Fig. 4: Cross-feeding in the stable cycle of the coculture in weak buffer.

Measurements of various quantities of the coculture throughout the fifth 24-h growth-dilution cycle. a The viable counts of 1A01 and 3B05 cells. The value shown was the average of three measurements on the same sample from a single co-culture; the error between these measurements was less than the size of the data marker. b The concentrations of GlcNAc and acetate in the medium from the same co-culture measured in (a). The measurement variability for the determination of sugar, organic acid, or amino acid concentrations by HPLC was ~2% on the basis of repeated measurements of the same spent media sample. The light gray regions in (a) and (b) indicate the period when 3B05 continued to grow after GlcNAc depletion. In (c)–(f), the duration around the “acetate peak” was densely sampled using a protocol that mimicked the stable cycle; see “Methods”. The data from all four panels were measured on the same coculture. The pH measurement was accurate to ±0.02 pH unit on the basis of repeated measurements of pH standards. The dotted vertical line at time “0” indicates the time of GlcNAc depletion, around 12 h into the cycle (b). Gray-shaded regions are the same as those in (a) and (b). The same symbols are used in (c) and (d) as in (a) and (b). In (c) the dashed-dotted line indicates exponential growth of 3B05 at rate ~0.35/h before the acetate peak; the solid line indicates a growth rate ~0.55/h after the acetate peak. The filled orange circles in (d) indicate the culture pH (right vertical axis). e Concentrations of pyruvate, lactate, and glutamate in the medium as measured by HPLC. f Scaled intensities of metabolites in the medium as measured by untargeted metabolomics; see “Methods”. Metabolites consumed (defined as those with the scaled intensity of the last timepoint <0.5) are plotted in purple. Other detected metabolites are plotted in gray. Identities of the metabolites are given in Supplementary Table 2 and their values are provided in the Source Data file.