Transcript Slide 1

BACTERIAL TYPING
Applications:
@ To identify the source of infection.
@ To distinguish infectious from non-infectious
organisms (i.e. a pathogenic or a commensal).
@ To distinguish relapse from reinfection cases.
Bacterial typing methods:
# Phenotypic methods
# Genotypic methods:
Phenotypic methods
1. Antimicrobial susceptibility typing according
to the susceptibility pattern
2. Biotyping: according to biochemical
reactions.
3. Bacteriocine Typing:
@ Bacteriocine is a protein produced by
certain bacterial species.
@ It is a toxin that inhibits the growth of
bacterial strains of the same family but not
of the same species secreting it.
@ This toxin is regarded as an antibiotic.
@ Common bacteriocines are:
# Colicine: secreted by Gram-negative bacteria:
e.g. E.coli & coliforms.
# Megacine: secreted by Gram-positive bacteria
e.g. B.megaterium.
@ Production is controlled genetically by a
plasmid factor.
Applications of bacteriocines:
@ They are used to type bacteria.
@ Treatment and diagnosis of cancer
@ Treatment of listeriosis.
@ Food preservation.
4. Pyocine Typing:
@ Pyocine is produces by some strains of
Pseudomonas; and used to type other
Pseudomonas.
Pyocine typing is done as follows:
@ Streak test strain across a tryptone soya
blood agar medium
@ Incubate at 37 °C overnight. Collect all the
crop by a sterile slide.
@ Expose the remaining growth on the plate to
chloroform for 15 minutes
@ Streak Ps. pyocyanea indicator strains at
right angles to test strain streak.
@ Re-incubate the plate at 37 °C overnight.
@ Read test by looking for zones of inhibitions.
5. Serological Typing:
@ Here patient serum is used to study the
antigen-antibody reaction
@ Antigens used are known, and disease is
diagnosed by identifying the antibody in serum
@ Different types of serological tests:
# Agglutination
# ELISA
# Immunoflurescence
# Precipitation
# C.F.T.
Agglutination test:
@ Agglutination occurs when the antigen unites
with the antibody and clumps together.
@ It will diagnose brucellosis, enteric fever, etc
@ Agglutination needs :
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Presence of salts, e.g. Na Cl
Neutral pH
Temperature between 37 °– 50 °C
Serum is heated (50°C) to destroy complement
Precipitation test:
@ A precipitation ring occurs between the
serum (antibody) and the fluid antigen.
Applications:
@ Lancefield typing
@ Plate precipitation test.
@ In immuno-electrophoresis
@ To diagnose infections of Strep., Neisseria,
B. anthracis, Haemophilus, pneumococci, etc.
Immunoflurescence :
@ Antigen unites with a fluorescent dye,
exposed to U.V. rays & examined under
fluorescence microscope to see a fluorescent
antigen.
ELISA:
@ Very popular, highly sensitive, more safe,
more cheap, and constant in its results.
C.F.T.:
@ First activate complement in serum by adding
Ca++ & Mg++ salts.
@ Positive CFT:
# Ag + specific Ab + C = complement fixation
# Fixed complement + RBC = no RBC haemolysis
Negative CFT:
# Ag + non specific Ab + C = no C fixation
# Non fixed complement + RBC = RBC
haemolysis
6. Phage Typing:
@ Phage typing is used to trace sources of
infections caused by: S.aureus, P.aeruginosa,
Salmonella species, B.abortus, M.tuberculosis
Staph. aureus phage typing:
@ 24 phages known to destroy S. aureus.
@ S. aureus is divided into > 100 phage types.
Procedure of phage typing:
1-Inoculate S. aureus on a nutrient agar plate
divided into 24 squares from outside
2-Use standard concentration of phages that
gives confluent areas of S. aureus killing.
3-Drop each phage in a separate square and
incubate at 37 °C overnight
4-Look for areas of no growth on each square,
i.e. complete destruction of S. aureus cells.
5-Record the phage number that made the
destruction.
6-This phage number is the S. aureus type.
BACTERIAL GENOTYPING
Applications:
@ To determine if bacterial isolates are
genetically related.
@ To trace the source of infection outbreaks
@ To determine multidrug-resistant pathogens.
@ GENOTYPIC METHODS INCLUDE:
1.
2.
3.
4.
Pulsed-field gel electrophoresis typing
PCR typing
Multilocus sequence typing
Plasmid analysis.
1. Pulsed-field gel electrophoresis typing:
@ It is similar to standard gel electrophoresis
except that instead of running the current
in one direction, it is run in 3 directions.
@ It is applied for genotyping and tracing
sources of infections.
2. PCR typing:
Application:
# Identification of slow-growing organisms,
# Early diagnosis of infectious diseases
# Diagnosis of hereditary diseases
# Identification of fingerprints and paternity
3. Multilocus sequence typing:
@ It is used to differentiate and type
many isolates, e.g. Campylobacter,
N. meningiditis, S. aureus, S. pyogenes,
C. albicans
4. Plasmid typing:
@ A plasmid is a small DNA molecule that is
separate from cell chromosomal DNA
@ Plasmids carry genes of antibiotic
resistance and can be transmitted from one
bacterium to another.
@ Plasmids are extracted by PCR and used to
differentiate and type bacteria.