Bacterial Genetics

Question 1

How does genetic material in bacteria differ from our genetic material

Answer 1

Bacteria-

• single circular chromosome, less than 5Mb
• DNA NOT compartmentalized, localized in bacterial cytoplasm
• often contain plasmids in addition to chromosome
• chromosome condensed by supercoiling DNA

Eukaryotes-

• several to many linear chromosomes, greater than 5 Mb
• DNA compartmentalized into organelles: nucleus and mitochondria
• plasmids NOT normally found
• chromosomes condensed by wrapping DNA around histones

Question 2

Describe bacterial replication

Answer 2

• single origin
• the second round of replication can begin before the cell divides
• Replicated DNA partitioned into daughter cell-binary fission

Question 3

Describe some differences between prokaryote and eukaryote transcription/translation

Answer 3

• Prokaryote
• genes encoded within operons
• NO inrtons or exons aka no splicing
• only 1 RNA polymerase
• transcription and translation are coupled in bcteria bc there is no compartmentalization
• ribosomes are 50s+30s=70s
• Eukaryotes
• NO operons
• introns and exons-splicing
• 3RNA polymerases

Question 4

how do bacteria generate genetic diversity

Answer 4

Following binary fission, all daughter cells are clones

BUT you can identify clones with genetic alteration

for example some clones are antibiotic resistant

Question 5

How does drug resistance develop?

Answer 5

replication errors introduce mutation into genes-mediated by DNA polymerase

• DNA polymerase misincorporates nucleotide and does not correct the mistake through proof reading activity
• frequency is 1 mutation per 300 chromosomes replicated so 10-6 or 10-7 mutations per genome per generation
• however most mutations do not confer a selective advantage

But sometimes they do!!

Question 6

Besides random mutations, how else can bacteria develop drug resistance

Answer 6

• exchanging genetic material (ie DNA)
• note there is no natural species boundary in prokaryotes like there is for eukaryotes (aka they can share DNA very easily)
• bacteria do both horizontal transmission and vertical transmission

Question 7

List 4 important concepts about gene exchange in bacteria

-think why is gene exchange helpful, what does it cause and why is ti possible

Answer 7

• The species barrier in bacteria is much less stringent than in eukaryotes, so bacteria are promiscuous with their DNA
• provides bacteria with mechanism to create genetic diversity. you only need a single organism in a population to survive
• gene exchange provides bacteria with selective growth/survival advantage in certain environments. it helps outcompete or kill other organisms
• horizontal gene transfer is largely responsible for rapid spread of antibiotic resistance. Vancomycin resistance from VRE to VRSA
• Exchange of genetic material between bacteria occurs with great frequency and efficiency outside and within a host

Question 8

bacteria exchanage a lot of genes. What genes are of most concern to physicians

Answer 8

virulence factors and antibiotic resistance determinants

Question 9

what are the 3 main types of mobile genetic elements by which genes are carried between bacteria

Answer 9

1. plasmids
2. transposable genetic elements
3. pathogenicity islands

Question 10

What is a plasmid

Answer 10

• ss or ds DNA molecule that replicates independently of the bacterial chromosome
• ost often circular, but can also be linear
• can be single (F plasmids or episomes) to multiple copies in the cell (~500 copies)
• vary in size from 1500bp to 400,000bp
• can be transferred between bacteria by transformation, conjugation, and transduction

Question 11

Whats another way to think of plasmids

Answer 11

You can think of plasmids as small chromosomes that carry virulence or antibiotic resistance genes and that can be transferred between bacteria

Question 12

What are transposable genetic elements

Answer 12

-transposable gentic elements are linear DNA segments that can be mobilized from one location in the genome to another. They often disrupt gene(s) in recipient bacterium following their transposition

Question 13

Tell me about the replication of transposable elements

Answer 13

• they cannot replicate on their own.
• once transferred the element can transpose from one location to another.
• they must be present on a replicon (ie plasmid or chromosome) to be maintained and passed on to daughter cells

Question 14

How are genetic elements able to transpose from one location to another

Answer 14

• they posses inverted terminal repeats at their ends
• There is an enzyme called a transposase (tnp) that recognizes the ITR and cuts the DNA allowing transposition from one location to another

Question 15

What are the four types of transposable elements

Answer 15

• Insertion sequences (see example of fimbriae production in UPEC)
• Composite transposons
• TnA family transposons (transposase enzyme is regulated)
• Mu bacteriophage (part of bacteriophage genome)

Question 16

Describe the example of an insertion sequence transposon (fimbriae)

Answer 16

• this is an example of an insertion sequence in bacterial genome that regulates production of a virulence determinant
• there is phase variation of fimbriae production (meaning not always happening) in uropathogenic Excherichia coli
• the insertion sequence undergoes site-specific inversion
• this causes spontaneous switch between non-fimbriated and fimbriated forms
• fimbriae promote attachment to urinary tract epithelial cell surface

Question 17

What are genomic pathogenicity islands

Answer 17

-large segment of bacterial genome that is carried on
a plasmid or by a bacteriophage
-often encode determinants that give bacteria survival advantage within a specific environment
-G and C content differs from majority of chromosome bc it was acquired by horizontal gene transfer from another organism
-they cannot replicate by itself. Must transpose onto a replicon (ie plasmid or chromosome) in the recipient to be maintained and passed on to daughter cells following replication
-vary in size from 10,000bp to 20,000 bp

Question 18

How do you identify genomic pathogenicity islands

Answer 18

Following sequencing of bacterial genomes

Question 19

in one sentence what are genomic pathogenicity islands

Answer 19

large DNA segment that differs in GC content from rest of genome

Question 20

What do pathogenicity islands often encode for

Answer 20

adherence factors, invasion genes, iron uptake systems, protein secretion systems and toxins

Question 21

How do pathogenicity island replicate

Answer 21

they cannot replicate by itself. Must transpose onto a replicon (ie plasmid or chromosome) in the recipient to be maintained and passed on to daughter cells following replication

Question 22

What are the three main mechanisms by which plasmids, transposable elements and pathogenicity island are transferred between bacteria

Answer 22

1. Transformation- naked DNA taken up from the environment (no specificity, just being in right place at right time)
2. Conjugation-direct cell to cell contact (mating bridge)
3. Transduction- mediated by bacteriophage

Question 23

Explain how transformation was first identified in the study of capsulated strains of Streptococcus

Answer 23

• living non encapsulated (not virulent) bacteria was injected in a mouse and the mouse was fine
• heat killed virulent bacteria was injected in a mouse and the mouse was fine
• BOTH living non-virulent DNA and heat killed virulent DNA were injected into a mouse and all mice died and all living bacteris was virulent
• this is bc even though the DNA was dead it stil transferred all of it virulence factors to the living bacteria making them virulent!

Question 24

Bacterial transformation

Answer 24

• uptake of DNA from teh environment
• Occurs in both gram positive and gram negative bacteria
• some bacteria are naturally “competent” others can be induced to be competent for DNA uptake by environmental conditions
• DNA is released from donor bacteria following lysis
• dsDNA bound b the recipient is then processed to ssDNA before it is internalized
• uptake of plasmids or segments of chromosomal DNA/stable inheritance of chromosomal DNA in recipient requires recombination with host chromosome

Question 25

Conjugation

Answer 25

• DNA is intentionally passed unidirectionally from donor to recipient
• requires physical contact*****
• occurs in both gram positive and gram negative bacteria
• requires a complex set of genes
• DNA is transferred through a mating bridge***** (formed from pilus)
• the donor retains a copy of the original genetic material as well.

-think of this like sharing notes. you aren’t just going to throw them out into the world so that you don’t have them anymore and anyone could find random notes that might not even help them. You would find your friend who you know they would help. You would make a copy so you still had a copy and then you would directly hand it to them to make sure they got it

Question 26

What are bacteriophages

Answer 26

• obligate intracellular parasites ( bacterial viruses)
• they are single nucleic acid molecules protected by protein coat or capsid
• they have a variable lifecycle: lytic, lysognic, or temperate (capabale of both lytic and lysogenic cycles)
• responsible for transduction-transfer of DNA

Question 27

Lytic phase

Answer 27

• make new progeny phage=phage genome replicated independently of host genome
• phage DNA maintained and replicated separately from bacterial genome. Unlikely to be transferred vertically to progeny. Goal is for phage to produce more progeny bacteriophage

Question 28

Lysogenic

Answer 28

-vertical transmission of phage=phage genome integrated and replicated with bacterial genome
- phage DNA integrates into bacterial genome and is replicated along with the
bacterial genome. Phage DNA able to be transferred vertically to daughter cells.

Question 29

Temperate

Answer 29

• temperate phage can switch between these lifecycles (lytic and lysogenic)
• phage can do both lytic and lysogenic lifecycles.

Question 30

Tranducing pahge

Answer 30

• sometimes, during the lytic replication of phage, a segment of bacterial genome is accidentally packaged into a phage particle rather than phage genome. The resulting progeny are the transducing phage
• the transducing page then goes and enters other bacteria. upon infection (injection/entering) a new bacterium, the transducing phage releases fragment of bacterial genome into the host cytoplasm. this is called transduction
• recombination between introduced bacterial DNA and endogenous host DNA results in stable inheritance of transferred material

Question 31

Transduction

Answer 31

the transducing phage releases fragments of the bacterial genome into host cytoplasm

Question 32

Transductant

Answer 32

transferred material?

Question 33

Some bacteriophages carry virulence mechanisms what are two examples

Answer 33

Cholera toxin and Shiga toxin

-note this is different than a transducing phage bc toxins are a part of the page genome

Question 34

Cholera phage toxin narration

Answer 34

-infects vibrio cholera
-comprised of core element and repetitive sequence
-phage genome contains ctxAB which encodes the Cholera toxin. Strains of
Vibrio cholera are infected with this bacteriophage. Cholera toxin produced by the organism is released
during infection and acts to ADP-ribosylates ganglioside GM1 on the surface of gut epithelial cells,
leading to activation of adenylate cyclase and secretion of water and electrolytes resulting in watery
stools (i.e. rice water stools).

Question 35

Shiga Phage toxin

Answer 35

phage genome contains stxAB which encodes Shiga toxin.
-the phage is a temperate phage**it is both lytic and lysogenic

Strains of Shigella
dysenteriae and EHEC carry this bacteriophage.

Shiga is toxin produced and released by these bacteria, and binds Gb3 glycolipid on the surface of host cells,

it is trafficked into the cell, and modifies ribosome
acceptor site to disrupt protein synthesis.

This toxin can cause severe bloody diarrhea and hemolytic-uremic
syndrome. (HUS)

Question 36

Acquisition of new determinants via horizontal gene transfer allows bacteria to

Answer 36

gain selective advantage within a given niche

Question 37

Plasmids, transposable genetic elements, and pathogenicity islands can……

Answer 37

be exchanged and often encode virulence determinants or antibiotic resistance genes

Question 38

Transformation, conjugation, and transduction are all……

Answer 38

mechanisms by whic DNA is transferred from donor bacterium to recipient bacterium

Question 39

Cholera toxin and Shiga toxin are both examples of

Answer 39

virulence factors that are carried on a bacteriophage and that are part of the normal bacteriophage genome