Bacterial growth and genetics

Question 1

How do Bacteria multiply?

Answer 1

By Binary fission

Question 2

What does Binary fission result in?

Answer 2

Results in two identical daughter cells

Question 3

What does the time of division depend on?

Answer 3

Environmental and species.

For example
E. coli 37oC with lots of food = 20 mins »(chromosome replication takes 30 min!)

Question 4

What does Rapid growth allow?

Answer 4

Rapid evolution of traits by mutations or recombination

For example - Resistance to chemical ages -antibiotics and biocides.

Question 5

What is the first step for Bacterial growth?

Answer 5

Chromosome replication

Question 6

In Bacterial growth what follows after Chromosome replication?

Answer 6

Septation (Continued growth of the cell) and division into two cells

Question 7

What are the 4 phases in Bacterial growth and replication?

Answer 7

Lag phase

Exponential log phase

Stationary phase

Death phase

Question 8

Why is the idealised growth curve of bacteria in a culture important experimentally?

Answer 8

because at different phases cells exhibit different characteristics.

Question 9

What are the two types of measuring growth or bacterial number in a sample?

Answer 9

Direct or indirect

Question 10

How does Direct count work?

Answer 10

Uses Microscopy
Membrane filtration which is mostly for water
Plate counts

Question 11

How does indirect count work?

Answer 11

Turbidity using spectrophotometry
Biomass
Measuring cell products - CO2

Question 12

Explain viable count

Answer 12

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

Question 13

How does spectroscopy estimate bacterial number?

Answer 13

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

Question 14

When is Turbidity used?

Answer 14

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.

Question 15

How can growth be used to identify an organism?

Answer 15

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

Question 16

How can growth be used to identify an organism- What are the 2 of many systems developed to exploit the differences?

Answer 16

Manual –agar plates – biochemical tests

Mechanised – robotic biochemical assays

All rely on differences in cellular biochemistry

Question 17

List the parts of a bacteria

Answer 17

Capsule
Ribosomes
Cytoplasm
Plasma membrane
Pili
Cell wall 
DNA
Flagellum

Question 18

List the parts of the bacterial chromosome

Answer 18

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

Question 19

Explain Chromosome replication

Answer 19

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

Question 20

What phase in the growth curve is of most danger to humans?

Answer 20

Exponential phase

Question 21

What is required for each phase in the growth curve?

Answer 21

A different subset of genes is required for each phase.

Question 22

How does chromosome vary?

Answer 22

it varys significantly between species

Question 23

Explain the recent research into genome sequencing

Answer 23

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.

Question 24

What are Operons?

Answer 24

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.

Question 25

How is gene expression controlled?

Answer 25

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.

Question 26

What are Regulon's?

Answer 26

A group of operons that is co-regulated Toxins and Pathogenicity islands.

Question 27

What are plasmids?

Answer 27

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

Question 28

How do plasmids spread?

Answer 28

Spread naturally by conjugation between donor cells and recipient cells (Sex pilus)

Question 29

What is the typical size and shape of a plasmid?

Answer 29

Typically circular, DS and <5% size of chromosomes - 1 to 5 x 10^5 bp

Question 30

How do plasmids replicate?

Answer 30

Replication is similar to that of the chromosome and starts at origin of replication (Ori V).

Question 31

In research what can plasmids be used for?

Answer 31

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

Question 32

Medical importance of plasmids- How is it involved with Antibiotic resistance?

Answer 32

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

Question 33

Medical importance of plasmids- How is it involved with Virulence factors?

Answer 33

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

Question 34

Explain mutation in a bacterial gene

Answer 34

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

Question 35

Explain the types of mutation

Answer 35

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

Question 36

Explain what mutations can do

Answer 36

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.

Question 37

Explain Recombination

Answer 37

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.

Question 38

Explain Transformation

Answer 38

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.

Question 39

Explain Transduction

Answer 39

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

Question 40

Explain Conjugation

Answer 40

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.