Download REVIEW: Lab Quiz #2 Wed. April 12

REVIEW: Lab Quiz #2 Wed. April 12th
Lab 21 Anaerobes etc. For each of the types of oxygen utilization (obligate anaerobes, obligate aerobes,
indifferent/aerotolerant, facultative anaerobes) understand: the type of metabolism performed; relative
growth in various environments (atmosphere 16-20%/candle jar 6-7%/anaerobe jar 0%). Thioglycollate
& pyrogallol: reducing agents which chemically decrease free oxygen content in media. Thioglycollate:
semisolid broth; gradient of oxygen from highest at top to lowest at bottom. Pyrogallol: itself is toxic,
must be applied to cotton plug; acts when in presence of a strong base. Agar slant below is kept in
anaerobic environment.
Lab 22. Define selective & differential.
EMB, MacConkeys: define what type of bacteria are selected for/selected against (Gram --/Gram
+); differential for lactose fermentation. Acid, ppt Eosin: red/pink; strong: Eosin + methylene blue:
metallic green. Azide agar poisons cytochromes; indifferent bacteria OK. Facultative anaerobes: growth
on azide is variable, depends on whether they survive the transition to fermentation only.
Lab 23. Antibiotic sensitivity can be determined by disk diffusion method. Measure DIAMETER of
zone of inhibition (absolutely no growth), consult a table of values (don’t directly compare the sizes of
two different zones). Size of zone is affected by rate of diffusion of the antibiotic, and the concentration,
as well as bacterial sensitivity/resistance.
Lab 24 UV light. What species of bacteria is very sensitive to UV damage, and grows red colonies?
Serratia maracescens. UV light kills cells by damaging DNA (it can also cause cancer by this
mechanism). Two kinds of structural damage: DNA strand breaks; formation of pyrimidine dimers,
especially between adjacent thymines (T’s). Thymidine dimers must be repaired, or the DNA cannot be
transcribed or replicated. Some forms of repair result in excision of the dimers and deletion of the bases.
UV damage is dose-dependent: the longer the exposure, the more damage. UV light can be used to
sterilize surfaces of surgical suites.
Lab 25/26. Normal flora. Skin: mainly Gram + cocci (Staphylococus epidermidis, Staphylococcus
aureus, Micrococcus). Know all the selective/differential media used: Blood agar (alpha hemolysis:
partial, appears as green/brown discoloration in the media around the colonies. Beta: total hemolysis;
zone of clearing. Gamma: no hemolysis.) Alpha & gamma are associated with the normal flora; Beta is
associated with group A streptococci (e.g. Streptococcus pyogenes). Sucrose-gelatin agar: differential for
capsule formation (looking for Streptococcus salivarius). Tellurite agar: selective for Staphylococci, esp.
S. aureus; differential for S. aureus (black colonies)
Lab 27 Bacteriophage. Phages are viruses that infect bacteria. A plaque is a “hole” in the lawn of
bacteria that grew on a plate. The plaque is loaded with phage. Any bacteria in that area failed to grow
because they were killed by virus. PFU=plaque forming unit; calculate original PFU/mL by serial
dilution & plaque count.
Lab 28 Coagulase. A test for Staphylococcus aureus. Coagulase converts fibrinogen into fibrin (clot).
The normal enzyme for this is thrombin. Thrombin requires the cofactor Calcium. EDTA chelates
(binds) calcium. Coagulase will coagulate plasma in the presence of EDTA. Coagulase is a virulence
factor that walls off Staph infection to hide it from the immune system.
Lab 29 Water analysis. See handout.
Lab 31/32 Sauerkraut & cheese. Fermentation of vegetables increases shelf life; acid produced
inhibits growth of microbes that would spoil the food. Anaerobic conditions (fermenters don’t need
oxygen, some rotters do), salt (to draw out water & sugar), bacteria (often catalase --) needed. Halophile:
salt-loving (or salt-tolerant). Two phase fermentation for sauerkraut: Leuconostoc mesenteroides:
heterofermenter; self-inhibits because doesn’t like low pH. Lactobacillus: takes pH down to 3.5 Cheese:
fermented milk. Casein (protein) lactose (carbohydrate). Curd (solid) Whey (liquid). Rennet: cleaves
casein, contributes to curd formation. Found in stomach of young mammals. Streptococcus lactis is
found in buttermilk. Acid produced from fermentation of lactose produces curd. Various products of
fermentation produce interesting flavors.