LAB - Colonial & Gram Morphology, Culture Media Flashcards
where we can see a visual difference between different species
differential medium
Inhibitory reagents such as dyes, chemicals or antimicrobials are added to make the medium …
selective
This can be added for anaerobic medium
A reducing agent (e.g. cysteine or thioglycollate)
“nutrient” broth
a simple fluid media designed to support the growth of most bacteria
with non-fastidious growth requirements. The constituents of the broth are peptone, beef extract, sodium chloride and distilled water
what makes a nutrient broth a solid medium?
the addition of a complex polysaccharide AGAR
- extracted from seaweed, acting as a source of calcium, but it is not utilized by most bacteria
BAP
- enriched medium, blood agar is prepared by adding sterile, defibrinated sheep blood to a sterile, molten nutrient agar base (TSB) cooled to approximately 50°C
- also considered a non-selective media,
- as well as differential, as it is used to differentiate different types of hemolysis
- to maximize recovery of fastidious bacteria, blood agar bases can be enriched by the addition of yeast extract
- vitamin K, hemin and cysteine (reducing agent) supplements may be added for isolation of strict anaerobes
CHOC
- chocolate agar
- BAP that has been heated before plates are poured (BROWN)
- enriched & non-selective
- used for culture of fastidious organisms (such as Haemophilus & gonococci)
- growth is enhanced if supplemented w yeast extract (V factor + X factor); sheep blood inactivates NAD - will not support growth of Haemophilus
Alternative method of supplying NAD on unheated blood agar
cross-streaking inoculated cultures with Staphylococcus aureus (“Staph. streak”)
- staphylococci lyse RBCs (beta hemolysis) and release NAD into the medium, allowing the Haemophilus to grow as “satellitic” colonies in the area adjacent to the staphylococci
Colisin-Nalidixic Acid Agar (CNA)
selective medium used for the recovery of aerobic
and anaerobic Gram positive bacteria
- basically a BAP with antibiotics colisin and nalidixic acid, which inhibit Gram-negative bacteria
MAC
- selective-differential medium; selective as it inhibits growth of gram-positive bacteria but allows growth of the Enterobacteriaceae and the non-fermentative gram-negative rods
- differentiates between “lactose fermenters” (pink colonies) and “non-lactose fermenters” (white or colorless colonies)
this media exist for the selective and differential isolation of specific bacteria and yeasts
Chromogenic media
grows best at cold temperatures (10-20°C)
psychrophile
mesophile
– grows best at moderate temperatures (20-40°C)
grows best at high temperatures (50-60°C)
thermophile
obligate aerobes
require oxygen (at least 18% O2)
grows with or without oxygen
facultative anaerobes
capnophilic
grows better when atmosphere enriched with CO2 (5-10%, CO2, ~18% O2)
require a reduced level of oxygen (5-6% O2)
microaerophilic
humidophilic
require increased humidity (70-80%)
Incomplete haemolysis of red blood cells with partial breakdown of haemoglobin.
alpha-hemolysis
- visible as a zone of partial clearing with green discoloration surrounding colonies
beta-hemolysis
Complete lysis of RBC with complete breakdown of haemoglobin. There are distinct, totally clear zones around colonies
There is no zone of hemolysis surrounding colonies, i.e., the bacteria are nonhemolytic. Normally this is not recorded as a significant finding.
gamma-hemolysis
gv
gram variable
pleo
pleomorphic
basal nutrient media
designed to support growth of most organisms (non-fastidious growth requirements)
- various sugars + indicator added to make it differential
- inhibitory reagents such as dyes, chemicals, or antimicrobials added to make selective
- may be enriched with blood, serum, yeast for growth of fastidious bacteria
- reducing agent (eg. cysteine or thioglycollate) for anaerobic medium
“nutrient” media
- designed to support growth of most organisms (non-fastidious)
- peptone, beef extract, sodium chloride, and distilled water
- addition of complex polysaccharide AGAR, it becomes solid or semi-solid (from seaweed); acts as a source of calcium but not used by most bacteria
Sheep Blood Agar (BAP)
- enriched medium
- sterile, defibrinated sheep blood to a sterile, molten nutrient agar base (TSB) cooled to approximately 50 C (5% v/v)
- non-selective media, as well as differential = differentiate different types of hemolysis
- maximize recovery of fastidious bacteria = add yeast extract
- reducing agents: vit K, hemin, and cysteine to isolate strict anaerobes
Chocolate agar (CA/CHOC/CAP)
- BAP that has been heated before plates are poured
- enriched and non-selective
- used for culture of fastidious organisms (Haemophilus and gonococci)
- growth enhanced w yeast
- X factor (hemin) and V factor (NAD)
- sheep enzyme has an enzyme that inactivares NAD so unheated sheep blood will not support the growth of H. influenzae (requires NAD)
Alternative method of supplying NAD on unheated blood agar
is by cross-streaking inoculated cultures with S. aureus (“staph streak”
- staph lyse RBCs = beta
and release NAD into medium
= Haemophilus grow as satellitic colonies in area adjacent to staph
these media can include antibiotics and can be used to isolate drug-resistant bacteria
Chromogenic media
- MRSA, VRE, etc.
key features of bacterial colonies that serve as important criteria for identification
- size
- shape
- elevation
- pigmentation
- surface texture
- opacity
these are used to detect single colonies from mixed cultures in order to obtain pure cultures
sterile wires
NOT loops
colonies are examined for these characteristics
- surrounding zone of hemolysis
- surface morphology including colour, opacity, and size
