42 research outputs found
Evidence for a bimodal distribution of Escherichia coli doubling times below a threshold initial cell concentration
Abstract 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 (τ) only up to a certain threshold cell density. Below this cell concentration (≤ 100 -1,000 CFU mL-1 ; ≤ 27-270 CFU well-1) we observed an obvious increase in the τ scatter. Results Working with a food-borne E. coli isolate we found that τ 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°C, τ ranged between 17-18 (LB) or 51-54 (MM) min. Making use of such OD[t] data we collected many observations of τ as a function of manifold initial or starting cell concentrations (CI). We noticed that τ appeared to be distributed in two populations (bimodal) at low CI. When CI ≤100 CFU mL-1 (stationary phase cells in LB), we found that about 48% of the observed τ values were normally distributed around a mean (μτ1) of 18 ± 0.68 min (± στ1) and 52% with μτ2 = 20 ± 2.5 min (n = 479). However, at higher starting cell densities (CI>100 CFU mL-1), the τ values were distributed unimodally (μτ = 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 τ 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.</p
Reliability of Transcriptional Cycles and the Yeast Cell-Cycle Oscillator
A recently published transcriptional oscillator associated with the yeast cell cycle provides clues and raises questions about the mechanisms underlying autonomous cyclic processes in cells. Unlike other biological and synthetic oscillatory networks in the literature, this one does not seem to rely on a constitutive signal or positive auto-regulation, but rather to operate through stable transmission of a pulse on a slow positive feedback loop that determines its period. We construct a continuous-time Boolean model of this network, which permits the modeling of noise through small fluctuations in the timing of events, and show that it can sustain stable oscillations. Analysis of simpler network models shows how a few building blocks can be arranged to provide stability against fluctuations. Our findings suggest that the transcriptional oscillator in yeast belongs to a new class of biological oscillators
An Environment-Sensitive Synthetic Microbial Ecosystem
Microbial ecosystems have been widely used in industrial production, but the inter-relationships of organisms within them haven't been completely clarified due to complex composition and structure of natural microbial ecosystems. So it is challenging for ecologists to get deep insights on how ecosystems function and interplay with surrounding environments. But the recent progresses in synthetic biology show that construction of artificial ecosystems where relationships of species are comparatively clear could help us further uncover the meadow of those tiny societies. By using two quorum-sensing signal transduction circuits, this research designed, simulated and constructed a synthetic ecosystem where various population dynamics formed by changing environmental factors. Coherent experimental data and mathematical simulation in our study show that different antibiotics levels and initial cell densities can result in correlated population dynamics such as extinction, obligatory mutualism, facultative mutualism and commensalism. This synthetic ecosystem provides valuable information for addressing questions in ecology and may act as a chassis for construction of more complex microbial ecosystems
Culturing Aerobic and Anaerobic Bacteria and Mammalian Cells with a Microfluidic Differential Oxygenator
In this manuscript, we report on the culture of anaerobic and aerobic species within a disposable multilayer polydimethylsiloxane (PDMS) microfluidic device with an integrated differential oxygenator. A gas-filled microchannel network functioning as an oxygen−nitrogen mixer generates differential oxygen concentration. By controlling the relative flow rate of the oxygen and nitrogen input gases, the dissolved oxygen (DO) concentration in proximal microchannels filled with culture media are precisely regulated by molecular diffusion. Sensors consisting of an oxygen-sensitive dye embedded in the fluid channels permit dynamic fluorescence-based monitoring of the DO concentration using low-cost light-emitting diodes. To demonstrate the general utility of the platform for both aerobic and anaerobic culture, three bacteria with differential oxygen requirements (E. coli, A. viscosus, and F. nucleatum), as well as a model mammalian cell line (murine embryonic fibroblast cells (3T3)), were cultured. Growth characteristics of the selected species were analyzed as a function of eight discrete DO concentrations, ranging from 0 ppm (anaerobic) to 42 ppm (fully saturated)
Noise Reduction by Diffusional Dissipation in a Minimal Quorum Sensing Motif
Cellular interactions are subject to random fluctuations (noise) in quantities of interacting molecules. Noise presents a major challenge for the robust function of natural and engineered cellular networks. Past studies have analyzed how noise is regulated at the intracellular level. Cell–cell communication, however, may provide a complementary strategy to achieve robust gene expression by enabling the coupling of a cell with its environment and other cells. To gain insight into this issue, we have examined noise regulation by quorum sensing (QS), a mechanism by which many bacteria communicate through production and sensing of small diffusible signals. Using a stochastic model, we analyze a minimal QS motif in Gram-negative bacteria. Our analysis shows that diffusion of the QS signal, together with fast turnover of its transcriptional regulator, attenuates low-frequency components of extrinsic noise. We term this unique mechanism “diffusional dissipation” to emphasize the importance of fast signal turnover (or dissipation) by diffusion. We further show that this noise attenuation is a property of a more generic regulatory motif, of which QS is an implementation. Our results suggest that, in a QS system, an unstable transcriptional regulator may be favored for regulating expression of costly proteins that generate public goods
Cooperative Adaptation to Establishment of a Synthetic Bacterial Mutualism
To understand how two organisms that have not previously been in contact can establish mutualism, it is first necessary to examine temporal changes in their phenotypes during the establishment of mutualism. Instead of tracing back the history of known, well-established, natural mutualisms, we experimentally simulated the development of mutualism using two genetically-engineered auxotrophic strains of Escherichia coli, which mimic two organisms that have never met before but later establish mutualism. In the development of this synthetic mutualism, one strain, approximately 10 hours after meeting the partner strain, started oversupplying a metabolite essential for the partner's growth, eventually leading to the successive growth of both strains. This cooperative phenotype adaptively appeared only after encountering the partner strain but before the growth of the strain itself. By transcriptome analysis, we found that the cooperative phenotype of the strain was not accompanied by the local activation of the biosynthesis and transport of the oversupplied metabolite but rather by the global activation of anabolic metabolism. This study demonstrates that an organism has the potential to adapt its phenotype after the first encounter with another organism to establish mutualism before its extinction. As diverse organisms inevitably encounter each other in nature, this potential would play an important role in the establishment of a nascent mutualism in nature
Synthetic biology: Understanding biological design from synthetic circuits
An important aim of synthetic biology is to uncover the design principles of natural biological systems through the rational design of gene and protein circuits. Here, we highlight how the process of engineering biological systems — from synthetic promoters to the control of cell–cell interactions — has contributed to our understanding of how endogenous systems are put together and function. Synthetic biological devices allow us to grasp intuitively the ranges of behaviour generated by simple biological circuits, such as linear cascades and interlocking feedback loops, as well as to exert control over natural processes, such as gene expression and population dynamics