2nd year Medical Students - JU2 year Medical Students - JU Bacterial genetics

Dr. Hamed Al ZoubiAssociate Professor of Medical Microbiology.

MBBS / J.U.S.T M S / J U SMSc, PhD/ UK

B t i l tiBacterial genetics

ILOs:

• bacterial genome and replication

ff• Mutations and effects on bacteria

• Genetic exchange and role in virulence• Genetic exchange and role in virulence

• Transposons definition and significance p g

The bacterial genome:g

• Chromosome ( a must)• Chromosome ( a must)

Pl id• Plasmids

• Bacteriophage

• Transposons

Bacterial Genetics

Significance of bacterial genetics?Significance of bacterial genetics?

St t P th i it d i lStructure, Pathogenicity and virulence Antibiotic resistanceToxinsEnzymesyMolecular cloning

Bacterial genetics

• Bacterial Genome: Chromosome, single circular double-stranded DNAcircular double-stranded DNA.

•1300 um long contains 2-5 x 106

l tid bnucleotide bases

• enough DNA to encode 1- 3 gthousand different genes. According to bacteria types.

•Genes are expressed as required

•Gene expression is controlled by many factors such as amount of end product metabolism presenceend product metabolism, presence of repressors (operators inducer binding)

Bacterial Genetics

E.coli:

• Single circular chromosome• 5 106 base pairs5 10 base pairs• Codes for nearly 2000 proteins

A human cell:

• 3 X 109 base pairs • 100, 000 proteins

G t / Wild T R t ll• Genotype / Wild Type : Represents allpotential genes of bacteria cell.. Its genome All Inherited essential biologicalgenome.. All Inherited essential biological features & growth patterns.

• Phenotype: The expressed genes..The observed characteristics of the of theobserved characteristics of the of the individual bacteria species/strain. Expressed by physical & biochemicalExpressed by physical & biochemical properties. Growth patterns, Fermentation products, Antibiotic resistance, Toxinsproducts, Antibiotic resistance, Toxins production. .etc.

1 M t ti•1. Mutation:• inaccuracies during DNA replication lead to change in the

base sequence of DNA that may result in insertion of amino acid into a protein and phenotypic alterationSpontaneous and induced• Spontaneous and induced

• Spontaneous:Spontaneous:

a. Happens every 104-109 cell division (1 colony isa. Happens every 10 10 cell division (1 colony is 1010)

b. happen all of the time, regardless of growth conditions: Spontaneous, pressure..co d t o s Spo ta eous, p essu e

• Induced mutations

•Chemicals, X-rays, UV light Viruses.., y , gDirect damage of nucleotidesAlteraion of the nucleotide bonds

•Mutations outcome:

No effect / silent

Change in protein structure such as an antibiotic g ptarget causing resistance to antibiotics

change or inhibition of a functional protein synthesis which can be lethal to bacteriay

Types of mutationsTypes of mutations• Multisite: many nucleotides

Inversion, duplication or deletion

• Point mutation: single or few bases• Point mutation: single or few bases

Insertion, substitution or deletion

1 base substitution:1. base substitution:

• DNA polymerase error or due to mutagens • Outcome: Missense vs nonsense mutations or silent

2 Frame shift mutation:2. Frame shift mutation:

• One or more base are added or deleted• One or more base are added or deleted

• Shift in the reading frameg

• Corrupting the reading codons downstream mutationsl di t i ti t ileading to inactive protein

• 2. Genetic exchange:

Importance:i tibi ti i ta. moving antibiotic resistance genes among

bacteriab i i l b t ib. moving virulence gene among bacteriac. changing the antigenic make-up to avoid immunity

Mechanisms

1. Transformation - uptake of naked DNA2 T d ti b t i h t2. Transduction - bacteriophage as vectors3. Conjugation - plasmids moved by cell-cell

t tcontact4. Transposons

• 1. Transformationa. recipient cell must be competentp pb.Naked DNA will be uptakenc. natural competence versus artificial competenced l t i b t i t ll t f bld. only certain bacteria are naturally transformable e.g

Streptococcus pneumoniae, Haemophilus influenzae, Neisseria gonorrhoeae VibrioNeisseria gonorrhoeae, Vibrio

What is transfection?

2. Transduction

a. bacteriophage (phage) are viruses of bacteria - can be either lytic or temperate (Lysogenic)y ( y g )i. lytic - always lyse (kill) host bacterial cellii. temperate - can stably infect and coexist within bacterial cell (lysogeny) until a lytic phase is induced

b. lysogenyi. the phage genome during lysogeny is called the prophage, and the bacterial cell is called a lysogenii. if the phage genome encodes an observable function, the lysogen will be altered in its phenotype -lysogenic conversion (e.g.,cholera toxin, group A streptococcus, botulinum toxin diphtheria toxin in Corynebacterium diphtheriae)

Lytic: replicate and bursty pLysogenic: prophage <> lytic

Diphtheria, Cholera, botulinum and erythrogenic toxins

A. specialized transduction (in temperate bacteriophages)bacteriophages)

i some prophages integrate into the bacterial genomei. some prophages integrate into the bacterial genome at a specific location

ii. when a prophage is induced to lytic phase, it may drag along a piece of the bacterial genome next todrag along a piece of the bacterial genome next to the integration site and move that bacterial sequence into the new recipient host cell changing theinto the new recipient host cell, changing the recipient's genome

Usually non medically significant

B. generalized transduction (LYTIC)

• bacterial chromosome fragments when transduction occur > phage packages phage p g p g p ggenome into the capsid.

• BUT sometimes the bacteriophage capsid is accidentally filled with random pieces of bacterial y pgenome, possibly including plasmids

• when the capsid injects the host genes into a new recipient, the new gene can recombine into the p grecipient genome and cause a change

iv. toxins and antibiotic resistance genes can be moved by generalized transduction

C. Conjugation (mediated by plasmids)

What is a plasmid?A small DNA molecule that is physically separate fromA small DNA molecule that is physically separate from,

and can replicate independently of bacterial chromosome / circular double strandedchromosome / circular double stranded

Three possible states for plasmid conjugationThree possible states for plasmid conjugationi. conjugative - the plasmid encodes all of the functions for conjugation and can move itself from thefunctions for conjugation and can move itself from the donor cell to the recipient cellii mobilizable - the plasmid cannot move itself butii. mobilizable - the plasmid cannot move itself, but can be moved with help from a conjugative plasmidiii non-transmissible - can't move by conjugationiii. non-transmissible - can t move by conjugation

Conjugation steps in generali. synthesis of sex pilusy pii. cell to cell contact via pilusiii. copying plasmid DNA and transfer of copy into py g p pyrecipient cell

bacteria containing a conjugative plasmid are called donor, male, (F+), , ( )

bacteria receiving the plasmid are called g precipient, female, (F-)

importance of conjugation - moving plasmids encoding multiple antibiotic resistance genes (R g p g (plasmids) among diverse bacterial

Types of conjugation;Types of conjugation;1 . F+ conjugation

G ti bi ti i hi h th i t f f FGenetic recombination in which there is a transfer of an F+ plasmid but not chromosomal DNA from a male donor bacterium to a female recipient bacterium Other plasmidsbacterium to a female recipient bacterium. Other plasmids present in the cytoplasm of the bacterium, such as those coding for antibiotic resistance, may also be transferred g , yduring this process.

2 . High frequency recombination (Hfr)

Genetic recombination in which fragments of chromosomal DNA from a male donor bacterium are transferred to a female recipient bacterium following insertion of an F+ l id i t th l id f th d b t iplasmid into the nucleoid of the donor bacterium.

terms of interest:terms of interest:O i ll th F f t i t t i t d iti i• Occasionally, the F factor integrates into a random position in the bacterial chromosome. When this happens, the bacterial cell is called Hfr instead of F+cell is called Hfr instead of F+.

• Hfr bacteria are still able to initiate conjugation with F- cells, j gbut the outcome is completely different from conjugation involving F+ bacteria. IT IS called an F’ instead

F' l id i t t i t th h i iF' - plasmid integrates into the chromosome, excision drags along piece of the chromosome. F' can move h t DNA b t ll ( i il t i li dhost DNA between cells (similar to specialized transduction)

3. Transposons:

• Jumping genes/copy and paste (Class1) or cut and paste (2)p g g py p ( ) p ( )

• Between plasmids or between chromosomes and plasmids

• medical importance since many antibiotic resistancemedical importance since many antibiotic resistance genes are encoded by transposons in antibiotic resistance plasmids

Th E d•The End