L8 - Horizontal Gene Transfer

Question 1

What are the sources of genetic diversity?

Answer 1

• spontaneuous mutagenesis = DNA polymerase has proofreading properties but its not completely perfect
• errors in DNA repair
• homologous recombination = deletion/duplication/inversion
• transposons = jumping genes, affects functions, occurs naturally (transposable elements)
• exposure to mutagens (chemical agents and radiation)
• horizontal gene transfer

Question 2

What is horizontal gene transfer?

Answer 2

movement of genetic material between organisms other than by the transmission of DNA from parent to offspring

Question 3

What does HGT do?

Answer 3

• genome diversification
• evolution
• adaptation
• survivability
• selection = antibiotic resistance

Question 4

How can HGT be advantageous to human?

Answer 4

pros to human :
1. evolution = discovered bacterias that breaks down plastic
2. genomes in japan: gut biome have acquired genes that help them break down
seafood (marine bacteria), agliases

Question 5

In terms of HGT definition (mobilome extracellular DNA), What is a mobilome?

Answer 5

mobile genetic elements e.g. lysogenic phages, transposons, plasmids
- chromosomal rearrangements, chromosomal islands
extracellular DNA (eDNA)

Question 6

What are lysogenic phages?

Answer 6

phages that become incorporated into bacteria

Question 7

What are the 4 mechanisms of HGT

Answer 7

transformation, conjugation, transduction, vesiduction

Question 8

What is transformation?

Answer 8

when bacteria die their DNA is released into the environment for other bacterias to take up. it is taken up by recipient (incorporated into chromosome) by homologous recombination. recipient has to be “competent” (have ability to take up eDNA)

Question 9

How is competency achieved in labs?

Answer 9

cold shock to make membranes permeable

Question 10

How is transformation achieved in gram positive organisms?

Answer 10

• have membrane proteins that are specific for identifying extracellular dna
• bound to site, nuclease switched on to convert dsDNA to ssDNA
• ssDNA binds to diff proteins
• eclipse phase bc the ssDNA cant be seen by nucleases that breaks down free roaming DNA (covered and protected)
• rec-A = if there are any homology, the foreign strand incorporates and recombines into chromosome
  usually more competnet bc only one membrane

Question 11

Transformation in gram negative organism

Answer 11

• membrane proteins identifies extracellular DNA
• needs to have these signatures T tract (AT rich) to be able to bind to gram negative membrane proteins (gram negative is less competent usually)
• as the DNA gets pulled and tract, it enteres the periplasmic space
• translocated into cytoplasm
• Rec2 makes competency proteins and works together to replicate
• if there are any homology, the foreign strand incorporates and recombines into chromosome

Question 12

What is conjugation?

Answer 12

transfer of DNA mediated by conjugative plasmid in donor. requires cell to cell contact , cytoplasmic bridges passes plasmid from donor to recipient (mating). recipient can receive conjugative plasmid.

Question 13

Describe the conjugative plasmid in conjugation

Answer 13

• F plasmid (e. coli)
• tra genes allow the conjugation of the pills (F pills assembly and F plasmid transfer
• insertion sequences (IS) and Tn1000 = integration with chromosome
• oriV = origin of replication
• oriT = origin of transfer

Question 14

Conjugation step by step using F plasmid

Answer 14

• F positive cell = carries conjugative plasmid
• has to take place with F- cell
• pillus connects cytoplasm of 2 cells
• pillus retracts so they become closer
• F posoitive cell knicks the DNA of one strand and DNA gets transferred by rolling motion
• receipient cell starts replicating as info gets exchanged until a whole plasid is replicated

Question 15

Conjugation step by step using Hfr strain (integrated F plasmid)

Answer 15

• The plasmid can become integrated into the bacterial chromosome.
• F+ bacteria have insertion sequences that allow integration at different sites of the chromosome.
• oriT (origin of transfer)
• When the plasmid integrates into the cell, it is no longer called an F+ cell but becomes an Hfr strain.
• During conjugation, single-stranded DNA gets pulled into the recipient cell.
• The recipient does not become F+ because the complete plasmid is not incorporated.

Question 17

What are integrative conjugative elements

Answer 17

alternative mechanism (not conjugation machinery)
- ICEs are integrated into chromosome and is commonly through the type IV secretion syste,
- transferred like conjugative plasmids
- carries AMR, virulence, metabolic genes that enhance survival

Question 18

What is staphylococcus aureus pathogenicity islands (SaPIs)

Answer 18

• ICE like elements
• have integration and excision genes
• do not have machinery for transfer so uses prophage as helpers
• when stress initiates SOS response, phage DNA pops out and gets carried along with it in phage particles and enters recipient

Question 19

What is transduction

Answer 19

transfer of DNA via bacterial virus (bacteriophage- virus that affects bacteria)

Question 20

What are the 2 types of transduction

Answer 20

• generalised = carried out by phages that infect bacterial cell, destroys it and uses it as its own machinary (lyric/viorulent phages)
• specialised = phages that gets incorporated into bacterial chromosome (by lysogenic/temperate phages)

Question 21

What is generalised transduction?

Answer 21

any gene from donor chromosome -> transductant by lytic/virulent phage

Question 22

How does generalised transduction work?

Answer 22

• After transduction, the resulting cell is called a transductant.
• A bacterial cell is infected by a phage particle.
• The phage inserts its DNA into the bacterial cell and uses the bacterial machinery to make more copies of its genome and proteins needed for new phage particles.
• The host chromosome is broken into pieces.
• At the end of infection, new phage particles form and exit the cell.
• Occasionally, at low frequency, when these phage particles are being assembled, DNA pieces get packed into the phage heads along with viral DNA - bacterial DNA can get incorporated with viral DNA.
• This unusual phage is called a transducing particle (contains bacterial chromosomal DNA).
• While it remains infectious like a normal phage, it now carries bacterial DNA. When it infects a new bacterial cell and injects this DNA, the bacterial DNA can be replicated. However, since the viral DNA is incomplete, it cannot hijack the host’s machinery.

Question 23

What is specialised transduction and how does it work ?

Answer 23

• selected region of donor chromosome becomes transductant by lysogenic/temperate phage
• phage is integrated at specific site in chromosome
• phage replication is controlled by bacterial chromosome
• lamda phage
• when replicated along w bacterial chromosome, normally, phage gets circularised and goes out making new phages (normal infection)
• sometimes, bacterial chromosome gets incorporated to phage (not random it is specific)

Question 24

What is vesiduction?

Answer 24

vesicle driven HGT
- gram negative lets out vesicles filled with protein and linear DNA
- vesicles picked up and is incorporated through fusion

Question 25

What are biofilms

Answer 25

multicellular communities with members belonging to same/different groups adhered to a biotic/abiotic surface
- composed of extracellular polymeric substances - eDNA, proteins, polysaccharides, membrane vesicles

Question 26

Biofilms are ___ for HGT

Answer 26

hotspots

Question 27

How are biofilms formed?

Answer 27

• motile cells
• attachment
• microcolony
• environmental stresses -> mature biofilms

Question 28

Biofilms: eDNA

Answer 28

• actively secreted or released from eukaryotic and prokaryotic cells (autolysis or fratricide)
• role in pathogenicity, fitness, evolution
• nutrient source
• DNA recombination and repair
• role in structural integrity of biofilms
• resistance to host antimicrobial peptides
• accompanied by secreted nucleases

Question 29

How HGT is used in intestines?

Answer 29

• Main Mechanisms of HGT:
  
  1. Conjugation:
     
     • Transfer of plasmids carrying genes (e.g., antibiotic resistance) between bacteria via a pilus.
     • Enables intraspecies and interspecies gene exchange.
  2. Transformation:
     
     • Uptake of free DNA from the environment (e.g., plasmids released when a bacterium dies).
     • This process helps bacteria acquire new traits.
  3. Transduction:
     
     • Bacteriophages (viruses that infect bacteria) transfer genes between bacteria during infection.
     • Can activate prophages, causing bacterial lysis and spreading genetic material.
• Implications in Gut Microbiota:
  
  • Antibiotic resistance genescan spread rapidly in the gut.
  • Opportunistic pathogens can acquire resistance and escape the gut to infect other tissues or individuals.

Question 30

How is HGT present in soil

Answer 30

role of HGT in the rhizosphere and its impact on plant-microbe interactions and bacterial evolution.

• Root Exudates:
  
  • Plants release exudates (nutrients, signaling molecules) that influence microbial activity and gene transfer.
• Mechanisms of HGT in Soil:
  
  1. Conjugation:
     
     • Transfer of plasmids between bacteria, promoting traits like nutrient utilization and stress resistance.
  2. Transformation:
     
     • Uptake of free DNA containing beneficial genes (e.g., for plant growth promotion or bioremediation).
  3. Transduction:
     
     • Phages facilitate gene transfer among soil bacteria, contributing to genetic diversity.
• Other Factors Influencing HGT:
  
  • Moisture contentandwater flowaffect bacterial motility and interactions.
  • Biofilm formationaround roots enhances bacterial communication and stability.
  • Mobile genetic elements (MGEs):Insertions, transposons, and phage elements accelerate bacterial evolution.