bacteria identification : gram positive cocci. bile-esculin enterococcus identification –esculin ...
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Bacteria identification :Gram positive cocci
Bile-Esculin
• Enterococcus identification– Esculin Esculitin + Glucose– esculitin + Fe Ferric citrate (dark brown)
• Selective media– Bile added to inhibit gram + bacteria– Enerococci can survive
Bile-Esculin
- +
Novobiocine Sensitivity
• Kirby Bauer Test• Discrimanates S. saprophyticus from other
staphylococci• S. saprophyticus is the only resistant
staphylococci
Mannitol + Salt Agar
• High salinity (7.5%) : enriches staphylococcus• Indicator : Phenol Red• Carbon Sources : Mannitol and Proteins• Detects Mannitol Fermention
– Mannitol acid formation
Mannitol + Salt Agar
A) Positive for mannitol fermentationB) Negative for mannitol fermentationC) Positive for mannitol fermentationD) No growth
Tellurite/Baird Parker Agar
• Selective Media:– Lithium chloride– 1% Pottasium Tellurite Solution
• Differential Media:– Egg Yolk : lecithinase (clearing)– Pottasium Tellurite : coagulase-positive
(blackening)
Tellurite/Baird Parker Agar
PYR Test
• Detected Enzyme : Pyrrolidonyl peptidase– L-pyrrolidonyl-β-napthylamide (PYR) L-
pyrrolidone carboxylic acid + β-napthylamine• Detection of reaction:
– β-napthylamine + p-dimethyl-aminocinnamaldehyde pink precipitate
PYR Test
- +
Cellular Aggregation of Gram Positive Cocci
Micrococcus & Streptococcus
Streptococcus
Micrococcus
Staphylococcus
-Aggregation can be used for distinguishingbetween genera
Diagnostic of Medically important Gram Negative Bacteria
Using the Identification Flow Chart
http://mysite.science.uottawa.ca/jbasso/microlab/IDFlowcharts.pdf
Diagnostics : Gram Positive Cocci
Gram positive cocci
Streptococcaceae
Catalase -
Aerobes & facultative anaerobes
Streptococcus
Micrococcaceae
Catalase +Aerobes
Micrococcus
Staphylococcus
Characteristics of Gram Positive Cocci
• All are non sporulating• Mainly found amongst the
natural flora of humans and animals
• Fastidious (‘picky’) nutritional requirements– Use simple carbon sources
Gram Positive Cocci of Medical Importance
• Micrococcaceae– Staphylococcus aureus
• Causes several types of infections, food infections and toxic shock (skin and respiratory tract)
– Staphylococcus epidermidis• Cause opportunistic infections (catheters with biofilms)
– Staphylococcus saprophyticus• Major cause of cystitis in women (bladder infection)
Gram Positive Cocci of Medical Importance
• Streptococcaceae– Streptococcus pyogenes
• Strep throat and flesh eating disease– Streptococcus agalactiae
• Genital infections– Streptococcus mutans
• Endocarditis– Streptococcus pneumonia
• Otitis, meningitis, and pneumonia– Enterococcus spp.
• Opportunistic infections
Diagnostics : Gram Positive Rods
Gram positive rods
Spore formers
Aerobes & facultative
aerobesBacillus
Strict anaerobes Clostridium
Non spore formers Aerobes Listeria
Medically Important Bacilli
• Bacillus– Mostly harmless– A few opportunistic species
• Bacillus cereus & Bacillus subtilis– Food poisoning
– One pathogenic species• Bacillus anthracis
– Anthrax Gram positive
rods
Spore formers
Aerobes & facultative
aerobesBacillus
Strict anaerobes Clostridium
Non spore formers Aerobes Listeria
Medically Important Bacilli
• Clostridium– Several pathogenic species
• Clostridium perfringens– Gas gangrene
• Clostridium tetani– Tetanus
• Clostridium botulinum– Botulism
• Clostridium difficile– Diarrhea
Canned food that has not been sterilized properly; paralytic illness
prolonged contraction of skeletal muscle fibers, neurotoxin produced by the bacteria
Diagnostics : Gram Negative Bacteria
Oxydase
Pseudomonaceae
NeisseriaceaeFermentation
of glucose
Neisseriaceae
Pseudomonaceae
EnterobacteriaceaeFermentation
of lactose
McConkey
Enterobacter
Escherichia
Klebsiella
Serratia
Proteus
Salmonella
Shigella
Morganella
Representative Gram Negative Bacteria
• Gram-negative Cocci– Neisseria gonorrhoea– Neisseria meningitis
• Gram-negative rods– Enterobacteriaceae family
• Escherichia, Enterobacter, Salmonella, Shigella, Klebsiella, Proteus, Morganella
– Pseudomonaceae family• Pseudomonas
Immunology
Immunology
• Purpose of the immune system:– Discriminate self from non-self
• Non-self –Antigens
• Immunity:– All mechanisms used by the host to protect itself
and fight non-self
Non-Self - Antigens• Anything that can react with the participants of
the immune system– Ex. antibodies
• Epitope: Characteristic of the antigen which allows its recognition as being non-self– Ex. Lipids, proteins, lipopolysaccharides
The Antigen
Virus=Antigen
Epitopes
Immunological Diagnostic Methods
• Determine the presence of an antigen:• An organism• A protein• A toxin• An antibody
– ELISA method to determine quantity– Immunochromatography (Rapid tests)
ELISA
• Used to detect the presence of antigens or antibodies– High sensitivity– Quantitative
ELISAMethod
ENZ ENZENZ ENZ ENZ
ENZENZ
ENZ ENZ ENZ
Serum (source of Ag) is added to plastic wells
Antigen Present Antigen Absent
Blocking agent addedAb against Ag is addedWashDetecting Ab addedWashSubstrate added
Interpretation of Results• Serums of patients tested for the presence of HIV
– 1o Ab anti-HIV from mouse– 2o Ab anti-Ab of mouse
• Conclusions– Patients 1 & 3 are positive for HIV– Patient 3 has a higher titer– Patient 2 is negative for HIV
1/2 1/4 1/8 1/16 1/32 1/64 1/128Control serums + —
1/8
Next Week
• Final Quiz• Practical Exam:
– Determine which partner will complete the exam fist
– Make sure everything on the exam is clear, you can ask any question concerning the methods required for the practical exam