Synchronous culture
Until now growth patterns of bacterial populations have been described, i.e. this is a reflection of the average state of the culture. From these studies one cannot gain any information regarding an individual cell because a bacterial culture at any timepoint probably contains cells in all growth phases. However, information on the growth behaviour of individual cells can be obtained from synchronous culture because all the organisms are at the same stage of growth.

Synchronous cultures can be obtained by:

i) inducing synchrony by such methods as manipulating the environmental conditions (usually in a cyclic fashion. e.g. by repeated shifts in growth temperature).
ii) selecting synchronous cells from a random population e.g. cell cycle stage may provide some physical conditions (density differences).

Synchronous cultures rapidly lose synchrony because various cells within a population do not all divide at the same stage.

Continuous culture of microorganisms
Types of cultures discussed so far are called batch cultures, and nutrients are not renewed. Consequently, exponential growth is only for a few generations. However, microbial populations can be maintained in exponential growth over a long time by using a system of continuous culture. Generally done in a Chemostat, here the growth chamber is connected to a reservoir of sterile growth medium and a siphon overflow spout (see Figure 6:1). So once growth starts, fresh medium is supplied at a constant rate, with the volume in the chamber kept constant - in essence the growth of bacteria is equivalent to the number that are lost through the spout. Growth is highly dependent on the rate of inflow - however, no matter how fast the media goes in the bacteria cannot grow faster than they would in batch culture under the conditions employed within the chamber.

Question arises: how can the growth rate be a function of input of fresh medium? The explanation is that continuous cultures are set up so that one nutrient is always limiting.

Environmental Factors Affecting Bacterial Growth
(A) physical
(B) nutritional factors.

(A) Physical Factors Affecting Growth:

i) pH - measurement of the acidity or alkalinity of the medium
Microorganisms have an optimum pH for growth and generally most bacteria grow best at neutral pH. However, some grow best at acidic pH and are called acidophiles e.g.Lactobacillus, whereas, other grow best at high pHÕs and are called alkaliphiles e.g.Vibrio cholerae

ii) temperature - most species can grow over a 30°C temperature range, with the maximum and minimum temps varying greatly for different species, e.g because sea water remains liquid below 32°F, bacteria living in the sea can tolerate temperatures below freezing, likewise, some bacteria can tolerate and grow in temperatures around 75°C (a very hot bath for us is about 60-65°C).

a) psychrophiles - cold-loving bacteria that grow best at 15-20°C. These bacteria generally live in cold water and soil - none can survive in the human body. These are the bacteria that cause food to spoil in the fridge.

b) mesophiles - majority of bugs, grow best between 25-40°C. Human pathogens are included in this group.

c) thermophiles - heat-loving bacteria, grow best at temperatures 50-60°C. Some can survive temps as high as 110°C in boiling hot springs, or, in deep-sea vents.

The temperature effect is generally on enzyme stability i.e. high temperatures tend to denature proteins. However, thermophiles are becoming attractive to industry because chemical reactions tend to be more efficient at higher temps. This means less money need be spent to obtain the final product.

(B) Nutrient factors Affecting Bacterial Growth

Generally the concentration of solutes (i.e. chemical growth components) is higher within the microbial cell than in the extracellular environment. The major barrier governing this differential passage of chemical components is the cell membrane.

Membrane function is:
i) keep essential nutrients and macromolecules inside the cell.
ii) pump certain nutrients inside the cell against a concentration gradient.
iii) permit free flow of nutrients across the membrane.
iv) exclude some solutes within the environment from entry into the cell.

For the great majority of bacteria the cytoplasmic membrane is the sole unit membrane within the cell (exception are the blue-green bacteria).

Cell membranes are a complex mixture of protein, phospholipids and glycolipids (see Figure 6:2).
A) lipid components: The bacterial lipid components form a typical bimolecular leaflet, with the hydrophobic components facing each other and the polar head groups interacting with the internal and external aqueous environments. Water is excluded from a membrane due to the hydrophobic core. A major difference between prokaryotic and eucaryotic membrane systems is that bacterial membranes lack sterols such as cholesterol (exception is the Mycoplasma group). Secondly, the fatty acid components of the membrane phospholipids are generally saturated fatty acids (i.e. donÕt contain polyunsaturated fatty acids - eucaryotic membranes do).

B) protein components: protein components can either span the membrane or alternatively be exposed on a single face. Therefore, proteins must contain polar regions (i.e. these parts can interact with the water molecules on the external and internal surfaces) as well as contain hydrophobic regions so that they can insert into the hydrophobic lipid core of the membrane. The protein components in the cell perform specialized tasks, e.g.energy-yielding metabolism (i.e. they contain the components of the respiratory electron transport system), or, alternatively, be involved in nutrient uptake.

C) glycolipid components: these are specialised lipids that contain a carbohydrate moiety as well as fatty acids. These molecules perform special functions such serving as attachment sites to eucaryotic cells.

Movement of Nutrients Across a Membrane
Problem: The translocation of polar molecules across a membrane is thermodynamically unfavorable due to the hydrophobic nature of a membrane. Consequently, very small polar molecules such as water, ions etc need pores to pass through the membrane. Non-polar molecules such as lipids or non-charged molecules will dissolve into the membrane and subsequently pass through. The mechanisms by which they achieve this are either passiveÊor active.

Passive transport mechanisms, which require no energy input, include:
a) simple diffusion.
b) facilitated diffusion.
c) osmosis.

Active transport mechanisms, which require energy input, include:
a) transport against a concentration gradient.
b) endocytosis.
c) exocytosis.

Passive Mechanisms
a) Simple Diffusion (see Figure 6:3)
All molecules contain kinetic energy (which means they are in constant motion) and move from regions of high concentration to low concentration. This is a random process. Why do they move from high to low? As they move they collide with other molecules, therefore they dont travel in straight lines, and get bumped around. Therefore, in regions of low concentration, they have less bumping going on to knock them back into regions of high concentration. Also they molecules in the high concentration will present a ÒwallÓ of molecules that will knock them back into the low concentration region. The length of time required to reach equilibrium depends on the volume of the cell (small size will reach equilibrium faster than a larger sized object).

Diffusion through a membrane is affected by several parameters:
i) solubity of the solute with the membrane.
ii) temperature (higher the temp the more kinetic energy in the fatty acids chains in the membranes, therefore more wiggling around (makes the membrane more fluid).
iii) The concentration difference (i.e. if concentration difference is extremely high then more molecules will be participating).

Therefore, non-polar substance such as steroids and gases (CO2 and O2) diffuse rapidly across a membrane. Polar solutes (H+, K+, Na+, and Cl-) diffuse through pores (i.e. protein tubes). Diffusion through pores depends upon the diameter of the pore.

b) Facilitated Diffusion (see Figure 6:4)
Facilitated diffusion is diffusion down a concentration gradient (i.e moves from high to low). This is achieved with the assistance of a carrier protein that spans the lipid bilayer. The carrier molecules are proteins that bind to one or more molecules, and through a mechanism like a revolving door provides a convenient channel for the passage of molecules. However, the system can become saturated when all external sites are occupied - once saturated, diffusion reaches a maximum and cannot go any higher. The carrier molecules donÕt have much specificity, with different molecules competing against each other. Movement does not require the expenditure of energy.

 

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