Growth of the syntrophic anaerobic acetogen, strain PA-1, with glucose or succinate as energy source
Applied and Environmental Microbiology
Strain PA-1 (S. Barik, W. J. Brulla, and M. P. Bryant, Appl. Environ. Microbiol. 50:304-310, 1985) is an anaerobic, gram-negative rod that in pure culture decarboxylates succinate to propionate and that grows syntrophically as an acetogen with the H2 utilizer Methanospirillum hungatei if glucose, pyruvate, aspartate, or fumarate is provided. In pure culture, strain PA-1 grows optimally in a medium containing 5% ruminal fluid, 0.1% yeast extract, a 4:1 N2-CO2 gas phase, and 20 mM succinate. With
... mM succinate. With the PA-1 plus M. hungatei coculture, good growth was obtained with 7.5 mM glucose and tryptophan could replace the yeast extract. Strain PA-1 in pure culture grew quite well in glucose medium if the large headspace was flushed intermittently with N2. Flushing with H2 inhibited this growth. We previously reported isolation from a methanogenic enrichment of sewage of strain PA-1, which was able to degrade phenylacetic acid (1). A coculture of an H2-utilizing fumarate reducer, Wolinella succinogenes, and strain PA-1 degraded a range of lowly substituted aromatic compounds such as phenol, benzoate, and phenylacetate, but it lost these abilities during long-term subculture. However, strain PA-1 in pure culture grew by decarboxylating succinate to propionate according to the following equation: -OOCCH2CH2COO-<-> CH3CH2COO-+ HC03-7 AGO' = -20.52 kJ (1) When PA-1 was cocultured with the H2-utilizing methanogen Methanospirillum hungatei (JF-1), good growth was obtained with pyruvate, aspartate, fumarate, or glucose as the substrate (1). On the basis of the products, the following stoichiometry was proposed for glucose: , AGO' = -341.9 kJ (4) Without an H2 utilizer, strain PA-1 grew very poorly. Syntrophococcus sucromutans is the only other anaerobic bacterium reported that cannot ferment sugars, unless it is cocultured with an H2 utilizer or with formate or methyl groups of methoxy benzenoids as electron acceptor systems (2). We present data showing that the inhibition of the growth of strain PA-1 in pure culture on glucose is due to a buildup of H2. Also, optimal substrate levels for growth of strain PA-1 plus M. hungatei on glucose and for growth of strain PA-1 in pure culture on succinate are presented. * Corresponding author. t Present address: Kech Laboratories 138-78, California Institute of Technology, Pasadena, CA 91125. Strain PA-1 and PA-1 plus M. hungatei were grown anaerobically (1) in RF medium (Table 1) . Triplicate serumstoppered tubes (18 by 150 mm), each with 10 ml of medium, were used for most experiments. The effect of H2 on the growth of strain PA-1 was determined in duplicate 150-ml sidearm (18-mm-diameter) flasks, each with 10 ml of medium. All incubations were at 35°C or 37°C. Culture purity was checked by phase-contrast microscopy. When grown in the RF medium with 20 mM succinate and various concentrations of yeast extract, strain PA-1 exhibited maximum growth with 0.05% yeast extract (A6. of 0.28 after 9 days of incubation). In the same medium but with 0.1% yeast extract and various concentrations of succinate, optimum growth (A6. of 0.31 after 16 days of incubation) was obtained with 20 mM succinate; succinate at 30 mM was somewhat inhibitory. The coculture of strain PA-1 plus M. hungatei grew optimally in the RF medium with 0.1% yeast extract and 7.5 mM glucose (A6. of 1.50 after 8 days of incubation). Tryptophan (5 mM) could replace the yeast extract, but the A600 was somewhat lower (1.2 after 9 days). The results showed that strain PA-1 is auxotrophic for tryptophan as M. hungatei does not have this requirement.