Bacterial Evolution

Question 1

Bacterial classification levels

Answer 1

over 90 phylum
Under species there is strain
Under strain there is isolates
- obtained from pure cultures

Question 2

Biochemical profiling

Answer 2

Species have set of specific enzymes
Use API strips to test which enzymes present in culture
If species infectious can use antibiotic susceptibility test

Question 3

Different phenotypic species ID methods

Answer 3

• MALDI-TOF mass spectrometry
• Chemotaxonomic markers
• Expressed features

Question 4

MALDI-TOF mass spectrometry

Answer 4

Samples are ionised into changed molecules
Measure their ratio mass to charge
= each species unique ratio

Question 5

Chemo taxonomic markers

Answer 5

Variation between species in:
Fatty acids
Cell wall compounds
Exopolysaccharide

Question 6

Expressed features in bacteria

Answer 6

Physiology
Morphology
Serology
Antibiotic resistance

Question 7

Genotypic methods of bacterial species ID

Answer 7

DNA. 
- need >70% to be similar 
DNA segments 
- sequence discriminatory/housekeeping genes to see variation 
- e.g. recA, gryB
rRNA 
- bac = 16s rRNA (eukarya = 18s rRNA) 
- need >97% similarity 
Whole genome sequencing 
- look at average nucleotide identity (ANI)
- want >95% ANI between genomes

Question 8

Species

Answer 8

Large number of strain with similar phenotypic/genetic properties
- >70% DNA-DNA hybridisation, >97% 16s rRNA gene sequencing similarity, >95% ANI whole genome sequencing

Question 9

Strain

Answer 9

When there is knowledge of clonality or genetic identity

Using strain typing techniques to identify

Question 10

Isolate

Answer 10

Cultures from an infection or elsewhere

When you have no genetic knowledge but its a single pure colony

Question 11

Phenotypic strain typing techniques

Answer 11

Serotyping
Resistotyping
Biotyping
MALDI-TOF mass spec

Question 12

Serotyping

Answer 12

Serotype = distinct variation within bacteria species
- classified via cell surface antigens = epidemiological classification
- take sample and mix with blood containing antibodies = antiserum
- blood will agglutinate (clump)
Resulting combination of antigens defines serotype

Question 13

Resistotyping

Answer 13

• testing bacterial strains against arbitrarily chosen chemicals
• show the selective toxicity at a critical concentration

Question 14

Biotyping

Answer 14

Identifying bacteria based on a series of bios chemical tests
The same as biochemical profiling ?

Question 15

Genotypic strain typing techniques

Answer 15

DNA finger printing 
- Restriction analysis 
- PCR based methods 
Sequence based ID
- gene sequencing 
- multi locus sequence typing (housekeeping genes)
- whole genome sequencing

Question 16

Restriction analysis

Answer 16

Using restriction enzymes to digest DNA
Will cleave DNA at restriction sites
Look at sizes using gel electrophoresis

Question 17

Desirable trains when strain typing

Answer 17

• typeability: how many isolates an you type?
• reproducibility: consistent results?
• accuracy
• discrimination: diff strains from same species?

= TRAD criteria

Question 18

Phenotypic or genotypic better at ID?

Answer 18

Genotypic more likely to meet TRAD criteria
- discriminatory
- more stable
Phenotypic relies on growth and can be variable

Question 19

Describe the bacterial genome

Answer 19

Highly variable in size from 100-10 mil bases
G + C content = 20-75%
One double circular DNA chromosome
Almost all coding material

Question 20

Accessory genome elements

Answer 20

• Non essential genes
• foreign DNA
• drives diversity between species = rapid evolutionary change e.g. plasmids
• multiple genetic applications
• contains bacteriophages, genomic islands, ICEs

Question 21

Core genome elements

Answer 21

• essential genes
• stable GC content
• shared within species but also suggests diversity
• slow evolution
• stable basis for genotype growth: wont change often but therefore good at showing diversity between species

Question 22

The bacterial pan genome

Answer 22

• entire collection of genes found in a species
• overlay strains and there will be a core section of shared genes
• the moe strains you compare the smaller the core genomes

Question 23

What drives bacterial genome evolution?

Answer 23

• horizontal gene transfer
• recombination
• gene duplication
• mutation
• additive evolution
• reductive evolution

Question 24

Additive evolution

Answer 24

Gene acquisition e.g. plasmids, transposons, genomic islands
Gene duplication and rearrangement

Question 25

Reductive evolution

Answer 25

Deletion: loss of 1+ genes

Inactivation and pseudogene formation

Question 26

Horizontal gene transfer

Answer 26

Transformation
Transduction or transfection
Conjugation
Other mobile genetic elements

Low freq but causes rapid evolution

Caused by plasmids, bacteriophages, transposable elements genomic islands

Question 27

Genomic horizontal transfer and genomic islands

Answer 27

• large regions of DNA integrated into the genome
• not present in all strains of a species
• can cause selective advantage

Question 28

Transformation

Answer 28

Foreign DNA incorporated into genome

Question 29

Transduction

Answer 29

Bacteriophage mediated transfer

Question 30

Conjugation

Answer 30

DNA transferred by cell-cell contact

E.g. plasmids

Question 31

Gene cloning vectors

Answer 31

Plasmids
Phases
Cosmics (phage and plasmid hybrid)

Question 32

Transposon mutagenesis

Answer 32

• deliver transposon into host cell
• transposon integrates into host genome
• disrupts gene function
• one copy per cell. Integration is in a random location
• if a gene is disrupted you can see where its location by identifying transposons location

Good for finding genes of interest