Exercise 18: Motility Determination Read results Lab 7 Goals and Objectives: Exercise 18: Motility Determination Read results Exercise 27: Effect of Oxygen on Growth Understand FTM patterns of growth Unknown Project Data collection for profiles Each pair get one organism to collect data on to share with the class to develop profiles of all possible organisms. Each person inoculate one BHIA streak for isolation plate and one BHI broth of assigned organism

Exercise 18: Motility Determination Motility Test Medium Inoculation method: vertical single stab with straight needle Contains: nutrient medium with low (0.5%) agar concentration (semisolid) and TTC which changes from colorless to dark pink in the presence of bacterial growth (enhances visualization) Discriminates motility (presence of flagella), ability to “swim” through media Results: organism growing only in line of inoculation = non-motile organism appears as haze beyond line of inoculation = motile Figure 18.4

Motility Test Media Results Non-motile Motile Non Motile

Exercise 27: Effect of Oxygen on Growth Obligate aerobes: 20% O2 Obligate anaerobes: O2 toxic Facultative: w/ or w/o, better with O2 Microaerophiles: 5-10% O2 Aerotolerant: ignore O2 Fig 27.1

Fig 27.1 Fluid Thioglycolate Medium (FTM) Inoculation method: loop transfer, careful mixing, screw cap must be loose Contains: rich medium with very low agar content (viscous) Sodium thioglycolate (removes oxygen) Resazurin oxygen indicator (pink when oxidized: O2 present) Discriminates oxygen requirements: obligate aerobes, obligate anaerobes, facultative anaerobes, microaerophiles, aerotolerant Results: growth only at top = obligate aerobe growth only at bottom = obligate anaerobe even growth throughout = aerotolerant heavy growth at top, lighter growth at bottom = facultative growth only in middle = microaerophile Fig 27.1

Supplement pg 52 Growth Patterns Observed in FTM Tubes

The gas pack releases H2. This combines with the O2 in the air creating water. The oxygen is chemically removed from the air as it gets bound by the hydrogen to form water molecules, resulting in an anaerobic environment in the jar.

Fig 27.4 Fig 27.2 Brewer’s Anaerobic Agar Inoculation method: surface streak with loop, must be incubated in Brewer’s anaerobic jar (water + gas pack = H2 + CO2, H2 combines with O2 creating H2O, sealed jar is oxygen free). Inoculate in conjunction with a second plate in aerobic 20% oxygen atmosphere to determine O2 needs. Contains: Nutrient agar with sodium thioglycolate and resazurin (see FTM) Discriminates oxygen requirements if read in conjunction with normal aerobic incubated plate: obligate aerobes, obligate anaerobes, facultative anaerobes, aerotolerant Results: growth on aerobic agar only = obligate aerobe growth on anaerobic agar only = obligate anaerobe even growth on both = aerotolerant heavy growth on aerobic, lighter growth on anaerobic = facultative Fig 27.4 Fig 27.2

Obligate anaerobes: O2 toxic Obligate aerobes: 20% O2 Which Groups Grow on Brewer’s Anaerobic Agar In the Anaerobic Jar? Which Groups Grow on Brewer’s in 20% Oxygen? Brewer’s No Oxygen Brewer’s 20% Oxygen Obligate anaerobes: O2 toxic Obligate aerobes: 20% O2 Aerotolerant: ignore O2 Facultative: w/ or w/o, better with O2 Microaerophiles: 5-10% O2

Klebsiella pneumoniae Proteus vulgaris Pseudomonas aeruginosa Possible Unknowns: Bacillus subtilis Enterobacter cloacae Escherichia coli Klebsiella pneumoniae Proteus vulgaris Pseudomonas aeruginosa Salmonella typhimurium Shigella flexneri Staphylococcus aureus Staphylococcus epidermidis Enterococcus (Streptococcus) faecalis Streptococcus pyogenes

Exercise 18: Motility Determination Read results Lab 7 Goals and Objectives: Exercise 18: Motility Determination Read results Exercise 27: Effect of Oxygen on Growth Understand FTM patterns of growth Unknown Project Data collection for profiles Each pair get one organism to collect data on to share with the class to develop profiles of all possible organisms. Each person inoculate one BHIA streak for isolation plate and one BHI broth of assigned organism