Microbial physiology and genetics - Essay Example

Microbial physiology and genetics Experiment MPG2: Continuous culture. Abstract. For one to model and follow the microbial evolution inside a chemostat, different parameters are required to give identification of the direct evolution extent given a high time of resolution. For this experiment, the maximum specialized growth rate evolution (μmax) together with the concentration of the residual glucose was observed for Escherichia coli populations under limited glucose conditions and at specific rates of dilution in the continuous culture. μmax was seen to improve in the initial hours, whereas the concentration of the residual glucose reduced at a steady rate in the 500h of cultivation. This worked as parameters that are convenient for monitoring the population evolution in at a high-resolution time in regards to its affinity for the limiting substrate growth. Aim. The prime aim of this experiment was to show out the equilibrium dynamics of a continuous culture. In this case, it is expected that, by the end of the practice session, a learner would be should be able to fully describe the washout kinetics in a continuous culture. Additionally, should be able to describe the growth kinetics in a continuous culture. The concept behind the calculation shall be mastered through the participation of the student, in which case, he or she should calculate the washout data of μ_max. Introduction. Every living microorganism needs nutrients for reproduction, growth, and maintenance (Kreyazig, 2007). The role of nutrients is to provide energy, as well as raw materials that would be used in building new components of the cell for their replication (Kreyazig, 2007). For replication, the cell needs to carry out a number of chemical synthesis and reaction for particular cellular structures. This explains why in the 1960s the reproducibility and versatility of the continuous culture were used in addressing basic problems in the microbial field that are diverse (Maaloe, 2004). To date, the utility of the technique of continuous culture helps in acquiring reliable data in biologically data seta that are homogenous. It is worth noting that a continuous culture gives out many advantages compared to the biologically batch culture that is heterogeneous. This occurs in situations where the secondary growth and effects of stress provide subtle mask trends and differences (Mahler, 2002). In order to understand the characteristics of the continuous culture, an experiment was set to investigate the equilibrium dynamics of a continuous culture. Method. The participants for this experiment were divided into groups having five individuals. The subjects were then given a mock continuous culture and an Escherichia coli continuous culture. A chemostat that was small was assembled out of a Schott flask having a volume of about 75 ml. The standard magnetic Stirrer was used to stir the chemostat. Air was introduced into the culture by use of headspace. In this case, the pH was maintained at pH 7 with potassium peroxide and sodium peroxide (Kessler, 2004). After this, the chemostats were inoculated with a given number of cells. This medium had glucose flow of about 1mg. Large chemostats medium was started at the same time. The number of cells inside the small chemostat was analyzed by the counts of the plate. Additionally, the total number of cells in large chemostats was analyzed by the counts of the plate. The crystal washout was observed from the system of the mock continuous culture. The data was presented in a suitable graphical form. Each of the group needed to discuss the data and establish a conclusion concerning the kinetic vessel washout. The biomass was monitored in the E. coli continuous instructed fermenter. The participants’ group compared the generated data from this culture from that of the system of mock and reach a conclusion concerning the growth kinetic description within the continuous culture. Additionally, the rate of dilution of the continuous culture was increased, and the biomass monitored in accordance with the given instructions. The collected data was plotted, and μmax calculated. The absorbance at 500nm was measured over a given period of time. The data was plotted as abs against time. The data collected was recorded in table 1. Results. Table 1: Absorption at 500nm Time (min) Abs’(500nm) Abs’(500nm) 0 -0.23 0.502 2 -0.69 0.202 4 -0.99 0.102 6 -1.41 0.039 8 -1.89 0.013 10 -2.52 0.003 At 660nm, abs are 2/3. If abs are greater than 0.8, dilute it in the abs then multiply it by a dilution factor. The first abs are 0.925. It was diluted with nutrient broth to give 0.486 having das 0.34h-1. Table 2: Absorption at 660nm. Time (min) Abs’(660nm) 0 0.966 14 0.964 38 0.942 54 0.972 This continuous culture is a steady state condition. Table 4: Chemostat culture at D 2.0h-1 Time (min) Abs’(660nm) 0 0.763 8 0.604 10 0.546 15 0.522 Data analysis. Steady state. At steady state, the change in biomass will be given by dx/DT = x-Dx. However in this state, the concentration of the biomass is a constant, thus dx/dt is equal to zero and x-dx=0. This implies that u will be equal to D. =max S/KS + S . S represents the substrate residual concentration where as Ks is the saturation constant half rate. From the experiment the volume left in the tube was 314 ml and after 30 seconds the volume of water was 72ml. The volume will be 314/1000= 0.314 dm3 72ml/1000 = 0.072dm3 Flow rate would be given as 0.072/8.33 x 10-3 = 8.642dm3h-1 But v is equal to 0.314dm3 D=8.643/0.314 = 27.53n-1 At steady state, D = = 27.53n-1 max= 27.53n-1 (0.386)/0.072= 1.476 x 102 Chemostat culture at D 2.0h-1 Discussion. This experiment gave out data for a process of evolution that can be described by the model of deterministic in populations that are large. This data shifts to be stochastic whenever the size of the population is in a range of about 1/(mutation rate) or minimum. It has also been displayed that an evolution that is selectively driven can move on as a component of the absolute time taken rather than a component of the generation numbers. This experiment gives a demonstration of the special case of chemostat evolution in a limitation of glucose. However, this does not act as a molecular clock seen in different substitution rates of amino acids. This behavior is evident from the plotted graphs as the improvement of K is seen. In the 0.3 and 0,1 dilution rates, the K evolution appears to be similar with an exception of the initial 150h. This experiment gives a prediction of equal improvements for K in different mutations (Kessler, 2004). A number of models have been highlighted for describing the kinetic growth of populations of a microbial that grow with a unit substrate. Different studies have shown that in different models that have tested the Shehata & marry (2001), Monod (2000) and Westerhoff (2003) identified their data in a recommendable way. These models give out an appropriate description of getting glucose concentration steady states in E. coli cultures that are cultivated by chemostats with limited glucose in various temperatures and rates of dilution. In these models, the concentration of the residual glucose is calculated showing out that the E. coli populations improve their affinity for glucose in the process of growth under the conditions of limited glucose. Using these models, the periodic take-over can be modeled by populations that are mutant having glucose that has an improved affinity (Beek, 2008). The periodic take-over obtained result into a decrease that is steady in the residual concentration of glucose with a continuous culture. References. Beek, C., 2008. The pattern of enzymes and Aerobic growth. New York: Oxford publishers. Kessler, P., 2004. Escherichia coli growth. New York: Jack and sons publishers. Kreyazig, E., 2007. Continous Culture. New York: John Willy & sons press. Maaloe, O., 2004. Microbial synthesis. New York: Benjamin. Inc. Mahler, R., 2002. Biological chemistry. New York: Harper & Row Press. Monod, J., 2000. Research on bacteira cultures. Paris : Hermann press. Shehata, T., & Marr, G., 2001. Effects of nutrient concentration on the growth of Escherichia coli. Journal of Bacteriol 107. Westerhoff, H., 2003. Thermodynamic of growth, non-equilibrium thermodynamic of bacteria growth, the phenomenology and the mosaic approaches. Biochim Biophys Acta 683, 181-220. Read More