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http://dx.doi.org/10.5713/ajas.2013.13134

Production of Citrate by Anaerobic Fungi in the Presence of Co-culture Methanogens as Revealed by 1H NMR Spectrometry  

Cheng, Yan Fen (Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University)
Jin, Wei (Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University)
Mao, Sheng Yong (Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University)
Zhu, Wei-Yun (Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University)
Publication Information
Asian-Australasian Journal of Animal Sciences / v.26, no.10, 2013 , pp. 1416-1423 More about this Journal
Abstract
The metabolomic profile of the anaerobic fungus Piromyces sp. F1, isolated from the rumen of goats, and how this is affected by the presence of naturally associated methanogens, was analyzed by nuclear magnetic resonance spectroscopy. The major metabolites in the fungal monoculture were formate, lactate, ethanol, acetate, succinate, sugars/amino acids and ${\alpha}$-ketoglutarate, whereas the co-cultures of anaerobic fungi and associated methanogens produced citrate. This is the first report of citrate as a major metabolite of anaerobic fungi. Univariate analysis showed that the mean values of formate, lactate, ethanol, citrate, succinate and acetate in co-cultures were significantly higher than those in the fungal monoculture, while the mean values of glucose and ${\alpha}$-ketoglutarate were significantly reduced in co-cultures. Unsupervised principal components analysis revealed separation of metabolite profiles of the fungal mono-culture and co-cultures. In conclusion, the novel finding of citrate as one of the major metabolites of anaerobic fungi associated with methanogens may suggest a new yet to be identified pathway exists in co-culture. Anaerobic fungal metabolism was shifted by associated methanogens, indicating that anaerobic fungi are important providers of substrates for methanogens in the rumen and thus play a key role in ruminal methanogenesis.
Keywords
Piromyces sp. F1; Methanobrevibacter thaueri; Nuclear Magnetic Resonance; Citrate;
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1 Mountfort, D. O., R. A. Asher, and T. Bauchop. 1982. Fermentation of cellulose to methane and carbon dioxide by a rumen anaerobic fungus in a triculture with Methanobrevibacter sp. Strain RA1 and Methanosarcina barkeri. Appl. Environ. Microbiol. 44:128-134.
2 Nakashimada, Y., K. Srinivasan, M. Murakami, and N. Nishio. 2000. Direct conversion of cellulose to methane by anaerobic fungus Neocallimastix frontalis and defined methanogens. Biotechnol. Lett. 22:223-227.   DOI   ScienceOn
3 Orpin, C. G. 1975. Studies on the rumen flagellate Neocallimastix frontalis. J. Gen. Microbiol. 91:249-262.   DOI   ScienceOn
4 O'Fallon, J. V., R. W. Wright, and R. E. Calza. 1991. Glucose metabolic pathways in the anaerobic rumen fungus Neocallimastix frontalis EB188. J. Biochem. 274:595-599.
5 Pham, L. H., J. Vater, W. Rotard, and C. Mugge. 2005. Identification of secondary metabolites from Streptomyces violaceoruber TU22 by means of on-flow LC-NMR and LC-DAD-MS. Magn. Reson. Chem. 43:710-723.   DOI   ScienceOn
6 Teunissen, M. J., E. P. W. Kets, H. J. M. Op den Camp, J. H. J. Huis in't Veld, and G. D. Vogels. 1992. Effect of coculture of anaerobic fungi isolated from ruminants and non-ruminants with methanogenic bacteria on cellulolytic and xylanolytic enzyme activities. Arch. Microbiol. 157:176-182.
7 Theodorou, M. K., D. R. Davies, and C. G. Orpin. 1995. Nutrition and survival of anaerobic fungi. In: Anaerobic fungi: Biology, Ecology and Function (Ed. D. O. Mountfort and C. G. Oprin). Marcel Dekker, New York, USA. pp. 107-128.
8 Theodorou, M. K., G. Mennim, D. R. Davies, W. Y. Zhu, A. P. Trinci, and J. L. Brookman. 1996. Anaerobic fungi in the digestive tract of mammalian herbivores and their potential for exploitation. Proc. Nutr. Soc. 55:913-926.   DOI   ScienceOn
9 Chikayama, E., M. Suto, T. Nishihara, K. Shinozaki, T. Hirayama, and J. Kikuchi. 2008. Systematic NMR analysis of stable isotope labeled metabolite mixtures in plant and animal systems: coarse grained views of metabolic pathways. PLoS ONE 3: e3805.   DOI   ScienceOn
10 Grivet, J. P. 2001. NMR and microorganisms. Curr. Issues Mol. Biol. 3:7-14.
11 Grivet, J. P. and A. M. Delort. 2009. NMR for microbiology: In vivo and in situ applications. Prog. Nucl. Magn. Reson. Spectrosc. 54:1-53.   DOI   ScienceOn
12 Kwon, M., J. Song, J. K. Ha, H. S. Park, and J. Chang. 2009. Analysis of functional genes in carbohydrate metabolic pathway of anaerobic rumen fungus Neocallimastix frontalis PMA02. Asian-Aust. J. Anim. Sci. 22:1555-1565.   과학기술학회마을   DOI
13 Hungate, R. E. 1982. Methane formation and cellulose digestion biochemical ecology and microbiology of the rumen ecosystem. Experimenta 38:189-192.   DOI   ScienceOn
14 Jin, W., Y. F. Cheng, S. Y. Mao, and W. Y. Zhu. 2011. Isolation of natural cultures of anaerobic fungi and indigenously associated methanogens from herbivores and their bioconversion of lignocellulosic materials to methane. Bioresour. Technol. 102:7925-7931.   DOI   ScienceOn
15 Joblin, K. N., G. E. Naylor, and A. G. Williams. 1990. Effect of Methanobrevibacter smithii on xylanolytic activity of anaerobic ruminal fungi. Appl. Environ. Microbiol. 56:2287-2295.
16 Liu J. H. 2009. Metabolic profiles of natural co-cultures of anaerobic fungi and methanogens in vitro and the production of lactate. Ma.D. Thesis, Nanjing Agricultural University, Nanjing, China.
17 Marvin-Sikkema, F. D., A. J. Richardson, C. S. Steward, J. C. Gottschal, and R. A. Prins. 1990. Influence of hydrogen-consuming bacteria on cellulose degradation by anaerobic fungi. Appl. Environ. Microbiol. 56:3793-3797.
18 Akhmanova, A., F. G. J. Voncken, K. M. Hosea, H. Harhangi, J. T. Keltjens, H. J. M. op den Camp, G. D. Vogels, and J. H. P. Hackstein. 1999. A hydrogenosome with pyruvate formate-lyase: anaerobic chytrid fungi use an alternative route for pyruvate catabolism. Mol. Microbiol. 32:1103-1114.   DOI   ScienceOn
19 Balch, W. E., G. E. Fox, L. J. Magrum, C. R. Woese, and R. S. Wolfe. 1979. Methanogens: reevaluation of a unique biological group. Microbiol. Rev. 43:260-296.
20 Bauchop, T. and D. O. Mountfort. 1981. Cellulose fermentation by a rumen anaerobic fungus in both the absence and the presence of rumen methanogens. Appl. Environ. Microbiol. 42:1103-1110.
21 Beckonert, O., H. C. Keun, T. M. Ebbels, J. Bundy, E. Holmes, J. C. Lindon, and J. K. Nicholson. 2007. Metabolic profiling, metabolomic and metabonomic procedures for NMR spectroscopy of urine, plasma, serum and tissue extracts. Nat. Protoc. 2:2692-2703.   DOI   ScienceOn
22 Boxma, B., F. Voncken, S. Jannink, T. van Alen, A. Akhmanova, S. W. van Weelden, J. J. van Hellemond, G. Ricard, M. Huynen, A. G. Tielens, and J. H. Hackstein. 2004. The anaerobic chytridiomycete fungus Piromyces sp. E2 produces ethanol via pyruvate: formate lyase and an alcohol dehydrogenase E. Mol. Microbiol. 51:1389-1399.   DOI   ScienceOn
23 Carrieri, D., K. McNeely, A. C. De Roo, N. Nennette, I. Pelczer, and G. C. Dismukes. 2009. Identification and quantification of water-soluble metabolites by cryoprobe-assisted nuclear magnetic resonance spectroscopy applied to microbial fermentation. Magn. Reson. Chem. 47:S138-S146.   DOI   ScienceOn
24 Cheng, Y. F., J. E. Edwards, G. G. Allison, W. Y. Zhu, and M. K. Theodorou. 2009. Diversity and activity of enriched ruminal cultures of anaerobic fungi and methanogens grown together on lignocellulose in consecutive batch culture. Bioresour. Technol. 100:4821-4828.   DOI   ScienceOn
25 Cheng, Y. F., S. Y. Mao, C. X. Pei, J. X. Liu, and W. Y. Zhu. 2006. Detection and diversity analysis of rumen methanogens in co-cultures with anaerobic fungi. Acta Microbiologica Sinica 46: 879-883.