MICROBIAL GENETICS

1
1
MICROBIAL GENETICS PLASMIDS OTHER MOBILE GENETIC
ELEMENTS
2
2
MICROBIAL GENETICS - PLASMIDS / OTHER MOBILE
GENETIC ELEMENTS
PLASMIDS Characteristics, Resistance Factors,
Resistance Transfer Factors Drug Resistance
Mechanisms Penicillin Resistance, Penicillinase,
Beta-lactamase Multiple Drug Resistance Transfor
mation by Plasmids OTHER MOBILE GENETIC
ELEMENTS Insertion Sequences Transposons Integ
rons Superintegrons Conjugative
Transposons Genomic Islands The Busy Genome
3
3
PLASMIDS SMALL, CIRCULAR, DOUBLE-STRAND
DNA MOLECULES 5 - 50
GENES, CYTOPLASMIC LOCATION, NOT
ESSENTIAL (NORMALLY) REPLICATION GENES AND
SITES 1 - 20 COPIES EACH / CELL SEVERAL
DIFFERENT PLASMIDS / CELL RESISTANCE FACTORS -
PLASMIDS WHICH CARRY GENES WHICH ENCODE PROTEINS
WHICH MAKE THE BACTERIAL HOST RESISTANT TO
ANTIBIOTIC - CALLED DRUG RESISTANCE RESISTANCE
TRANSFER FACTORS - ALL ABOVE PLUS ABILITY TO
TRANSFER PLASMID IN MATING CONJUGATION OTHER
PLASMID GENES HYDROCARBON CATABOLISM TOXIN
PRODUCTION MINERAL UPTAKE
4
4
MECHANISMS OF DRUG RESISTANCE

• MUTATION RESULTS IN ALTERED BACTERIAL PROTEIN.
• IT NO LONGER RECOGNIZES ANTIBIOTIC BUT
• CONTINUES TO PERFORM NORMAL FUNCTION
• IN BACTERIAL GROWTH
• EX STREPTOMYCIN
• BACTERIA ACQUIRE NEW GENE WHICH CODES FOR
• ENZYME WHICH DESTROYS ANTIBIOTIC
• EX PENICILLINASE DESTROYS PENICILLIN
• BACTERIA ACQUIRE NEW GENE WHICH CODES FOR
• ENZYME WHICH PUMPS ANTIBIOTIC BACK OUTSIDE
• CELL
• EX TETRACYCLINE RESISTANCE

5
5
PENICILLIN CLEAVAGE INACTIVATION BY
PENICILLINASE
b - LACTAM RING
PENICILLINASE b - LACTAMASE
INACTIVE
6
6
MULTIPLE DRUG RESISTANCE JAPAN, 1957, SHIGELLA
DYSENTERIAE, DYSENTERY RESISTANT
TO SULFONAMIDES, STREPTOMYCIN, CHLORA
MPHENICOL, NEOMYCIN JAPAN 1957 - 1
CASE 1964 - 50 LONDON 1962 - 3 1964
- 61 NEISSERIA GONORRHOEAE 1945
PENICILLIN-SENSITIVE 1975 PENICILLIN-RESISTANT
, BUT SPECTINOMYCIN-SENSITIVE 1985 -
1990 STREPTOMYCIN-RESISTANT, CEFTRIAXONE-SENSIT
IVE
7
8
BACTERIUM RESISTANT TO PENICILLIN
PENICILLINASE
GENE FOR PENICILLINASE b - LACTAMASE
PLASMID
PHENOTYPE ORGANISM GROWS ON MEDIUM CONTAINING
PENICILLIN
8
9
TRANSFORMATION BY PLASMIDS
PENICILLIN RESISTANCE PLASMID
PENICILLIN - SENSITIVE CELLS
MIX, SOME PLASMIDS ENTER CELLS
PENICILLIN - RESISTANT TRANSFORMANT
SELECT PLATE ON MEDIUM AND PENICILLIN PLASMID
ONLY NO COLONIES RECIPIENT CELLS ONLY
NO COLONIES RECIPIENT CELLS AND PLASMIDS

COLONIES PENICILLIN - R
9
10
PEN - R TRANSFORMANTS
MEDIUM AND PENICILLIN
10
11
OTHER MOBILE DNA
11
12
TRANSPOSONS 5 - 10 KB, LINEAR TRANSPOSE TO
MULTIPLE SITES CARRY ANTIBIOTIC (DRUG)
RESISTANCE GENES BRACKETED BY 2 INVERTED
INSERTION SEQUENCES TRANSFERS DRUG RESISTANCE
12
12
RESISTANCE GENES ON TRANSPOSONS
KANAMYCIN NEOMYCIN
Tn5
STREPTOMYCIN
IS1
SULFONAMIDE
TETRACYCLINE
Tn3 PENICILLIN
Tn10
IS3
IS3
Tn4
IS2
NOTE SEVERAL INSERTION SEQUENCES
IS1
TRANSFER GENES
REPLICATION GENES AND SITES
PLASMID R1 100,000 BP
13
(No Transcript)
14
(No Transcript)
15
(No Transcript)
16
(No Transcript)
17
(No Transcript)
18
(No Transcript)
19
(No Transcript)
20
(No Transcript)
21
13
INTEGRONS - GENE CAPTURE ELEMENTS

• CODE FOR INFORMATION NECESSARY TO INSERT HOST
  GENE INTO ITSELF
• INTEGRASE TO MOVE GENE
• ATTACHMENT SITE TO PUT THE GENE INTO
• HAVE A PROMOTER TO EXPRESS CAPTURED GENE
• ACQUIRE VARIOUS HOST GENES
• EXIST IN MANY FORMS - DIFFER IN NUMBER AND
  IDENTITY OF CAPTURED GENES

22
14

• INTEGRON

• INTEGRON WITH INSERTED HOST GENE

• INTEGRON WITH MULTIPLE INSERTIONS OF HOST GENES

23
15

• SUPER INTEGRON - VIBRIO CHOLERAE CHROMOSOME 179
  INSERTED GENES
• HOW ARE INTEGRONS INVOLVED IN MOBILITY OF DNA?
• BECAUSE INTEGRONS CAN BE LOCATED ON TRANSPOSONS
  AND ON CONJUGATIVE PLASMIDS, WHEN THEY
  CATALYZE MOVEMENT OF HOST GENES INTO
  THEMSELVES, THE OVERALL RESULT IS MOVEMENT OF
  GENES INTO TRANSPOSONS AND CONJUGATIVE PLASMIDS

24
16
CONJUGATIVE TRANSPOSONS

• MOVE FROM FIRST HOST (DONOR) GENOME
  SECOND HOST (RECIPIENT) GENOME
• CELL-CELL CONTACT
• SOME MOVE TO NEW SITES
• SOME MOVE TO IDENTICAL SITES
• 20 - 100 KB WITH GENES FOR
• INTEGRATION AND EXCISION
• CONJUGATION
• ANTIBIOTIC RESISTANCE
• TRANSFER VIA CIRCULAR INTERMEDIATE AND THEN
  INTEGRATE

25
17
TRANSFER ORIGIN oriT
DRUG RESISTANCE
TRANSFER OPERON
INTEGRASE
EXCISIONASE
oriT
Tn916
TET-R
EXCISION (IN DONOR CELL)

• SINGLE STRAND TRANSFERRED TO RECIPIENT
• CIRCULARIZES IN RECIPIENT
• COPIED INTO DOUBLE STRAND
• INTEGRATES INTO RECIPIENT CHROMOSOME

oriT
26
18
GENOMIC ISLANDS
LINEAR, 10-200 KB, IN CHROMOSOMES INTEGRATION /
TRANSFER ENZYMES INTEGRATE IN 3' ENDS OF tRNA
GENES (SIMILAR IN SEQUENCE TO PROPHAGE
ATTACHMENT SITES) BOUNDARIES ARE DIRECT REPEATING
SEQUENCES DIFFER FROM HOST CHROMOSOME IN G AND C
CONTENT OF DNA - INDICATES ORIGINATION IN
ANOTHER GENUS PERMIT EVOLUTION IN QUANTUM LEAPS
tRNA GENE
GENOMIC ISLAND
DIRECT REPEAT
INTEGRATION GENE
DIRECT REPEAT (SEQUENCE)
27
19
BRING IN GENES WHICH HELP SURVIVAL (OF BACTERIA)
28
20
UROPATHOGENIC E. COLI
MOST COMMON CAUSE OF BLADDER AND KIDNEY
INFECTIONS E. COLI (06 K15 H31) FOUR
PATHOGENICITY ISLANDS
29
21
THE BUSY GENOME ELEMENTS OF HORIZONTAL EXCHANGE
GENOMIC ISLANDS
e.g. ESCHERICHIA COLI COMMON 4.1 MB K12 ISLANDS
0.53 MB 0157H7 ISLANDS 1.34 MB
PROPHAGES
CONJUGATIVE TRANSPOSONS (gram ve)
MINIMAL SPECIES GENOMIC BACKBONE
SUPER INTEGRONS (MAINLY g PROTOBACTERIA)
INSERTION SEQUENCES
INTEGRONS
TRANSPOSONS