Methane fermentation is a useful process for liquid wastewater treatment because it reduces the chemical oxygen demand before disposal and yields the valuable fuel methane. Although most of the problems concerning the hardware technology for full-scale application of this fermentation process have been overcome, understanding of the microbial basis of methanogenie fermentation of wastes should facilitate process control and improve reactor performance. Since methanogenic bacteria contain a number of unique cofactors, it might be possible to use these compounds as specific parameters for the identification and quantitation of methanogens in anaerobic sludge. The presence ofcoenzyme F4zo allows the identification ofmethanogenic bacteria in mixed populations by epifluorescenee microscopy; in this way information is obtained concerning the location and interactions of methanogens. Chromatographic analysis of coenzyme F4z0 derivatives, which differ in the number of glutamate groups present in the molecule, yields information on the presence of Methanosarcina species among the methanogens. Since an aberrant corrinoid and a specific group of 7-methylpterins, among which methanopterin, are present in methanogens, the analysis of such compounds is now successfully used in studies on the activity ofmethanogens in various anaerobic reactors.Episymbiotic association ofmethanogenic bacteria and rumen ciliates H. J. GIJZEN, K. B. ZWART and C. K. STUMM Department of Microbiology, Faculty of Science, University of Nijmegen, Toernooiveld, 6525 ED Nijmegen, The NetherlandsOne of the many interrelationships that exists between tureen microorganisms is the association of methanogenic bacteria with rumen ciliates (Vogels et aL, 1980; Stumm et al., 1982). This episymbiotic association enables an efficient interspecies hydrogen transfer.We have studied changes in the association ofmethanogens with the ciliates Eudiplodinium maggii and Diplodinium dentatum under different conditions of feeding and at different partial hydrogen tensions. Methanogenic bacteria were identified by bright-field fluorescence microscopy according to Doddema and Vogels (1978). A relatively low association frequency (20-30~£) was observed when substrates containing easily degradable compounds (grass, concentrates) were given and also during flushing with hydrogen. A high association frequency (70-100~) was observed after feeding cellulose and at lo w hydrogen tensions caused by flushing with nitrogen. Apart from the frequency of association the number ofmethanogens attached to the ciliate surface varied. After feeding grass the number of associated methanogens was low, whereas after feeding cellulose the ciliate surface was almost completely covered with long chains of methanogenic bacteria. Chains of these organisms were

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