Slide 1Slide 2Slide 3Slide 4Slide 5Slide 6Slide 7Slide 8Slide 9Slide 10Lab 7 Goals and Objectives:Gather all plates and tubes so we can discuss results together: ***Do NOT shake the FTM tubes!!!***Your Unknowns: Exercises 37 and 38Check plates, record appearance, check for contaminationCheck slants for growth and place in my “reserve” rackMake new inoculations for both unknowns from the plates:1. 2 FTM tubes for O2 requirements (Ex. 38 pg. 267) **leave caps loose to incubate!2. 2 BHI broth tubes for growth in liquid characteristics (Ex. 38 pg. 266 fig. 38.2) and for Gram stain (Ex. 37 pg. 260) 3. 2 Gelatin stab cultures for gelatin hydrolysis ability (Ex. 38 pg. 259)4. 3 BHI plates each unknown (6 total), streak for isolation, to grow at 25°C, 30°C, 37°C to determine optimal temp. (Ex. 37&38 had you using slants and broths; side by side plates is easier)5. 2 Motility test media stabs (Ex. 37 pg. 262) No: KOH, endospore, acid fast, other structures (Ex. 37 pg. 261-262)(From the Media List in the Supplemental packet)Motility Test MediumInoculation method: vertical single stab with straight needleContains: nutrient medium with low (0.5%) agar concentration (semisolid) and TTC which changes from colorless to dark pink (reduced) 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-motileorganism appears as haze beyond line of inoculation = motileFig 18.4Motility Test Media ResultsNon-motileMotileNonMotileBrewer’s Anaerobic AgarInoculation 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 plate in aerobic 20% oxygen atmosphere.Contains: Nutrient agar with sodium thioglycolate and resazurin (see FTM)Discriminates oxygen requirements if read in conjunction with normal incubated plate: obligate aerobes, obligate anaerobes, facultative anaerobes, aerotolerantResults: growth on aerobic agar only = obligate aerobegrowth on anaerobic agar only = obligate anaerobeeven growth on both = aerotolerantheavy growth on aerobic, lighter growth on anaerobic = facultative Fig 27.2Fig 27.4Which Groups Grow on Brewer’s Anaerobic Agar In the Anaerobic Jar?Which Groups Grow on Brewer’s in 20% Oxygen?Brewer’s No OxygenBrewer’s 20% OxygenObligate anaerobes: O2 toxic Obligate aerobes: 20% O2Aerotolerant: ignore O2Facultative: w/ or w/o, better with O2Microaerophiles: 5-10% O2Fig 27.1Obligate aerobes: 20% O2Obligate anaerobes: O2 toxicFacultative: w/ or w/o, better with O2Microaerophiles: 5-10% O2Aerotolerant: ignore O2Fig 27.1Fluid Thioglycolate Medium (FTM)Inoculation method: loop transfer, careful mixing, screw cap must be looseContains: 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, aerotolerantResults: growth only at top = obligate aerobegrowth only at bottom = obligate anaerobeeven growth throughout = aerotolerantheavy growth at top, lighter growth at bottom = facultativegrowth only in middle = microaerophile•Record all data for both unknowns: fill in on blank data report in lab•Type in data for each category on the data report on the day you collect it: do not wait until the end of the project!Data to collect for Exercise 37 & 38 For Next LabGram result, size, shape, arrangementTransfer to Data ChartMotilityTransfer to Data ChartAmount of growthColorOpacityFormSurface (broth)Subsurface (broth)Sediment (broth)Growth (broth)TemperatureTransfer to Data ChartOxygen requirements (FTM)Transfer to Data ChartGelatinTransfer to Data ChartColoniesLab 7 Goals and Objectives:Gather all plates and tubes so we can discuss results together: ***Do NOT shake the FTM tubes!!!***Your Unknowns: Exercises 37 and 38Check plates, record appearance, check for contaminationCheck slants for growth and place in my “reserve” rackMake new inoculations for both unknowns from the plates:1. 2 FTM tubes for O2 requirements (Ex. 38 pg. 267) **leave caps loose to incubate!2. 2 BHI broth tubes for growth in liquid characteristics (Ex. 38 pg. 266 fig. 38.2) and for Gram stain (Ex. 37 pg. 260) 3. 2 Gelatin stab cultures for gelatin hydrolysis ability (Ex. 38 pg. 259)4. 3 BHI plates each unknown (6 total), streak for isolation, to grow at 25°C, 30°C, 37°C to determine optimal temp. (Ex. 37&38 had you using slants and broths; side by side plates is easier)5. 2 Motility test media stabs (Ex. 37 pg. 262) No: KOH, endospore, acid fast, other structures (Ex. 37 pg.