Special Media forIsolating BacteriaOBJECTIVESBecause multiple methods and multiple mediaexist, you must be able to match the correct procedureto the desired microbe. For example, if bacterium B issalt-tolerant, a high concentration (5%) of salt couldbe added to the culture medium. Physical conditionscan also be used to select for a bacterium. If bacteri-After completing this exercise, you should be able to:1. Differentiate selective from differential media.2. Provide an application for enrichment and selec-tive media.um B is heat-resistant, the specimen could be heatedbefore isolation. Dyes such as phenol red, eosin, ormethylene blue are sometimes ineluded in differentialBACKGROUNDmedia. Products of bacterial metabolism can react withOne of the major limitations of dilution techniquesused to isolate bacteria is that organisms present inlimited amounts may be diluted out on plates filledwith dominant bacteria. For example, if the culture tobe isolated has 1 million of bacterium A and only 1 ofbacterium B, bacterium B will probably be limited tothe first sector in a streak plate. To help isolate organ-isms found in the minority, various enrichment andselective culturing methods are available that either MATErialsenhance the growth of some organisms or inhibit thegrowth of other organisms. Selective media con-tain chemicals that prevent the growth of unwantedbacteria without inhibiting the growth of the desired Petri plates containing mannitol salt agar (2)organism. Enrichment media contain chemicals thatenhance the growth of desired bacteria. Other bacteria SECOND PERIODwill grow, but the growth of the desired bacteria will bethese dyes to produce a color change in the medium orcolonies (FIGURE 1 and FIGURE 2). The dyes (eosin andmethylene blue) in eosin methylene blue (EMB) agarare also selective. These dyes inhibit the growth ofsome bacteria. Three culture media will be comparedin this exercise (TABLE 1).FIRST PERIODPetri plates containing nutrient agar (2)Petri plates containing EMB agar (2)Gram-staining reagentsBSL-1 Demonstration plate (sealed)with Staphylococcus aureus on one half andStaphylococcus epidermidison the other half.increased.Another category of media useful in identifyingbacteria is differential media. These media containvarious nutrients that allow the investigator to distin-guish one bacterium from another by how they metab-olize or change the media with a waste product.168OCEDNBETABLE 1MAJOR CHEMICAL COMPONENTS OF MEDIA USED IN THIS EXERCISEChemical ComponentsNutrient AgarMannitol Salt AgarEMB AgarPeptone0.5%1.0%1.0%NaCl0.8%7.5%Agar1.5%1.5%1.5%Mannitol1.0%Lactose, sucrose0.5% eachEosin0.04%Methylene blue0.0065%Phenol red0.025%From Exercise 12 of Laboratory Experiments in Microbiology, Eleventh Edition. Ted R. Johnson, Christine L. Case.Copyright © 2016 by Pearson Education, Inc. All rights reserved. SPECIAL MEDIA FOR ISOLATING BACTERIAFIGURE 2 Staphylococcus aureus on mannitol salt agar. Bacte-ria that produce acid from mannitol will cause the phenol red toturn yellow.FIGURE 1 EMB agar. Colonies of Escherichia and Citrobacter de-velop a metallic green sheen on EMB agar.CULTURESStaphylococcusEscherichiacolsnaune(BSL-2 Labs)Escherichia coliJAIRAlcaligenes faecalisStaphylococcus aureus BSL-2Staphylococcus epidermidisUnknown mixed cultureTECHNIQUES REQUIREDCompound light mieroscopySmear preparationGram stainingAseptic techniqueInoculating loop techniqueStreak plate procedureStaphylococcusepidermidisAlcaligenesfaecalisPROCEDURE First PeriodFIGURE 3 Inoculating the plates. Divide a Petri plate into four sec-tions by drawing lines on the bottom of the plate. Inoculate each sectionby streaking it with an inoculating loop.1. Using a marker, divide one nutrient agar plate intofour sections by labeling the bottom. (BSL-1 labs willneed to mark three sections on the plate.) Repeat tomark one mannitol salt plate and one EMB plate.Label one quadrant on each plate for each culture.2. Streak each culture on the agar, as shown in5. Incubate the plates in an inverted position at 35°C.PROCEDURE Second PeriodFIGURE 3.3. Label the remaining nutrient agar, mannitol salt, 1. Record the results after 24 to 48 hours.and EMB plates with the number of your unknown.Record the number of the unknown in the Labora-tory Report. The unknown contains two differentbacteria.4. Streak your "unknown" onto the appropriateplates.

Question

Special Media forIsolating BacteriaOBJECTIVESBecause multiple methods and multiple mediaexist, you must be able to match the correct procedureto the desired microbe. For example, if bacterium B issalt-tolerant, a high concentration (>5%) of salt couldbe added to the culture medium. Physical conditionscan also be used to select for a bacterium. If bacteri-After completing this exercise, you should be able to:1. Differentiate selective from differential media.2. Provide an application for enrichment and selec-tive media.um B is heat-resistant, the specimen could be heatedbefore isolation. Dyes such as phenol red, eosin, ormethylene blue are sometimes ineluded in differentialBACKGROUNDmedia. Products of bacterial metabolism can react withOne of the major limitations of dilution techniquesused to isolate bacteria is that organisms present inlimited amounts may be diluted out on plates filledwith dominant bacteria. For example, if the culture tobe isolated has 1 million of bacterium A and only 1 ofbacterium B, bacterium B will probably be limited tothe first sector in a streak plate. To help isolate organ-isms found in the minority, various enrichment andselective culturing methods are available that either MATErialsenhance the growth of some organisms or inhibit thegrowth of other organisms. Selective media con-tain chemicals that prevent the growth of unwantedbacteria without inhibiting the growth of the desired Petri plates containing mannitol salt agar (2)organism. Enrichment media contain chemicals thatenhance the growth of desired bacteria. Other bacteria SECOND PERIODwill grow, but the growth of the desired bacteria will bethese dyes to produce a color change in the medium orcolonies (FIGURE 1 and FIGURE 2). The dyes (eosin andmethylene blue) in eosin methylene blue (EMB) agarare also selective. These dyes inhibit the growth ofsome bacteria. Three culture media will be comparedin this exercise (TABLE 1).FIRST PERIODPetri plates containing nutrient agar (2)Petri plates containing EMB agar (2)Gram-staining reagentsBSL-1 Demonstration plate (sealed)with Staphylococcus aureus on one half andStaphylococcus epidermidison the other half.increased.Another category of media useful in identifyingbacteria is differential media. These media containvarious nutrients that allow the investigator to distin-guish one bacterium from another by how they metab-olize or change the media with a waste product.168OCEDNBETABLE 1MAJOR CHEMICAL COMPONENTS OF MEDIA USED IN THIS EXERCISEChemical ComponentsNutrient AgarMannitol Salt AgarEMB AgarPeptone0.5%1.0%1.0%NaCl0.8%7.5%Agar1.5%1.5%1.5%Mannitol1.0%Lactose, sucrose0.5% eachEosin0.04%Methylene blue0.0065%Phenol red0.025%From Exercise 12 of Laboratory Experiments in Microbiology, Eleventh Edition. Ted R. Johnson, Christine L. Case.Copyright © 2016 by Pearson Education, Inc. All rights reserved. SPECIAL MEDIA FOR ISOLATING BACTERIAFIGURE 2 Staphylococcus aureus on mannitol salt agar. Bacte-ria that produce acid from mannitol will cause the phenol red toturn yellow.FIGURE 1 EMB agar. Colonies of Escherichia and Citrobacter de-velop a metallic green sheen on EMB agar.CULTURESStaphylococcusEscherichiacolsnaune(BSL-2 Labs)Escherichia coliJAIRAlcaligenes faecalisStaphylococcus aureus BSL-2Staphylococcus epidermidisUnknown mixed cultureTECHNIQUES REQUIREDCompound light mieroscopySmear preparationGram stainingAseptic techniqueInoculating loop techniqueStreak plate procedureStaphylococcusepidermidisAlcaligenesfaecalisPROCEDURE First PeriodFIGURE 3 Inoculating the plates. Divide a Petri plate into four sec-tions by drawing lines on the bottom of the plate. Inoculate each sectionby streaking it with an inoculating loop.1. Using a marker, divide one nutrient agar plate intofour sections by labeling the bottom. (BSL-1 labs willneed to mark three sections on the plate.) Repeat tomark one mannitol salt plate and one EMB plate.Label one quadrant on each plate for each culture.2. Streak each culture on the agar, as shown in5. Incubate the plates in an inverted position at 35°C.PROCEDURE Second PeriodFIGURE 3.3. Label the remaining nutrient agar, mannitol salt, 1. Record the results after 24 to 48 hours.and EMB plates with the number of your unknown.Record the number of the unknown in the Labora-tory Report. The unknown contains two differentbacteria.4. Streak your &#34;unknown&#34; onto the appropriateplates.

Accepted Answer

Solid media (agar) is most often used to culture bacteria and fungi as discrete, single colonies – a…