Abstract
For anaerobic mixed cultures performing microbial chain elongation, it is unclear how pH alterations affect the abundance of key players, microbial interactions, and community functioning in terms of medium-chain carboxylate yields. We explored pH effects on mixed cultures enriched in continuous anaerobic bioreactors representing closed model ecosystems. Gradual pH increase from 5.5 to 6.5 induced dramatic shifts in community composition, whereas product range and yields returned to previous states after transient fluctuations. To understand community responses to pH perturbations over long-term reactor operation, we applied Aitchison PCA clustering, linear mixed-effects models, and random forest classification on 16S rRNA gene amplicon sequencing and process data. Different pH preferences of two key chain elongation species─one Clostridium IV species related to Ruminococcaceae bacterium CPB6 and one Clostridium sensu stricto species related to Clostridium luticellarii─were determined. Network analysis revealed positive correlations of Clostridium IV with lactic acid bacteria, which switched from Olsenella to Lactobacillus along the pH increase, illustrating the plasticity of the food web in chain elongation communities. Despite long-term cultivation in closed systems over the pH shift experiment, the communities retained functional redundancy in fermentation pathways, reflected by the emergence of rare species and concomitant recovery of chain elongation functions.