Biotecnología

 

A preliminary study on molecular characterization of the eubacteria in a thermophilic, poultry waste fed anaerobic digester

 

Un estudio preliminar de la caracterización molecular de eubacterias en un digestor anaerobio, termofílico alimentado con desechos de una pollería

 

N. Balagurusamy*

 

Escuela de Ciencias Biológicas, Carretera Torreon-Matamoros Km 7.5, Ciudad Universitaria, Universidad Autónoma de Coahuila, Torreón. CP 27000, México. * Corresponding author: E-mail: bnagamani@mail.uadec.mx Phone 871- 7571795, fax: (+52) 871- 7571785

 

Received 20^th February 2006
Accepted 6^th November 2007

 

Abstract

Biomethanation is a unique process which aids the recovery of carbon present in the wastes as methane, an energy rich product. The presence and activity of a variety of bacterial and archaeal microorganisms play a vital role in this process. An understanding of the microbial groups present in the anaerobic digester is important for augmenting the recovery of carbon and also will help in solving the digester related problems as the functioning of the digester depends on the microbial activity. Molecular characterization of the anaerobic bacteria present in a thermophilic (55°C) anaerobic digester fed with poultry waste was analyzed by DNA extraction followed by PCR amplification, cloning and sequencing of the obtained clones. Results showed that more than 75 per cent of the clones represented uncultured bacteria and among the different genera recorded, Clostridium sp. was dominant. Results demonstrated the need for obtaining more number of clones and a combination of different methods for reliable molecular characterization of an anaerobic digester.

Keywords: anaerobic digester, poultry waste, fermentative anaerobes, molecular analysis.

 

Resumen

La biometanación es un proceso único que ayuda la recuperación del carbono presente en la basura como el metano, un producto rico en energía. La presencia y actividad de una variedad bacteriana y los microorganismos del archaea juegan un papel vital en este proceso. Una comprensión de los grupos microbianos presentes en el digestor anaerobio es importante para aumentar la recuperación de carbono y también ayudará resolver los problemas relacionados con el digestor ya que depende de la actividad microbiana. La caracterización molecular de grupos de bacterias anaerobias presente en un digestor termofilico (55°C), anaerobio alimentado con los desechos de la pollería se analizó por extracto de ADN seguido por la amplificación de PCR, clonando y secuenciado de los clones obtenidos. Los resultados mostraron que más del 75 por ciento de los clones representaron las bacterias no cultivadas y entre diferente genero registrado, Clostridium sp. era dominante. Los resultados demostraron claramente la necesidad de obtener más número de clones y la combinación de los métodos diferentes para una caracterización molecular confiable de un digestor anaerobio.

Palabras clave: digestor anaerobio, desechos de pollería, anaerobias fermentativas, análisis molecular.

 

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Acknowledgement

The author wishes to thank Dr. David Huber, Dr. Umesh Reddy and Dr. Mark Chatfield of West Virginia State University, Institute, WV, USA for their help in providing lab facilities and all resources for this work.

 

References

Ahring, B.K. (2003). Perspectives for Anaerobic Digestion. Advances in Biochemical Engineering/Biotechnology 82, 1-29.         [ Links ]

Ali, M. K., Kimura, T., Sakka, K. and Ohmiya, K. (2001). The multidomain xylanase Xyn10B as a cellulose-binding protein in Clostridium stercorarium. FEMS Microbiology Letters 198, 79-83.         [ Links ]

Apajalahti, J., Kettunen, A. and Graham, H. (2004). Characteristics of the gastrointestinal microbial communities, with special reference to the chicken. World's Poultry Science Journal 60, 223-232.         [ Links ]

Burrell, P.C., O'Sullivan, C., Song, H., Clarke, W.P. and Blackall, L.L. (2004). Identification, detection, and spatial resolution of Clostridium populations responsible for cellulose degradation in a methanogenic landfill leachate bioreactor. Applied and Environmental Microbiology 70, 2414-2419        [ Links ]

Buzzini, A.P., Sakamoto, I.K., Varesche, M.B. and Pires, E.C. (2006). Evaluation of the microbial diversity in an UASB reactor treating wastewater from an unbleached pulp plant. Process Biochemistry 41, 168-176.         [ Links ]

Chouari, R., Le Paslier, D., Daegelen, P., Ginestet, P., Weissenbach, J. and Sghir, A. (2005). Novel predominant archaeal and bacterial groups revealed by molecular analysis of an anaerobic sludge digester. Environmental Microbiology 7, 1104-1115.         [ Links ]

Cole, J.R., Chai, B., Farris, R.J., Wang, Q., Kulam, S.A., McGarrell, D.M., Garrity, G.M. and Tiedje, J.M. (2005). The ribosomal database project (RDP-II): sequences and tools for high-throughput rRNA analysis. Nucleic Acids Research 33, D294-D296.         [ Links ]

Daims, H., Ramsing, N.B., Schleifer, K.H. and Wagner, M. (2001). Cultivation-independent, semiautomatic determination of absolute bacterial cell numbers in environmental samples by fluorescence in situ hybridization. Applied and Environmental Microbiology 67, 5810-5818.         [ Links ]

Dahllöf, I. (2002). Molecular community analysis of microbial diversity. Current Opinion in Biotechnology 13, 213-217.         [ Links ]

Drábková, L., Kirschner, J. and Vlcek, C. (2002). Comparison of seven DNA extraction and amplification protocols in historical herbarium specimens of Juncaceae. Plant Molecular Biology Reporter 20, 161-175.         [ Links ]

Espinosa-Solares, T., Bombardiere, J., Chatfield, M., Domaschko, M., Easter, M., Stafford, D.A., Castillo-Angeles, S. and Castellanos-Hernandez, N. (2006). Macroscopic mass and energy balance of a pilot plant anaerobic bioreactor operated under thermophilic conditions. Applied Biochemistry and Biotechnology 132, 959-968.         [ Links ]

Flora, J.R.V. and Riahi-Nezhad, C. (2006). Availability of poultry manure as a potential biofuel feedstock for energy production. p.23. South Carolina Energy Office, Columbia SC20201, USA.         [ Links ]

Godon, J.J., Zumstein, E., Dabert, P., Habouzit, F. and Moletta, R. (1997a). Molecular microbial diversity of an anaerobic digestor as determined by small-subunit rDNA sequence analysis. Applied and Environmental Microbiology 67, 2802-2813.         [ Links ]

Godon, J.J., Zumstein, E., Dabert, P., Habouzit, F. and Moletta, R. (1997b). Microbial 16S rDNA diversity in an anaerobic digester. Water Science and Technology 36, 40-55.         [ Links ]

Leung, K. and Topp, E. (2001). Bacterial community dynamics in liquid swine manure during storage: molecular analysis using DGGE/PCR of 16S rDNA. FEMS Microbiology Ecology 38, 169-177.         [ Links ]

Lynd, L. R., Weimer, P.J., van Zyl, W.H. and Pretorius, I.S. (2002). Microbial cellulose utilization: fundamentals and biotechnology. Microbiology and Molecular Biology Reviews 66, 506-577.         [ Links ]

McInerney, M.J. (1999). Anaerobic Metabolism and its Regulation. In: Biotechnology: a multi volume comprehensive treatise, (H.-J. Rehm, G. Reed, A. Pühler, and P.I.W. Stadler, eds.), Vol. 11a. Environmental Processes I -Wastewater Treatment, (J. Winter, ed.), pp. 455-478. Wiley-VCH Verlag GmbH, Weinheim, Germany.         [ Links ]

Mladenovska, Z., Dabrowski, S. and Ahring, B.K. (2003). Anaerobic digestion of manure and mixture of manure with lipids: biogas reactor performance and microbial community analysis. Water Science and Technology 48, 271-278        [ Links ]

O'Donnel, A.G. and Görres, H.E. (1999). 16S rDNA methods in soil microbiology. Current Opinion in Biotechnology 10, 225-229.         [ Links ]

O'Sullivan, C.A., Burrell, P.C., Clarke, W.P., Blackall, L.L. (2005). Structure of a cellulose degrading bacterial community during anaerobic digestion. Biotechnology and Bioengineering 92, 871-878.         [ Links ]

Schramm, A. and Amann, R. (1999). Nucleic acid-based techniques for analyzing the diversity, structure, and dynamics of microbial communities in wastewater treatment. In: Biotechnology: a multi volume comprehensive treatise, (H.-J. Rehm, G. Reed, A. Pühler, and P.I.W. Stadler, eds.), Vol. 11a. Environmental Processes I - Wastewater Treatment, (J. Winter, ed.), pp. 85-108. Wiley-VCH Verlag GmbH, Weinheim, Germany.         [ Links ]

Shiratori, H., Ikeno, H., Ayame, S., Kataoka, N, Miya, A., Hosono, K., Beppu, T., Ueda, K. (2006). Isolation and characterization of a new Clostridium sp. that performs effective cellulosic waste digestion in a thermophilic methanogenic bioreactor. Applied and Environmental Microbiology 72, 3702-3709.         [ Links ]

Smit, E., Leeflang, P., Gommans, S., van den Broek, J., van Mil, S. and Wernars, K. (2001). Diversity and seasonal fluctuations of the dominant members of the bacterial soil community in a wheat field as determined by cultivation and molecular methods. Applied and Environmental Microbiology 67, 2284-2291.         [ Links ]

Stewart, C. S., Flint, H. J. and Bryant, M.P. (1997). The rumen bacteria. In: The rumen microbial ecosystem, (P. N. Hobson and C. S. Stewart, eds.), p. 10-72. Blackie Academic and Professional, London, United Kingdom.         [ Links ]

Suau, A., Bonnet, R., Sutren, M., Godon, J.J., Gibson, G.R., Collins, M.D. and Dore, J. (1999). Direct analysis of genes encoding 16S rRNA from complex communities reveals many novel molecular species within the human gut. Applied and Environmental Microbiology 65, 4799-4807.         [ Links ]

Syutsubo, K., Nagaya, Y., Sakai, S. and Miya, A. (2005). Behavior of cellulose degrading bacteria in thermophilic anaerobic digestion process. Water Science and Technology 52, 79-84.         [ Links ]

Tang, Y. Shigematsu, T., Ikbal, M.S. and Kida, K. (2004). The effect of microaeration on the phylogenetic diversity of microorganisms in a thermophilic anaerobic municipal solid waste digester. Water Research 38, 2537-2550.         [ Links ]

Tauber, T., Berta, B., Székely, A.J., Gyarmati, I, Kékesi, K., Márialigeti, K. and Tóth, E.M. (2007). Characterisation of community structure of bacteria in parallel mesophilic and thermophilic pilot scale anaerobe sludge digesters. Acta Microbiologica et Immunologica Hungarita 54, 47-55.         [ Links ]

Van Dyke, M. I. and McCarthy, A.J. (2002). Molecular biological detection and characterization of Clostridium populations in municipal landfill sites. Applied and Environmental Microbiology 68, 2049-2053.         [ Links ]

Verstraete, W. and Vandevivere, P. (1998). New and broader applications of anaerobic digestion. Critical Reviews in Environmental Science and Technology 28, 151-173.         [ Links ]

Westlake, K., Archer, D.B. and Boone, D.R. (1995). Diversity of cellulolytic bacteria in landfill. Journal of Applied Bacteriology 79, 73-78.         [ Links ]