Bacterial growth and genetics Flashcards
How do Bacteria multiply?
By Binary fission
What does Binary fission result in?
Results in two identical daughter cells
What does the time of division depend on?
Environmental and species.
For example
E. coli 37oC with lots of food = 20 mins »(chromosome replication takes 30 min!)
What does Rapid growth allow?
Rapid evolution of traits by mutations or recombination
For example - Resistance to chemical ages -antibiotics and biocides.
What is the first step for Bacterial growth?
Chromosome replication
In Bacterial growth what follows after Chromosome replication?
Septation (Continued growth of the cell) and division into two cells
What are the 4 phases in Bacterial growth and replication?
Lag phase
Exponential log phase
Stationary phase
Death phase
Why is the idealised growth curve of bacteria in a culture important experimentally?
because at different phases cells exhibit different characteristics.
What are the two types of measuring growth or bacterial number in a sample?
Direct or indirect
How does Direct count work?
Uses Microscopy
Membrane filtration which is mostly for water
Plate counts
How does indirect count work?
Turbidity using spectrophotometry
Biomass
Measuring cell products - CO2
Explain viable count
Allows the approximate quantity of organisms that are “Alive” to be counted in a sample.
For example: bacteria and yeasts
Spread plates
Pour plates
Miles and Misera
However doesn’t take into account viable but non-culturable organisms (VBNC’s).
is an approximation only
How does spectroscopy estimate bacterial number?
It uses a light source to pass through the bacteria sample and hit a photocell detector. The more light that hits the photocell the less bacteria there is.
However the less light that hits the photocell the higher amount of bacteria
When is Turbidity used?
When a growth curve is plotted using plate counts along with the optical density of a sample from a growing culture at 550 – 600 nm, bacterial number can be determined using only turbidity.
How can growth be used to identify an organism?
As bacterial cells grow they utilise nutrients or carbon sources in different ways and produce enzymes.
Allows a metabolic fingerprint to be obtained- Characteristic to the organism
How can growth be used to identify an organism- What are the 2 of many systems developed to exploit the differences?
Manual –agar plates – biochemical tests
Mechanised – robotic biochemical assays
All rely on differences in cellular biochemistry
List the parts of a bacteria
Capsule Ribosomes Cytoplasm Plasma membrane Pili Cell wall DNA Flagellum
List the parts of the bacterial chromosome
Double stranded circular DNA molecule Condensed into the nucleiod No nuclear membrane Chromosome is unpaired - Haploid Encodes all the genes required for essential cell functions
Explain Chromosome replication
This must happen for cell division
Replication is semi-conservative - each copy made contains one new (daughter) and one old strand (parent).
Replication starts from a specific site the origin of Replication (ori C)
Bi-Directional
Uses a large collection of enzymes- Replisome
What phase in the growth curve is of most danger to humans?
Exponential phase
What is required for each phase in the growth curve?
A different subset of genes is required for each phase.
How does chromosome vary?
it varys significantly between species
Explain the recent research into genome sequencing
Efficient methods for genome sequencing available only from 1995.
Whole genome “Shot gun” sequencing.
Venter and Smith where the first to sequence Haemophilus influenzae and Mycoplasma genitalium.
What are Operons?
Functionally linked genes are grouped together. The expression is controlled by a single operator which together is called a Operon.
All genes in a operon are transcribed onto one large messenger RNA molecule. This ensures that all the genes required for a particular metabolic task are expressed and active at the same time.
How is gene expression controlled?
By proteins which regulate the rate of gene transcription.
Repressors (Negative control - down regulate).
Activators (positive control - up regulate)
Can be specific proteins or products such as levels of catabolites - glucose.
What are Regulon’s?
A group of operons that is co-regulated
Toxins and Pathogenicity islands.
What are plasmids?
Extra-chromosomal genetic elements that replicate independently from host chromosome
Not always essential for survival or the bacterium - lab vs host
Thousands of different plasmids exist
- 300 isolated from strains of E. coli alone
- Possible to make them in the lab - Plasmid vectors for cloning
How do plasmids spread?
Spread naturally by conjugation between donor cells and recipient cells (Sex pilus)
What is the typical size and shape of a plasmid?
Typically circular, DS and <5% size of chromosomes - 1 to 5 x 10^5 bp
How do plasmids replicate?
Replication is similar to that of the chromosome and starts at origin of replication (Ori V).
In research what can plasmids be used for?
A typical plasmid that is used as cloning vector genes of interest can be inserted in the multiple cloning site (MCS) and are expressed once introduced to a cell
Medical importance of plasmids- How is it involved with Antibiotic resistance?
Called R factors
First discovered in Japan in early 1940’s
Usually enzymatic inactivation of the antibiotic
Beta-lactamases - penicillin resistance
Prevent uptake by modifying cell wall
Increase of R plasmids correlates with increasing use of antibiotics
Medical importance of plasmids- How is it involved with Virulence factors?
Toxins
Enteropathogenic E.coli strains - enterotoxin
S.aureus - enterotoxin B
B. anthracis - PA, LF, EF and capsule.
Adherence factors - pili in E.coli
Siderophores - Iron important for pathogens
Explain mutation in a bacterial gene
In heritable change within a nucelotide (DNA) sequence.
A bacterial strain carrying such a change is called a mutant.
Call be
Spontaneous- natural radiation, errors during DNA replication
Induced- chemical mutagens
Explain the types of mutation
Point mutation - affects one base pair
Insertion or deletion of large DNA fragment.
usually due to a transposon
may result in loss of gene function
inversion and duplications
Explain what mutations can do
Can alter the function of a gene
Replacement of nucleotide resulting in amino acid substitution in a protein (missense).
Premature termination of protein synthesis (nonsense).
Change in expression levels due changed regulation - deletions or insertions.
Explain Recombination
Genes in two separate bacterial cells brought together in one.
Occurs by transfer of genetic information between bacteria - have a common gene pool
Swap genes through horizontal gene transfer
includes genes of plasmids
Happens through - Transformation, Conjugation and transduction.
Explain Transformation
Uptake of pure (naked) DNA from medium surrounding cell.
Released into medium from lysed cells
DNA must integrate into chromosome to transform recipient cell - Transformants
Can be induced in the lab - Plasmids
This can also be a way that plasmids are introduced into the cells, this process can happen naturally or can be induced in the lab to create mutants.
Explain Transduction
DNA transferred cell to cell by Bacteriophages (bacterial viruses).
bacteriophage picks up partially degraded bacterial DNA instead of its own by mistake when making new virus - transducing phage.
Not all bacteriphages can transduce and not all bacteria can be tranducible
Explain Conjugation
Cell to cell contact allowing transfer of DNA
Plasmids are self- transmissible by conjugation
Encode the sex pilus and other transfer genes (Tra region).
is the major mechanism of transfer of antibiotic resistance genes on R plasmids.
