Metabolic Commensalism and Competition in a Two-Species Microbial Consortium
Authors: Christensen Bjarke B. , Haagensen Janus A. J. , Heydorn Arne , Molin Søren 
Published: July 2002

We analyzed metabolic interactions and the importance of specific structural relationships in a benzyl alcohol-degrading microbial consortium comprising two species,
Pseudomonas putida
strain R1 and
Acinetobacter
strain C6, both of which are able to utilize benzyl alcohol as their sole carbon and energy source. The organisms were grown either as surface-attached organisms (biofilms) in flow chambers or as suspended cultures in chemostats. The numbers of CFU of
P. putida
R1 and
Acinetobacter
strain C6 were determined in chemostats and from the effluents of the flow chambers. When the two species were grown together in chemostats with limiting concentrations of benzyl alcohol,
Acinetobacter
strain C6 outnumbered
P. putida
R1 (500:1), whereas under similar growth conditions in biofilms,
P. putida
R1 was present in higher numbers than
Acinetobacter
strain C6 (5:1). In order to explain this difference, investigations of microbial activities and structural relationships were carried out in the biofilms. Insertion into
P. putida
R1 of a fusion between the growth rate-regulated rRNA promoter
rrnB
P1 and a
gfp
gene encoding an unstable variant of the green fluorescent protein made it possible to monitor the physiological activity of
P. putida
R1 cells at different positions in the biofilms. Combining this with fluorescent in situ hybridization and scanning confocal laser microscopy showed that the two organisms compete or display commensal interactions depending on their relative physical positioning in the biofilm. In the initial phase of biofilm development, the growth activity of
P. putida
R1 was shown to be higher near microcolonies of
Acinetobacter
strain C6. High-pressure liquid chromatography analysis showed that in the effluent of the
Acinetobacter
strain C6 monoculture biofilm the metabolic intermediate benzoate accumulated, whereas in the biculture biofilms this was not the case, suggesting that in these biofilms the excess benzoate produced by
Acinetobacter
strain C6 leaks into the surrounding environment, from where it is metabolized by
P. putida
R1. After a few days,
Acinetobacter
strain C6 colonies were overgrown by
P. putida
R1 cells and new structures developed, in which microcolonies of
Acinetobacter
strain C6 cells were established in the upper layer of the biofilm. In this way the two organisms developed structural relationships allowing
Acinetobacter
strain C6 to be close to the bulk liquid with high concentrations of benzyl alcohol and allowing
P. putida
R1 to benefit from the benzoate leaking from
Acinetobacter
strain C6. We conclude that in chemostats, where the organisms cannot establish in fixed positions, the two strains will compete for the primary carbon source, benzyl alcohol, which apparently gives
Acinetobacter
strain C6 a growth advantage, probably because it converts benzyl alcohol to benzoate with a higher yield per time unit than
P. putida
R1. In biofilms, however, the organisms establish structured, surface-attached consortia, in which heterogeneous ecological niches develop, and under these conditions competition for the primary carbon source is not the only determinant of biomass and population structure.