Background: In the process of developing a microplate-based growth assay, we discovered that our test organism, a native E. coli isolate, displayed very uniform doubling times (tau) only up to a certain threshold cell density. Below this cell concentration (<or= 100 -1,000 CFU mL-1 ; <or= 27-270 CFU well-1) we observed an obvious increase in the tau scatter.
Results: Working with a food-borne E. coli isolate we found that tau values derived from two different microtiter platereader-based techniques (i.e., optical density with growth time {=OD[t]} fit to the sigmoidal Boltzmann equation or time to calculated 1/2-maximal OD {=tm} as a function of initial cell density {=tm[CI]}) were in excellent agreement with the same parameter acquired from total aerobic plate counting. Thus, using either Luria-Bertani (LB) or defined (MM) media at 37 degrees C, tau ranged between 17-18 (LB) or 51-54 (MM) min. Making use of such OD[t] data we collected many observations of tau as a function of manifold initial or starting cell concentrations (CI). We noticed that tau appeared to be distributed in two populations (bimodal) at low CI. When CI <or=100 CFU mL-1 (stationary phase cells in LB), we found that about 48% of the observed tau values were normally distributed around a mean (mutau1) of 18 +/- 0.68 min (+/- sigmatau1) and 52% with mutau2 = 20 +/- 2.5 min (n = 479). However, at higher starting cell densities (CI>100 CFU mL-1), the tau values were distributed unimodally (mutau = 18 +/- 0.71 min; n = 174). Inclusion of a small amount of ethyl acetate to the LB caused a collapse of the bimodal to a unimodal form. Comparable bimodal tau distribution results were also observed using E. coli cells diluted from mid-log phase cultures. Similar results were also obtained when using either an E. coli O157:H7 or a Citrobacter strain. When sterile-filtered LB supernatants, which formerly contained relatively low concentrations of bacteria(1,000-10,000 CFU mL-1), were employed as a diluent, there was an evident shift of the two populations towards each other but the bimodal effect was still apparent using either stationary or log phase cells.
Conclusion: These data argue that there is a dependence of growth rate on starting cell density.