MMSC 428 Exam 4 Study Guide |Medical and Molecular
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Who am I? 1. Anaerobes
- I have special nutritional
requirements for vitamin K,
hemin, and yeast extract.
- My culture specimens
should be inoculated
onto one of these
primary isolation agar
plates.
1. Supports growth of almost all obligate anaerobes
Anaerobes: CDC ABAP &
Brucella agar and provides sheep RBCs, vitamin K, and yeast
extract
1. RBCs lysed through a series of freeze-thaw cycles.
Lysis releases more nutrients into the agar.
Anaerobes: Laked sheep 2. Selects for anaerobic GNB. Kanamycin inhibits most
blood (LSB)
facultative GNB & vancomycin inhibits most GP
organisms. Yeast and kanamycin-resistant GNB may also
grow "breakthrough"
T/F: Broths serve as a False, SECONDARY, PRIMARY, ALL
PRIMARY culture source. If
the SECONDARY agar plates
do not recover suspected
anaerobes, the broth can
be subcultured to anaerobic
plates for recovery. Broths
should be inoculated for
SOME body fluids and
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tissues.
1. Blood-based
a) Laked blood agar w/ kanamycin & Vancomycin (LKV)
Anaerobes: Selective Media b) Phenylethyl alcohol (PEA)
4. Non blood based
a) Bacteroides bile esculin agar (BBE)
1. Selects for anaerobic GP organisms and some
Anaerobes: Phenylethyl
alcohol (PEA) anaerobic GNB. Primarily suppresses
Enterobacterales and inhibits swarming by
Proteus spp.
1. Selects for Bacteroides spp. Contains gentamicin to
Anaerobes: Bacteroides bile inhibit aerobic organisms, 20% bile to inhibit most
esculin agar (BBE) other anaerobes, and esculin for differentiation.
2. BBE can be used to rapidly identify members of
the B. fragilis group (most common clinically isolated
anaerobes): growth in 20% bile and esculin
hydrolysis
Anaerobes: Optimal 1. Nonselectiveblood agar plate, PEA, LKV, & BBE.
Anaerobes Culture Setup 2. At minimum should include: Nonselective blood agar plate,
LKV, & BBE.
1. Room temperature plates (Agar plates should be
allowed to warm to room temperature before
specimen inoculation.)
Anaerobes: Inoculation
2. Holding jar, box, small chamber
- Ideally each culture should be inoculated and
incubated immediately. However, in laboratories
without the necessary resources a holding jar, box, or
small chamber can be used to batch inoculated
anaerobic cultures before incubation.
1. Anaerobic environment
2. Anaerobic jars and bags
3. 35-37C for 48 hours. Jars can hold 8-12 plates, bags can hold 1-4
plates.
Anaerobes: Incubation
5. Jars and bags can utilize a gas generating tube or
sachet containing CO2 and H2. The H2 binds oxygen
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inside to form water vapor.
6. Waterless gas generators absorb oxygen from the
air. These generally take 30 minutes-2 hours to
reach anaerobiosis.
1. Expensive and require gas
tanks. Best for large volume
Anaerobic Chamber
bacteriology labs.
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1. Gas evacuation system that creates anaerobiosis within 3
minutes.
Anaerobes: Anoxomat™ 2. Pros: Rapid anaerobic environment, reusable
system
components without consumables, mature anaerobic
colonies faster
3. Cons: Expensive, requires gas tanks
1. Slow growers compared to aerobes
2. Growth on anaerobic culture media
3. Aerotolerance test—aerobic and anaerobic cultures
Anaerobes: ID
4. Gram stain
5. Biochemical/enzyme testing systems/panels
6. MALDI & Sequencing
T/F: Aerotolerance can be True
performed with primary
isolation media or with
subcultured isolates
1. Gram-positive bacilli
2. Spores
3. Obligate anaerobes
4. Habitat: Some found as members of the normal
Clostridium/Clostridioides
microbiota of the GI tract of humans and animals.
Some species are also found in soil.
5. Infection can result from opportunistic endogenous
species, or exogenous species via ingestion of
organisms or their toxins, and from contamination of
wounds with soil containing the organisms.
T/F: Clostridium/Clostridioides False, >200
has <200 named species
1. Member of histotoxic or gas gangrene group
Clostridium/Clostridioides: C.
perfringens 2. Five different strains or types based on exotoxins
1. α-toxin, lecithinase (phospholipase C): catalyze the
splitting of host phospholipids (in cell membranes)
C. perfringens: Virulence 2. β-toxin: forms cation-selective pores in cell membranes
Mechanisms causing cell lysis
3. Perfringolysin O: Pore forming cytolysin
4. Enterotoxins
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