Changes in Cell Size and Shape during 50,000 Generations of Experimental Evolution with Escherichia coli

Abstract

Bacteria adopt a wide variety of sizes and shapes, with many species exhibiting stereotypical morphologies. How morphology changes, and over what timescales, is less clear. Previous work examining cell morphology in an experiment with Escherichia coli showed that populations evolved larger cells and, in some cases, cells that were less rod-like. That experiment has now run for over two more decades. Meanwhile, genome sequence data are available for these populations, and new computational methods enable high-throughput microscopic analyses. Here, we measured stationary-phase cell volumes for the ancestor and 12 populations at 2,000, 10,000, and 50,000 generations, including measurements during exponential growth at the last timepoint. We measured the distribution of cell volumes for each sample using a Coulter counter and microscopy, the latter of which also provided data on cell shape. Our data confirm the trend toward larger cells, while also revealing substantial variation in size and shape across replicate populations. Most populations first evolved wider cells, but later reverted to the ancestral length-to-width ratio. All but one population evolved mutations in rod-shape maintenance genes. We also observed many ghost-like cells in the only population that evolved the novel ability to grow on citrate, supporting the hypothesis that this lineage struggles with maintaining balanced growth. Lastly, we show that cell size and fitness remain correlated across 50,000 generations. Our results suggest larger cells are beneficial in the experimental environment, while the reversion toward ancestral length-to-width ratios suggests partial compensation for the less favorable surface area-to-volume ratios of the evolved cells. IMPORTANCE Bacteria exhibit great morphological diversity, yet we have only a limited understanding of how their cell sizes and shapes evolve, and of how these features affect organismal fitness. This knowledge gap reflects, in part, the paucity of the fossil record for bacteria. Here, we revive and analyze samples extending over 50,000 generations from 12 populations of experimentally evolving Escherichia coli to investigate the relation between cell size, shape, and fitness. Using this “frozen fossil record” we show that all 12 populations evolved larger cells concomitant with increased fitness, with substantial heterogeneity in cell size and shape across the replicate lines. Our work demonstrates that cell morphology can readily evolve and diversify, even among populations living in identical environments.

Primordial mimicry induces morphological change in Escherichia coli

“…The changes in cell shape toward the spherical form rather than other morphological features were highly crucial for the E. coli to use OAVs as the sole carbon source. So far, whether and how OAVs impacted cell morphology through metabolism mediated by molecular machinery is unclear, as both the short-term evolution in OAVs and the long-term evolution in glucose 47 , 48 cause changes in cell size. Alternatively, the changes in cell shape might be partially attributed to the changes in the plasma membrane capacity of E. coli caused by the carbon source changing from glucose to OAVs, because the changes in fatty acid synthesis due to such a nutritional alteration could influence the cell envelope capacity, thereby affecting cell size and morphology 49 .…”

Evolution with private resources reverses some changes from long-term evolution with public resources

“…Both cell size and total biomass increased in all 12 LTEE populations during selection for increased competitive ability for shared resources despite the fact that neither is directly a target of selection under those conditions (Grant et al 2021, Lenski & Mongold, 2000). This increase in cell size can be seen in our data for the three time points that we sampled from the Ara–5 population, by comparing the values of the three grey points along the x-axis in Figure 5.…”

“…All of the LTEE lineages show increases in competitive ability relative to their common ancestor, while simultaneously having lower numerical yields (Vasi et al 1994, Wiser et al 2013). The reductions in numerical yield result from the evolution of much larger individual cells (Lenski & Travisano 1994, Vasi et al 1994, Grant et al 2021). In many bacteria, increased cell size is a physiological response to increased growth rate (Johnston et al 1979; Pierucci, 1978).…”

Resource allocation to cell envelopes and the scaling of bacterial growth rate

“…First, we used S/V in combination with the relative fitness data for four LTEE time points spanning 50,000 generations [59]. Given that the relative fitness is the ratio of Malthusian parameters of evolved to ancestral line determined from the head-to-head competition (see [60]), we converted this metric to the growth rate by multiplying it with the growth rate of the ancestral strain (see V max in [61]).…”