Lecture 2: Bacterial Growth Flashcards
Learning objectives
To understand and be able to describe:
- energy and nutrition requirements
- the bacterial growth cycle
- methods of measured growth
- spores
Discuss bacterial nutrition.
- Major elements in cell (macro-nutrients, required in large amounts): C, H, O, N
- Minor elements in cell (macro-nutrients, required in smaller amounts): P, S, K, Mg, Ca, Na
- Micro-nutrients, required in trace amounts: Fe, other metals
70-80% wet weight of cells is water, most of dry weight is macromolecules, e.g. protein, RNA, DNA, polysaccharide, lipid.
What are the possible energy sources for bacteria?
Chemicals (chemotrophy) - most bacteria
Light (Phototrophy)
Organic chemicals are used by Chemoorganotrophs.
Inorganic chemicals are used by Chemolithotrophs.
Light is used by Phototrophs.
What is a heterotroph?
An organism which uses organic carbon.
Give 4 alternative sources of energy to carbon.
1) Sulphur
2) Nitrogen (nitrifying bacteria)
3) Iron
4) Light
Give the steps of bacterial cell division.
1) DNA replication
2) Cell elongation
3) Septum formation
4) Completion of septum formation with formation of distinct cell walls
5) Cell separation
What is an FtsZ ring?
A protein ring which forms at septum of dividing bacteria..
What is the generation time?
The doubling time
How long is the generation/doubling time in a normal bacterial culture?
30 minutes
What is the exponential phase?
The phase of bacterial growth during which the doubling/generation time is constant. The rate of increase of cell number is increasing. This phase only continues when there are no limiting factors. The growth rates are limited by temperature, nutrients, pH and genetic factors.
Which equation can be used to find the number of bacterial cell present in a culture during exponential phase?
N = N0 x 2^n
where:
N = final cell number
N0 = initial cell number
n = number of generations
What is the lag phase?
The lag phase happens at the beginning of introduction into a new environment. It occurs because the cells are still adapting to the environment. The inoculum is usually depleted of certain nutrients and time is required for resynthesis.
The length of the lag phase can vary greatly.
What is the stationary phase?
- The cell division rate is equal to the death rate, so there is no increase or decrease in cell number.
- Growth is limited by a limiting factor such as nutrients.
- Accumulation of storage products
- Certain genes are necessary for survival in stationary phase
- Sporulation commences in certain species.
- Not all the cells are dead
What assumptions are made in a viable cell count?
That each viable cell will give rise to one colony.
Explain the two types of plating used during viable cell counts.
Spread plate - cells put on top of agar (surface colonies)
Pour plate - cells mixed with agar (surface and subsurface colonies)
Describe the procedure of serial dilutions for viable cell count.
A series of test tubes containing 9 ml of broth are prepared. Then 1 ml of the original sample is removed and added to the first test tube. This creates the 10^-1 dilution. Then 1 ml of this mixture is added to the second test tube, creating the 10^-2 dilution. This is repeated.
1 ml samples of each dilution is then plated and incubated. The number of colonies on each plate is then counted. The highest number which is countable is chosen and the number of colonies is multiplied by the dilution factor (if it is the 10^-3 dilution, the dilution factor is 10^3) to give the number of viable bacterial cells in the original sample.
What is the effect of temperature on growth processes?
Low temperature - membranes tend to become gel (crystalline) and transport through the membranes becomes limiting.
High temperature - rate of chemical reactions and enzymes are high
Very high temperature - Protein denaturation, collapse of the cytoplasmic membrane, thermal lysis.
What are endospores?
Differentiated cells which are resistant to heat and very difficult to destroy. They are also resistant to drying, radiation, acids, chemical disinfectants. They are dormant so they do not grow or divide.
What can destroy spores?
Autoclaving (high temperature, high pressure)
Which bacterial species mostly commonly form spores?
Bacillus and Clostridia species (Gram positive)
e.g. Clostridium Botulism, Bacillus anthracis
How long does it take for a spore to made and released from a vegetative cell?
8 hours
How are spores and vegetative cells linked?
A vegetative cell can become a sporulating cell and then a spore.
A spore can germinated under the correct conditions, undergo outgrowth and become a vegetative cell.
What are the stages in sporulation?
Vegetative cells have membrane invaginations normally. 1) The DNA condenses.
2) Cytoplasmic membrane invaginates to form spore septum. This is asymmetric cell division then occurs. The cytoplasmic membrane engulfs the ‘forespore’.
5) The spore septum grows around protoplast (engulfment). The exosporium appears; primordial cortex is formed between the two membranes
6) The cell begins to dehydrate. Ca2+ incorporation into spore; further dehydration, production of SASPs (small acid-soluble spore proteins) and dipicolinic acid synthesis.
7) Coat layers are formed (proteins form outside spore)
8) The cortical layers mature and develop resistance to heat and chemicals)
9) Lysis of mother cell and release of free spore surrounded by exosporium.
What do SASPs do?
SASPs (small acid-soluble spore proteins):
- bind DNA and protect it from UV light, dessication and dry heat
- carbon and energy source for outgrowing vegetative cell
