The role of commercial probiotic (Bacillus subtilis and Bacillus coagulans) and halophilic (Bacillus sp.) bacteria isolated from Urmia Lake on water quality improvement, biofloc production and composition using different carbon sources

Document Type : Research Paper


1 Ph.D. Student in Aquaculture, Artemia and Aquaculture Research Institute, Urmia University, Urmia, Iran

2 Associate Professor in Artemia and Aquaculture Research Institute, Urmia University, Urmia, Iran

3 Assistant Professor in Artemia and Aquaculture Research Institute, Urmia University, Urmia, Iran

4 Professor in Artemia Reference Center, Ghent University, Ghent, Belgium


In this study, the role of probiotic and halophilic bacteria were studied in biofloc system using different carbon sources including corn starch (treatments 1 and 4), beetroot molasses (treatments 2 and 5), rice-bran (treatments 3 and 6), with sugar cane vinasses (used in all treatments) at presence of commercial probiotics (Bacillus subtilis and Bacillus coagulans) (treatments 1, 2 and 3) and combination of probiotics and halophylic bacteria (treatments 4, 5 and 6) at salinity of 60±5g/L during 42 days. Minimum concentrations of nitrite and nitrate were recorded in treatment 6 and lowest ammonia in treatment 1. The highest floc volume were produced in treatments 2 and 5 (41-48mL/L). In terms of nutritional value, significantly higher protein (20-25%) and ash (75-80%) were detected in treatments 2, 3, 5 and 6. But significantly higher carbohydrate (64%) was detected in treatments 1 and 4. Fatty acid analysis of the bioflocs showed that treatments 2 and 5 had highest PUFA n-3 (32%) and treatments 3 and 6 had significantly higher MUFA (22-25%). Results of this research showed that different carbon sources had significant effect on biofloc quality, but the use of probiotics separately or in combination with halophilic bacteria had no significant effect on biofloc quality.


APHA. 1989. Standard methods for the examination of water and wastewater. American Public Health Association, Washington. 164P.
Arias-Moscoso J.L., Espinoza-Barron L.G., Miranda-Baeza A., Rivas-Vega M.E. and Nieves-Soto M. 2018. Effect of commercial probiotics addition in a biofloc shrimp farm during the nursery phase in zero water exchange. Aquaculture Reports, 11: 47–52.
Avnimelech Y. 1999. Carbonrnitrogen ratio as a control element in aquaculture systems. Aquaculture, 176: 227–235.
Avnimelech Y. 2007. Feeding with microbial flocs by tilapia in minimal discharge bio-flocs technology ponds. Aquaculture, 264(1-4): 140–147.
Avnimelech Y. 2012. Biofloc Technology: A Practical Guide Book. World Aquaculture Scociety, USA. 272P.
Avnimelech Y. 2017. Adapting biofloc technology for use in small adapting biofloc technology for use in small-scale ponds with vertical substrate. World Aquaculture, 2017: 54–58.
Avnimelech Y., Mokady S. and Schroeder G.L. 1989. Circulated ponds as efficient bioreactors for single cell protein production. Israeli Journal of Aquaculture, 41(2): 58–66.
Azim M.E., Little D.C. and Bron J.E. 2008. Microbial protein production in activated suspension tanks manipulating C:N ratio in feed and the implications for fish culture. Bioresource Technology, 99: 3590–3599.
Brown M.R., Barrett S.M., Volkman J.K., Nearhos S.P., Nell J.A. and Allan G.L. 1996. Biochemical composition of new yeasts and bacteria evaluated as food for bivalve aquaculture. Aquaculture, 143(3–4): 341–360.
Burja A.M. and Radianingtyas H. 2007. Nutraceuticals and functional foods from marine microbes: An introduction to a diverse group of natural products isolated from marine macroalgae, microalgae, bacteria, fungi, and cyanobacteria. P: 367–404. In: Barrow C. and Shahidi F. (Eds.). Marine Nutraceuticals and Functional Foods. CRC Press, Taylor and Francis Group, UK.
Crab R., Lambert A., Defoirdt T., Bossier P. and Verstraete W. 2010. The application of bioflocs technology to protect brine shrimp (Artemia franciscana) from pathogenic Vibrio harveyi. Journal of Applied Microbiology, 109(5): 1643–1649.
De Paiva Maia E., Alves Modesto G., Otavio Brito L., Olivera Galvez A. and Vasconcelos Gesteira T.C. 2016. Intensive culture system of Litopenaeus vannamei in commercial ponds with zero water exchange and addition of molasses and probiotics. Revista de Biologia Marina y Oceanografia, 51(1): 61–67.
Decamp O., Moriarty D.J.W. and Lavens P. 2008. Probiotics for shrimp larviculture: Review of field data from Asia and Latin America. Aquaculture Research, 39(4): 334–338.
Defoirdt T., Sorgeloos P. and Bossier P. 2011. Alternatives to antibiotics for the control ofbacterial disease in aquaculture. Current Opinion in Microbiology, 14(3): 251–258.
Deng M., Chen J., Gou J., Hou J., Li D. and He X. 2018. The effect of different carbon sources on water quality, microbial community and structure of biofloc systems. Aquaculture, 482: 103–110.
Dube M.A., Tremblay A.Y. and Liu J. 2007. Biodiesel production using a membrane reactor. Bioresource Technology, 98: 639–647.
Effendy I., Al Deen S. and Chithambaran S. 2016. Semi intensive and semi biofloc methods for the culture of Indian white prawn, Fenneropenaeus indicus in high-density polyethylene liner ponds. Hayati Journal of Biosciences, 23: 106–110.
Ekasari J., Angela D., Waluyo S.H., Bachtiar T., Surawidjaja E.H., Bossier P. and De Schryver P. 2014. The size of biofloc determines the nutritional composition and the nitrogen recovery by aquaculture animals. Aquaculture, 426-427: 105–111.
Ekasari J., Crab R. and Verstraete W. 2010. Primary nutritional content of bio-flocs cultured with different organic carbon sources and salinity. Hayati Journal of Biosciences, 17(3): 125–130.
Emerenciano M., Gaxiola G. and Cuzo G. 2013. Biofloc Technology (BFT): A Review for Aquaculture Application and Animal Food Industry. P: 301–327. In: Matovic M.D. (Ed.). Biomass Now- Cultivation and Utilization. IntechOpen, UK.
Emerenciano M.G.C., Martinez-Cordova L.R., Martinez-Porchas M. and Miranda-Baeza A. 2017. Biofloc Technology (BFT): A Tool for Water Quality Management in Aquaculture. P: 91–109. In: Tutu H. (Ed.). IntechOpen, UK.
Esiobu N., Armenta L. and Ike J. 2002. Antibiotic resistance in soil and water environments. International Journal of Environmental Health Research, 12: 133–144.
Gomes Vilani F., Schveitzer R., Da Fonseca Arantes R., Do Nascimento Vieira F., Manoel Do Espirito Santo C. and Quadros Seiffert W. 2016. Strategies for water preparation in a biofloc system: Effects of carbon source and fertilization dose on water quality and shrimp performance. Aquacultural Engineering, 74: 70–75.
Hapsari F. 2016. The effect of fermented and non fermented biofloc inoculated with bacterium Bacillus cereus for catfish (Clarias gariepinus) juveniles. AACL Bioflux, 9(2): 334–339.
Hargreaves J.A. 2006. Photosynthetic suspended-growth systems in aquaculture. Aquacultural Engineering, 34(3): 344–363.
Hu X., Cao Y., Wen G., Zhang X., Xu Y., Xu W., Li Z. 2016. Effect of combined use of Bacillus and molasses on microbial communities in shrimp cultural enclosure systems. Aquaculture Research, 48(6): 2691–2705.
Jiao W., Liying S. and Yuangao D. 2014. Effects of Bioflocs on Artemia Growth and Water Quality. Acta Geologica Sinica, 88: 111–113.
Ju Z.Y., Forster I., Conquest L., Dominy W., Kuo W.C. and Horgen F.D. 2008. Determination of microbial community structures of shrimp floc cultures by biomarkers and analysis of floc amino acid profiles. Aquaculture Research, 39: 118–133.
Kaneda T. 1977. Fatty acids of the genus Bacillus: An example of branched-chain preference. American Society for Microbiology, 41(2): 391–418.
Krummenauer D., Poersch L., Romano L.A., Gabriele R.L., Encarnacao P. and Wilson Wasielesky J.R. 2014. The Effect of probiotics in a Litopenaeus vannamei biofloc culture system infected with Vibrio parahaemolyticus. Journal of Applied Aquaculture, 26(4): 370–379.
Lepage G. and Roy C.C. 1984. Improved recovery of fatty acid through direct transesterification without prior extraction or purification. Journal of Lipid Research, 25(12): 1391–1396.
Lopes-dos-Santos R.M.A., Groot R., Liying S., Bossier P. and Van Stappen G. 2019. Halophilic bacteria as a food source for the brine shrimp Artemia. Aquaculture, 17: 633–659.
Lopez-Elias J.A., Moreno-Arias A., Miranda-Baeza A., Martinez-Cordova L.R., Rivas-Vega M.E. and Marquez-Rios E. 2015. Proximate composition of bioflocs in culture systems containing hybrid red tilapia fed diets with varying levels of vegetable meal inclusion. North American Journal of Aquaculture, 77(1): 102–109.
Lowry O., Rosebrough J., Lewis A. and Randal R. 1951. Medicion de proteinas con el reactivo de fenol Folin. The Journal of Biological Chemistry, 193(1): 265–275.
Manush S.M., Pal A.K., Das T. and Mukherjee S.C. 2005. Dietary high protein and vitamin C mitigate stress due to chelate claw ablation in Macrobrachium rosenbergii males. Comparative Biochemistry and Physiology A, 142(1): 10–18.
Martinez-Cordova L.R., Emerenciano M., Miranda-Baeza A. and Martinez-Porchas M. 2014. Microbial-based systems for aquaculture of fish and shrimp: An updated review. Reviews in Aquaculture, 6: 1–18.
Miao S., Zhu J., Zhao C., Sun L., Zhang X. and Chen G. 2017. Effects of C/N ratio control combined with probiotics on the immune response, disease resistance, intestinal microbiota and morphology of giant freshwater prawn (Macrobrachium rosenbergii) effects of C/N ratio control combined with probiotics on the immun. Aquaculture, 476: 125–133.
Michaud L., Blancheton J.P., Bruni V. and Piedrahita R. 2006. Effect of particulate organic carbon on heterotrophic bacterial populations and nitrification efficiency in biological filters. Aquacultural Engineering, 34(3): 224–233.
Nichols D.S. 2003. Prokaryotes and the input of polyunsaturated fatty acids to the marine food web. FEMS Microbiology Letters, 219(1): 1–7.
Nurhatijah N., Muchlisin Z.A., Sarong M.A. and Supriatna A. 2016. Application of biofloc to maintain the water quality in culture system of the tiger prawn (Penaeus monodon). AACL Bioflux, 9(4): 923–928.
Oehmen A., Yuan Z. and Blackall L.L. 2004. Short-term effect of carbon source on the competition of polyphosphate accumulating organisms and glycogen accumulating organisms short-term effects of carbon source on the competition of polyphosphate accumulating organisms and glycogen accumulating orga. Water Science and Technology, 50: 139–144.
Oliveira S. and Pijoan C. 2004. Haemophilus parasuis: New trends on diagnosis, epidemiology and control. Veterinary Microbiology, 99(1): 1–12.
Pacheco-Vega J.M., Cadena-Roa M.A., Leyva-Flores J.A., Zavala-Leal O.I., Perez-Bravo E. and Ruiz-Velazco J.M.J. 2018. Effect of isolated bacteria and microalgae on the biofloc characteristics in the Pacific white shrimp culture. Aquaculture Reports, 11: 24–30.
Ray A.J. and Lotz J.M. 2014. Comparing a chemoautotrophic-based biofloc system and three heterotrophic-based systems receiving different carbohydrate sources. Aquacultural Engineering, 63: 54–61.
Russell N.J. and Nichols D.S. 1999. Polyunsaturated fatty acids in marine bacteria- A dogma rewritten. Microbiology, 145: 767–779.
Salehizadeh H. and Van Loosdrecht M.C.M. 2004. Production of polyhydroxyalkanoates by mixed culture : Recent trends and biotechnological importance. Biotechnology Advances, 22: 261–279.
Santhana Kumar V., Pandey P.K., Anand T., Bhuvaneswari G.R., Dhinakaran A. and Kumar S. 2018. Biofloc improves water, effluent quality and growth parameters of Penaeus vannamei in an intensive culture system. Journal of Environmental Management, 215: 206–215.
Satomi M., Oikawa H. and Yano Y. 2003. Note Shewanella marinintestina sp. nov., Shewanella schlegeliana sp. nov. and Shewanella sairae sp. marine bacteria isolated from sea-animal intestines. International Journal of Systematic and Evolutionary Microbiology, 53: 491–499.
Sekar A. and Packyam M. 2014. Screening, identification and antagonistic activity of halo stable Bacillus sp. Mk22 used as probiotic in Penaeus monodon Fabricius, 1798. African Journal of Food Science, 8(1): 48–53.
Spotte S. 1979. Seawater Aquariums: the Captive Environment. Wiley, USA. 413P.
Tacon A.G.J., Cody J.J., Conquest L.D., Divakaran S., Forster I.P. and Decamp O.E. 2002. Effect of culture system on the nutrition and growth performance of Pacific white shrimp Litopenaeus vannamei (Boone) fed different diets. Aquaculture Nutrition, 8: 121–137.
Van Den Hende S., Claessens L., De Muylder E., Boon N. and Vervaeren H. 2016. Microalgal bacterial flocs originating from aquaculture wastewater treatment as diet ingredient for Litopenaeus vannamei (Boone). Aquaculture Research, 47(4): 1075–1089.
Wei Y., Liao S.A. and Wang A. 2016. The effect of different carbon sources on the nutritional composition, microbial community and structure of bioflocs. Aquaculture, 465: 88–93.
Wylie J.L. and Currie D.J. 1991. The relative importance of bacteria and algae as food sources for crustacean zooplankton. Limnology and Oceanography, 36(4): 708–728.
Xu W.J., Pan L.Q., Zhao D.H. and Huang J. 2012. Preliminary investigation into the contribution of bioflocs on protein nutrition of Litopenaeus vannamei fed with different dietary protein levels in zero-water exchange culture tanks. Aquaculture, 350-353: 147–153.
Yuangao D., Gaochao X. and Liying S. 2015. Isolation and characterization of halophilic bacteria and archaea from salt ponds in Hangu Saltworks, Tianjin, China. Chinese Journal of Oceanology and Limnology, 33(4): 862–868.