labrenzia, Algiphilus, Hyphobacterium and Roseitalea were significantly more associated with the SS strains compared with the WT strains. WT strains showed higher abundance of ASVs assigned to the genera Fabibacter and Tropicimonas. We hypothesize that these compositional differences in associated bacterial communities between SS and WT strains also contribute to the thermal tolerance of the microalgae. Future research should explore functional potential between bacterial communities using metagenomics to unravel specific genomic adaptations.     

Drastic Attenuation of Pseudomonas aeruginosa Pathogenicity in a Holoxenic Mouse Experimental Model Induced by Subinhibitory Concentrations of Phenyllactic acid (PLA)

The discovery of communication systems regulating bacterial virulence has afforded a novel opportunity to control infectious bacteria without interfering with growth. In this paper we describe the effect of subinhibitory concentrations of phenyllactic acid (PLA) on the pathogenicity of Pseudomonas aeruginosa in mice. The animals were inoculated by oral (p.o.), intranasal (i.n.), intravenous (i.v.) and intraperitoneal (i.p.) routes with P. aeruginoasa wild and PLA-treated cultures. The mice were followed up during 16 days after infection and the body weight, mortality and morbidity rate were measured every day. The microbial charge was studied by viable cell counts in lungs, spleen, intestinal mucosa and blood. The mice batches infected with wild P. aeruginosa bacterial cultures exhibited high mortality rates (100 % after i.v. and i.p. route) and very high cell counts in blood, lungs, intestine and spleen. In contrast, the animal batches infected with PLA treated bacterial cultures exhibited good survival rates (0 % mortality) and the viable cell counts in the internal organs revealed with one exception the complete abolition of the invasive capacity of the tested strains. In this study, using a mouse infection model we show that D-3-phenyllactic acid (PLA) can act as a potent antagonist of Pseudomonas (P.) aeruginosa pathogenicity, without interfering with the bacterial growth, as demonstrated by the improvement of the survival rates as well as the clearance of bacterial strains from the body.     

Isolation and characterization of autoflocculating mutants of cyanobacterium Arthrospira platensis

Harvesting microalgae is a major concern for mass culture in industry. Flocculation is an easy and effective way to harvest microalgae. However, flocculation using chemical flocculants is not feasible for scaling-up due to their toxicity. As an alternative technique, mutation breeding of autoflocculating microalgae strain has been reported in this study. We characterized autoflocculating mutants of Arthrospira platensis (A. platensis) by ethyl methane sulfonate (EMS). The cells of mutants were aggregated during the culture and dry cell weight increased 1.2- to 1.8-fold compared to the wild type. Autoflocculation was induced highly at an optimal pH level of 9 and the flocculation efficiency reached almost 90%. Mutants showed higher flocculation efficiency irrespective of the addition of chemical flocculants. Thus, it is definitely useful to harvest microalgae using autoflocculating mutants in large-scale culture without any drawbacks of harvested algal biomass.     

Evaluation of Nitrogen Fixing Enzyme Activities in Response to Pyrene Bioremediation Efficacy by Defined Artificial Microalgal-Bacterial Consortium of Gujarat,India

This study was carried out to investigate the response and relationship between nitrogen fixing enzymes during the remediation of different concentrations of high molecular weight four rings Polynuclear Aromatic Hydrocarbon (PAH) Pyrene by microalgae Synechocystis sp. (cyanobacteria) with artificial developed indigenous bacterial consortium. One axenic microalgal culture of Synechocystis sp. and two pyrene degrading indigenous bacteria with efficient removal capabilities viz. Pseudomonas indoxyladons and Bacillus benzoevorans isolated from crude oil polluted site and common industrial effluent canal were used to construct the consortium. The effect of pyrene on algal growth in terms of chlorophyll-a was measured and it was found that in the presence of bacteria, the growth and bioremediation capacity of Synechocystis sp. raised tremendously, whereas Synechocystis sp. monoculture exhibited concentration dependent decrease. Moreover, the nitrogen fixing enzymes; nitrate reductase (NR), glutamine synthetase (GS), and succinate dehydrogenase (SDH) showed chronological decrease by 93%, 90%, and 98%, respectively. Increased Bioremediation of pyrene by consortium JPNKA7B2 (Mix culture of Synechocystis sp., Pseudomonas indoxyladons, and Bacillus benzoevorans) was eliminated at 94.1% in 50 mg/L, which indirectly retarded the nitrogen fixing enzymes – NR, GS, and SDH. However, Synechocystis sp. monoculture could remediate up to 36% at 1.5 mg/L after 16 days of incubation.     

Effect of bacterial inoculum ratio in mixed culture for decolourization and detoxification of pulp paper mill effluent

BACKGROUND: Effluent released from industry is a mixture of various pollutants. For the degradation of complex pollutants, mixed bacterial cultures can be more effective than a single culture. This study investigated the balance of bacterial populations in a mixed culture for maximum reduction of pollutants. RESULTS: This study deals with the degradation and detoxification of pulp paper mill effluent (PPME) by three bacterial strains, i.e. Serratia marcescens, Serratia liquefaciens and Bacillus cereus in different ratios, and found that two ratios, 4:1:1 and 1:4:1, were effective for the degradation of PPME. These ratios reduced the various pollution parameters. Enzyme bioassay revealed that more enzyme was produced during degradation for the ratio 4:1:1. High performance liquid chromatography (HPLC) analysis showed that the ratio 4:1:1 degraded 95% of lignin and related compounds, and chlorophenols up to 98%, whereas ratio 1:4:1 reduced lignin by 84% and chlorophenols by 58% after 7 days incubation. Degradation products were confirmed by gas chromatography–mass spectrometry (GC‐MS) analysis. A seed germination bioassay on Phaseolous mungo L. revealed that toxicity was reduced by the ratio 4:1:1. CONCLUSION: Due to variable potential of different bacteria show variation in their growth pattern at any contaminated site. This study shows that an appropriate ratio of mixed cultures is required for maximum degradation and detoxification of PPME. Copyright © 2012 Society of Chemical Industry     

Synthesis and characterization of poly(styrene-co-fluorescein O-methacrylate)/poly(N-isopropylacrylamide)-Fe3O4 core/shell composite particles

We demonstrate the synthesis and characteristics of multifunctional poly(styrene-co-fluorescein O-methacrylate)/poly(N-isopropylacrylamide)-Fe₃O₄ [P(St/FMA)/PNIPAAm-Fe₃O₄] core/shell composite particles, in which the core consists of fluorescent materials and the shell consists of magnetic and thermo-responsive components. First, core/shell particles consisting of a fluorescent P(St/FMA) core and thermo-responsive PNIPAAm-rich shell were prepared by two-stage shot-growth emulsion polymerization. Next, Fe₃O₄ nanoparticles were immobilized via electrostatic interactions and then covalently linked to the shell via surface coordinated Aphen by a coupling reaction in order to obtain magnetic properties. The morphology of P(St/FMA)/PNIPAAm-Fe₃O₄ composite particles, confirmed by transmission electron microscopy (TEM), reveals that Fe₃O₄ nanoparticles are located in the PNIPAAm shell. The thermo-sensitivity of composite particles to hydrodynamic diameter was confirmed by using dynamic light scattering (DLS). Photoluminescence (PL) spectra indicate that the fluorescence emission intensity of core/shell particles is highly sensitive to the pH of an aqueous medium. The core/shell composite particles exhibited a combination of fluorescent, magnetic, pH and thermo-responsive behavior.     

Fucoxanthin,a Marine-Derived Carotenoid from Brown Seaweeds and Microalgae: A Promising Bioactive Compound for Cancer Therapy

Fucoxanthin is a well-known carotenoid of the xanthophyll family, mainly produced by marine organisms such as the macroalgae of the fucus genus or microalgae such as Phaeodactylum tricornutum. Fucoxanthin has antioxidant and anti-inflammatory properties but also several anticancer effects. Fucoxanthin induces cell growth arrest, apoptosis, and/or autophagy in several cancer cell lines as well as in animal models of cancer. Fucoxanthin treatment leads to the inhibition of metastasis-related migration, invasion, epithelial–mesenchymal transition, and angiogenesis. Fucoxanthin also affects the DNA repair pathways, which could be involved in the resistance phenotype of tumor cells. Moreover, combined treatments of fucoxanthin, or its metabolite fucoxanthinol, with usual anticancer treatments can support conventional therapeutic strategies by reducing drug resistance. This review focuses on the current knowledge of fucoxanthin with its potential anticancer properties, showing that fucoxanthin could be a promising compound for cancer therapy by acting on most of the classical hallmarks of tumor cells.     

Central Carbon Metabolism,Sodium-Motive Electron Transfer,and Ammonium Formation by the Vaginal Pathogen Prevotella bivia

Replacement of the Lactobacillus dominated vaginal microbiome by a mixed bacterial population including Prevotella bivia is associated with bacterial vaginosis (BV). To understand the impact of P. bivia on this microbiome, its growth requirements and mode of energy production were studied. Anoxic growth with glucose depended on CO₂ and resulted in succinate formation, indicating phosphoenolpyruvate carboxylation and fumarate reduction as critical steps. The reductive branch of fermentation relied on two highly active, membrane-bound enzymes, namely the quinol:fumarate reductase (QFR) and Na⁺-translocating NADH:quinone oxidoreductase (NQR). Both enzymes were characterized by activity measurements, in-gel fluorography, and VIS difference spectroscopy, and the Na⁺-dependent build-up of a transmembrane voltage was demonstrated. NQR is a potential drug target for BV treatment since it is neither found in humans nor in Lactobacillus. In P. bivia, the highly active enzymes L-asparaginase and aspartate ammonia lyase catalyze the conversion of asparagine to the electron acceptor fumarate. However, the by-product ammonium is highly toxic. It has been proposed that P. bivia depends on ammonium-utilizing Gardnerella vaginalis, another typical pathogen associated with BV, and provides key nutrients to it. The product pattern of P. bivia growing on glucose in the presence of mixed amino acids substantiates this notion.     

Nitrate reducing CaCO3 precipitating bacteria survive in mortar and inhibit steel corrosion

Microbial healing of concrete cracks is a relatively slow process, and meanwhile the steel rebar is exposed to corrosive substances. Nitrate reducing bacteria can inhibit corrosion and provide crack healing, by simultaneously producing NO₂⁻ and inducing CaCO₃ precipitation. In this study, the functionality of one non-axenic and two axenic NO₃⁻ reducing cultures for the development of corrosion resistant self-healing concrete was investigated. Both axenic cultures survived in mortar when incorporated in protective carriers and became active 3 days after the pH dropped below 10. The non-axenic culture named “activated compact denitrifying core” (ACDC) revealed comparable resuscitation performance without any additional protection. Moreover, ACDC induced passivation of the steel in corrosive electrolyte solution (0.05 M NaCl) by producing 57 mM NO₂⁻ in 1 week. The axenic cultures produced NO₂⁻ up to 26.8 mM, and passivation breakdown and pitting corrosion were observed. Overall, ACDC appears suitable for corrosion resistant microbial self-healing concrete.     

Chemical Characterization of Six Microalgae with Potential Utility for Food Application

Microalgae contain high levels of proteins, carbohydrates, and lipids, and have found a useful application in enhancing the nutritional value of foods. These organisms can also synthesize long‐chain fatty acids in the form of triacylglycerols, such as α‐linolenic acid (ALA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), linolenic acid (LA), γ‐linolenic acid (GLA) and arachidonic acid (AA). The aim of this study was to determine the chemical composition and measure protein, carbohydrates, fibers, lipids as well as the fatty acids composition of six microalgae species with potential application in the food industry. Two freshwater species, Chlorella vulgaris and Spirulina platensis, and four marine species, Nannochloropsis oculata, Nannochloropsis gaditana, Porphyridium cruentum, and Phaeodactylum tricornutum, were used in the experiments. Intracellular protein was the most prominent algal component (42.8–35.4 %), followed by carbohydrate + fiber (32.3–28.6 %), and lipids (15.6–5.3 %). N. gaditana is rich in saturated fatty acids, mainly palmitic acid (5.1 g/100 g), while the cells of S. platensis and C. vulgaris algae are abundant in GLA (1.9 g/100 g) and ALA (2.8 g/100 g) acids, respectively. P. cruentum differs from other algae, because it contains a large amount of AA (3.7 g/100 g). The marine microorganisms N. oculata and P. tricornutum are also a source of essential long‐chain polyunsaturated fatty acids (LC‐PUFA‐?3), mainly composed of EPA and DHA. Our results suggest that the freshwater species C. vulgaris and S. platensis are attractive nutritional supplements because of their low fiber and high protein/carbohydrate contents, while the marine species P. tricornutum and N. oculata can enrich foods with LC‐PUFA‐ω3, because of their favorable ω3/ω6 ratio.     

Polycaprolactone degradation by mixed and pure cultures of bacteria and a yeast

The degradability of three high molecular weight polycaprolactones (M?_w = 35,000, 18,600, and 7,130) and one low molecular weight polycaprolactone diol(M?_w = 2060) by mixed and pure cultures of microorganisms was assayed. A yeast, Cryptococcus laurentii, a gram-negative rod, Acinetobacter calcoaceticus var. lwoffi, and a gram-positive coryneform rod were used in the pure culture assays. The analysis of degradation by gel permation chromatography (GPC) allowed for quantitation independent of the growth of the organisms or the addition of supplementary growth factors. GPC analysis showed that the degradation effected by pure cultures was often enhanced when alternate carbon sources were present. This was not the case for mixed cultures. Mixed cultures. Mixed cultures completely metabolized polymer breakdown products while in some cases pure cultures did not.     

Subcritical co‐solvents extraction of lipid from wet microalgae pastes of Nannochloropsis sp.

In this paper subcritical co‐solvents extraction (SCE) of algal lipid from wet pastes of Nannochloropsis sp. is examined. The influences of five operating parameters including the ratio between ethanol to hexane, the ratio of mixed solvents to algal biomass (dry weight), extraction temperature, pressure, and time were investigated. The determined optimum extraction conditions were 3:1 (hexane to ethanol ratio), 10:1 ratio (co‐solvents to microalgae (dry weight) ratio), 90°C, 1.4 MPa, and 50 min, which could produce 88% recovery rate of the total lipids. In addition, electron micrographs of transmission electron microscopy (TEM) and scanning electron microscopy (SEM) were conducted to show that the algal cell presented shrunken, collapsed with some wrinkles and microholes after SCE extraction. The main composition of total lipids extracted under the optimum conditions was TAG which represented more than 80%. And the fatty acid profile of triglycerides revealed that C16:0 (35.67 ± 0.2%), C18:1 (26.84 ± 0.044%) and C16:1 (25.96 ± 0.011%) were dominant. Practical applications: The reported method could save energy consumption significantly through avoiding deep dewatering (for example drying). The composition of the extracted lipid is suitable for the production of high quality biodiesel.     

Contrasting Health Effects of Bacteroidetes and Firmicutes Lies in Their Genomes: Analysis of P450s,Ferredoxins, and Secondary Metabolite Clusters

Species belonging to the bacterial phyla Bacteroidetes and Firmicutes represent over 90% of the gastrointestinal microbiota. Changes in the ratio of these two bacterial groups were found to have contrasting health effects, including obesity and inflammatory diseases. Despite the availability of many bacterial genomes, comparative genomic studies on the gene pools of these two bacterial groups concerning cytochrome P450 monooxygenases (P450s), ferredoxins, and secondary metabolite biosynthetic gene clusters (smBGCs) are not reported. This study is aimed to address this research gap. The study revealed the presence of diverse sets of P450s, ferredoxins, and smBGCs in their genomes. Bacteroidetes species have the highest number of P450 families, ferredoxin cluster-types, and smBGCs compared to Firmicutes species. Only four P450 families, three ferredoxin cluster types, and five smBGCs are commonly shared between these two bacterial groups. Considering the above facts, we propose that the contrasting effects of these two bacterial groups on the host are partly due to the distinct nature of secondary metabolites produced by these organisms. Thus, the cause of the contrasting health effects of these two bacterial groups lies in their gene pools.     

The Chitinolytic Activities of Streptomyces sp. TH-11

Chitin is an abundant biopolymer composed of units of N-acetyl-D-glucosamine linked by β-1,4 glycosidic bonds. Chitin is the main component of the shells of mollusks, the cell wall of fungi and yeast and of the exoskeleton of crustaceans and insects. The degradation of chitin is catalyzed by chitinases that occur in a wide range of organisms. Among them, the chitinases from microorganisms are extremely important for the degradation and recycling of the carbon and nitrogen trapped in the large amount of insoluble chitin in nature. Streptomyces sp. TH-11 was isolated from the sediment of the Tou-Chien River, Taiwan. The chitinolytic enzyme activities were detected using a rapid in-gel detection method from the cell-free preparation of the culture medium of TH-11. The chitinolytic enzyme activity during prolonged liquid culturing was also analyzed by direct measurement of the chitin consumption. Decomposition of the exoskeleton of shrimps was demonstrated using electron microscopy and atomic force microscopy.     

Inhibiting biofilm formation of Enterobacter sp. prevented premature withering in cut flowers

Early withering and premature flower fall are a growing menace to the cut flower industry, the reasons of which were considered to be varied from loss of water uptake, bacterial proliferation, decay in tap water etc. In the present study, we identified the bacterial biofilm formation by Enterobacter sp. and subsequent water uptake blockage as the root cause of early withering in cut flowers using Chrysanthemum, Yellow Daisy and Maroon Rose as model plants. The biofilm-forming Enterobacter sp was identified by 16 S rDNA sequencing data. Studies on biofilm were conducted by using field emission scanning electron microscope electron back scattering diffraction (FE-SEM-EBSD), Atomic force microscopy (AFM) and fluorescence microscopy. In vitro and in vivo studies were conducted with different antimicrobials to prevent biofilm formation in both conditions. Most antimicrobials were toxic to plants, but we found citric acid 1,000 μg/ml and calcium hypochlorite 50 μg/ml to be most effective in preventing biofilm formation and extending the vase life of cut flowers. We studied the synergistic action of different combinations in vivo and suggest citric acid 1,000 μg/mL, Ca hypochlorite 50 μg/mL and glucose 1,000 μg/mL as the best combination to be used for prolonging vase life of cut flowers from 10 days (non-treated) to 30 days (treated).