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Bacterial abundance and diversity in Microchloropsis salina (formerly Nannochloropsis salina) cultures in response to the presence of ammonium, nitrate and glycerol

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Abstract

Ammonium and nitrate were compared as nitrogen sources for mixotrophic growth of Microchloropsis salina (formerly Nannochloropsis salina) on glycerol. The effects of nitrogen source and glycerol on the abundance, composition and profile of bacterial communities were examined by 16S rRNA sequencing. The effect of bacteria on culture performance was assessed by comparing ampicillin-containing and ampicillin-free cultures. The productivity of M. salina was higher with ammonium than with nitrate, and lower with bacteria present. Glycerol utilization by M. salina was minimal in the absence of a nitrogen source and the addition of glycerol increased the abundance of bacteria. Bacterial profiles were influenced by nitrogen source, with Pseudomonadaceae proliferating with ammonium, Alteromonadaceae with nitrate and Beijerinckiaceae prevalent with either. The presence of nitrate and glycerol led to aerobic-denitrifying bacteria capable of diverting both resources away from algal metabolism. These findings demonstrate significant advantages of providing ammonium during mixotrophic cultivation of M. salina on glycerol.

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References

  • Alcantara C, Fernandez C, Garcia-Encina PA, Munoz R (2015) Mixotrophic metabolism of Chlorella sorokiniana and algal-bacterial consortia under extended dark-light periods and nutrient starvation. Appl Microbiol Biotechnol 99:2393–2404

    CAS  PubMed  Google Scholar 

  • Allen DK (2016) Assessing compartmentalized flux in lipid metabolism with isotopes. Biochim Biophys Acta 1861:1226–1242

    CAS  PubMed  Google Scholar 

  • Anderson KE, Russell JA, Moreau CS, Kautz S, Sullam KE, Hu Y, Basinger U, Mott BM, Buck N, Wheeler DE (2012) Highly similar microbial communities are shared among related and trophically similar ant species. Mol Ecol 21:2282–2296

    PubMed  Google Scholar 

  • Andriukonis E, Gorokhova E (2017) Kinetic 15N-isotope effects on algal growth. Sci Rep 7:44181

    PubMed  PubMed Central  Google Scholar 

  • Chen H, Zheng Y, Zhan J, He C, Wang Q (2017) Comparative metabolic profiling of the lipid-producing green microalga Chlorella reveals that nitrogen and carbon metabolic pathways contribute to lipid metabolism. Biotechnol Biofuels 10:153

    PubMed  PubMed Central  Google Scholar 

  • Croft MT, Lawrence AD, Raux-Deery E, Warren MJ, Smith AG (2005) Algae acquire vitamin B12 through a symbiotic relationship with bacteria. Nature 438:90–93

    CAS  PubMed  Google Scholar 

  • Danger M, Oumarou C, Benest D, Lacroix G (2007) Bacteria can control stoichiometry and nutrient limitation of phytoplankton. Funct Ecol 21:202–210

    Google Scholar 

  • de Oliveira ZM, Floh EIS, Ferrara FIS, Barbosa HR (2011) Diazotrophyc rhizobacteria isolated from sugarcane can release amino acids in a synthetic culture medium. Biol Fertil Soils 47:957–962

    CAS  Google Scholar 

  • Delmont TO et al (2018) Nitrogen-fixing populations of Planctomycetes and Proteobacteria are abundant in surface ocean metagenomes. Nat Microbiol 3:804–813

    CAS  PubMed  PubMed Central  Google Scholar 

  • Doyon G, Gaudreau G, Stgelais D, Beaulieu Y, Randall CJ (1991) Simultaneous HPLC determination of organic-acids, sugars and alcohols. Can I Food Sci Tech J 24:87–94

    CAS  Google Scholar 

  • Fawley MW, Jameson I, Fawley KP (2015) The phylogeny of the genus Nannochloropsis (Monodopsidaceae, Eustigmatophyceae), with descriptions of N. australis sp. nov. and Microchloropsis gen. nov. Phycologia 54:545–552

    CAS  Google Scholar 

  • Fulbright SP, Robbins-Pianka A, Berg-Lyons D, Knight R, Reardon KF, Chisholm ST (2018) Bacterial community changes in an industrial algae production system. Algal Res 31:147–156

    PubMed  PubMed Central  Google Scholar 

  • Gonzalez LE, Bashan Y (2000) Increased growth of the microalga Chlorella vulgaris when coimmobilized and cocultured in alginate beads with the plant-growth-promoting bacterium Azospirillum brasilense. Appl Environ Microbiol 66:1527–1531

    CAS  PubMed  PubMed Central  Google Scholar 

  • Griessmeier V, Bremges A, McHardy AC, Gescher J (2017) Investigation of different nitrogen reduction routes and their key microbial players in wood chip-driven denitrification beds. Sci Rep 7:17028

    PubMed  PubMed Central  Google Scholar 

  • Gutierrez J, Kwan TA, Zimmerman JB, Peccia J (2016) Ammonia inhibition in oleaginous microalgae. Algal Res 19:123–127

    Google Scholar 

  • Han J, Zhang L, Wang S, Yang G, Zhao L, Pan K (2016) Co-culturing bacteria and microalgae in organic carbon containing medium. J Biol Res (Thessalon) 23:8

    Google Scholar 

  • Hernandez J-P, de-Bashan LE, Rodriguez DJ, Rodriguez Y, Bashan Y (2009) Growth promotion of the freshwater microalga Chlorella vulgaris by the nitrogen-fixing, plant growth-promoting bacterium Bacillus pumilus from arid zone soils. Eur J Soil Biol 45:88–93

    CAS  Google Scholar 

  • Higgins BT, Gennity I, Samra S, Kind T, Fiehn O, VanderGheynst JS (2016) Cofactor symbiosis for enhanced algal growth, biofuel production, and wastewater treatment. Algal Res 17:308–315

    Google Scholar 

  • Higgins BT, Labavitch JM, VanderGheynst JS (2015) Co-culturing Chlorella minutissima with Escherichia coli can increase neutral lipid production and improve biodiesel quality. Biotechnol Bioeng 112:1801–1809

    CAS  PubMed  Google Scholar 

  • Higgins BT, VanderGheynst JS (2014) Effects of Escherichia coli on mixotrophic growth of Chlorella minutissima and production of biofuel precursors. PLoS One 9:e96807

    PubMed  PubMed Central  Google Scholar 

  • Hii YS, Soo CL, Chuah TS, Ambak M, Abol-Munafi AB (2011) Interactive effect of ammonia and nitrate on the nitrogen uptake by Nannochloropsis sp. J Sustainabil Sci Manage 6:60–68

    CAS  Google Scholar 

  • Law SQK, Halim R, Scales PJ, Martin GJO (2018) Conversion and recovery of saponifiable lipids from microalgae using a nonpolar solvent via lipase-assisted extraction. Bioresour Technol 260:338–347

    CAS  PubMed  Google Scholar 

  • Leflaive J, Danger M, Lacroix G, Lyautey E, Oumarou C, Ten-Hage L (2008) Nutrient effects on the genetic and functional diversity of aquatic bacterial communities. FEMS Microbiol Ecol 66:379–390

    CAS  PubMed  Google Scholar 

  • Leite GB, Paranjape K, Abdelaziz AEM, Hallenbeck PC (2015) Utilization of biodiesel-derived glycerol or xylose for increased growth and lipid production by indigenous microalgae. Bioresour Technol 184:123–130

    CAS  PubMed  Google Scholar 

  • Li K, Bihan M, Yooseph S, Methe BA (2012) Analyses of the microbial diversity across the human microbiome. PLoS One 7:e32118

    CAS  PubMed  PubMed Central  Google Scholar 

  • Liu H, Zhou Y, Xiao W, Ji L, Cao X, Song C (2012) Shifting nutrient-mediated interactions between algae and bacteria in a microcosm: evidence from alkaline phosphatase assay. Microbiol Res 167:292–298

    CAS  PubMed  Google Scholar 

  • Liu L, Zhao Y, Jiang X, Wang X, Liang W (2018) Lipid accumulation of Chlorella pyrenoidosa under mixotrophic cultivation using acetate and ammonium. Bioresour Technol 262:342–346

    CAS  PubMed  Google Scholar 

  • Liu Y, Ai GM, Miao LL, Liu ZP (2016) Marinobacter strain NNA5, a newly isolated and highly efficient aerobic denitrifier with zero N2O emission. Bioresour Technol 206:9–15

    CAS  PubMed  Google Scholar 

  • Martin GJ, Hill DR, Olmstead IL, Bergamin A, Shears MJ, Dias DA, Kentish SE, Scales PJ, Botté CY, Callahan DL (2014) Lipid profile remodeling in response to nitrogen deprivation in the microalgae Chlorella sp. (Trebouxiophyceae) and Nannochloropsis sp. (Eustigmatophyceae). PLoS One 9:e103389

    PubMed  PubMed Central  Google Scholar 

  • Moon M, Kim CW, Park WK, Yoo G, Choi YE, Yang JW (2013) Mixotrophic growth with acetate or volatile fatty acids maximizes growth and lipid production in Chlamydomonas reinhardtii. Algal Res 2:352–357

    Google Scholar 

  • Nakase G, Eguchi M (2007) Analysis of bacterial communities in Nannochloropsis sp cultures used for larval fish production. Fish Sci 73:543–549

    CAS  Google Scholar 

  • Olmstead IL, Hill DR, Dias DA, Jayasinghe NS, Callahan DL, Kentish SE, Scales PJ, Martin GJ (2013) A quantitative analysis of microalgal lipids for optimization of biodiesel and omega-3 production. Biotechnol Bioeng 110:2096–2104

    CAS  PubMed  Google Scholar 

  • Paerl RW, Hansen TNG, Henriksen N, Olesen AK, Riemann L (2018) N-fixation and related O2 constraints on model marine diazotroph Pseudomonas stutzeri BAL361. Aquat Microbial Ecology 81:125–136

    Google Scholar 

  • Pagnanelli F, Altimari P, Trabucco F, Toro L (2014) Mixotrophic growth of Chlorella vulgaris and Nannochloropsis oculata: interaction between glucose and nitrate. J Chem Technol Biotechnol 89:652–661

    CAS  Google Scholar 

  • Palacios OA, Gomez-Anduro G, Bashan Y, de-Bashan LE (2016) Tryptophan, thiamine and indole-3-acetic acid exchange between Chlorella sorokiniana and the plant growth-promoting bacterium Azospirillum brasilense. FEMS Microbiol Ecol 92:fiw077

    PubMed  Google Scholar 

  • Paranjape K, Leite GB, Hallenbeck PC (2016) Strain variation in microalgal lipid production during mixotrophic growth with glycerol. Bioresour Technol 204:80–88

    CAS  PubMed  Google Scholar 

  • Poddar N, Sen R, Martin GJO (2018) Glycerol and nitrate utilisation by marine microalgae Nannochloropsis salina and Chlorella sp and associated bacteria during mixotrophic and heterotrophic growth. Algal Res 33:298–309

    Google Scholar 

  • Podevin M, De Francisci D, Holdt SL, Angelidaki I (2015) Effect of nitrogen source and acclimatization on specific growth rates of microalgae determined by a high-throughput in vivo microplate autofluorescence method. J Appl Phycol 27:1415–1423

    CAS  Google Scholar 

  • Ramanan R, Kim BH, Cho DH, Oh HM, Kim HS (2016) Algae-bacteria interactions: evolution, ecology and emerging applications. Biotechnol Adv 34:14–29

    CAS  PubMed  Google Scholar 

  • Sanz-Luque E, Chamizo-Ampudia A, Llamas A, Galvan A, Fernandez E (2015) Understanding nitrate assimilation and its regulation in microalgae. Front Plant Sci 6:899

    PubMed  PubMed Central  Google Scholar 

  • Shu W, Pablo GP, Jun Y, Danfeng H (2012) Abundance and diversity of nitrogen-fixing bacteria in rhizosphere and bulk paddy soil under different duration of organic management. World J Microbiol Biotechnol 28:493–503

    PubMed  Google Scholar 

  • Simionato D, Block MA, La Rocca N, Jouhet J, Marechal E, Finazzi G, Morosinotto T (2013) The response of Nannochloropsis gaditana to nitrogen starvation includes de novo biosynthesis of triacylglycerols, a decrease of chloroplast galactolipids, and reorganization of the photosynthetic apparatus. Eukaryot Cell 12:665–676

    CAS  PubMed  PubMed Central  Google Scholar 

  • Subramanian G, Yadav G, Sen R (2016) Rationally leveraging mixotrophic growth of microalgae in different photobioreactor configurations for reducing the carbon footprint of an algal biorefinery: a techno-economic perspective. RSC Adv 6:72897–72904

    CAS  Google Scholar 

  • Sun Y, Feng L, Li A, Zhang X, Yang J, Ma F (2017) Ammonium assimilation: an important accessory during aerobic denitrification of Pseudomonas stutzeri T13. Bioresour Technol 234:264–272

    CAS  PubMed  Google Scholar 

  • Tran NAT, Seymour JR, Siboni N, Evenhuis CR, Tamburic B (2017) Photosynthetic carbon uptake induces autoflocculation of the marine microalga Nannochloropsis oculata. Algal Res 26:302–311

    Google Scholar 

  • Tzollas NM, Zachariadis GA, Anthemidis AN, Stratis JA (2010) A new approach to indophenol blue method for determination of ammonium in geothermal waters with high mineral content. Int J Environ Anal Chem 90:115–126

    CAS  Google Scholar 

  • Vadiveloo A, Moheimani N (2018) Effect of continuous and daytime mixing on Nannochloropsis growth in raceway ponds. Algal Res 33:190–196

    Google Scholar 

  • Venkata Mohan S, Devi MP (2014) Salinity stress induced lipid synthesis to harness biodiesel during dual mode cultivation of mixotrophic microalgae. Bioresour Technol 165:288–294

    CAS  PubMed  Google Scholar 

  • Yap BH, Crawford SA, Dagastine RR, Scales PJ, Martin GJ (2016) Nitrogen deprivation of microalgae: effect on cell size, cell wall thickness, cell strength, and resistance to mechanical disruption. J Ind Microbiol Biotechnol 43:1671–1680

    CAS  PubMed  Google Scholar 

  • Zheng HY, Liu Y, Gao XY, Ai GM, Miao LL, Liu ZP (2012) Characterization of a marine origin aerobic nitrifying-denitrifying bacterium. J Biosci Bioeng 114:33–37

    CAS  PubMed  Google Scholar 

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Acknowledgments

The authors are grateful to the University of Melbourne for a Melbourne International Engagement Award Scholarship for Ms. Poddar. We acknowledge the contribution of Australian Genomic Research Facility (AGRF) in performing the diversity profiling of bacteria.

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Authors and Affiliations

  1. Algal Processing Group, Department of Chemical Engineering, The University of Melbourne, Parkville, Victoria, 3010, Australia

    Nature Poddar & Gregory J. O. Martin

  2. Department of Biotechnology, Indian Institute of Technology Kharagpur, Kharagpur, West Bengal, 721302, India

    Ramkrishna Sen

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Contributions

N. Poddar developed and conducted the experimental program, interpreted the results, drafted, revised and approved the final manuscript. R. Sen contributed to the conceptual development and interpretation of the research and revised the final manuscript. G. Martin assisted in the design of the experimental program, conceptual development, data interpretation and preparation of the manuscript. G. Martin takes responsibility for the integrity of the entire work and can be contacted as gjmartin@unimelb.edu.au.

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Correspondence to Gregory J. O. Martin.

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Poddar, N., Sen, R. & Martin, G.J.O. Bacterial abundance and diversity in Microchloropsis salina (formerly Nannochloropsis salina) cultures in response to the presence of ammonium, nitrate and glycerol. J Appl Phycol 32, 839–850 (2020). https://doi.org/10.1007/s10811-019-02007-0

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