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Chapter 3
Microbiology
| Question | Answer |
|---|---|
| How is the sample processed and profiled? (hint: the five I's) | 1. Inoculation 2. Incubation 3. Isolation 4. Inspection 5. Identification |
| inoculation | to introduce a tiny sample into a container of nutrient medium, which provides an environment in which they multiply or grow |
| incubation | involves placing inoculated media in a temperature controlled environment for a specified time to enhance growth |
| isolation | techniques based on the concept that if an individual bacterial cell is separated from other cells and provided adequate space on a nutrient surface, it will grow into a colony |
| inspection | macroscopic observations of colonies and morphological observations using a microscope |
| identification | biochemical tests to determine nutrient requirements, metabolism, presence of certain enzymes, and mechanism for deriving energy |
| what three properties classifies media? | 1. physical state 2. chemical composition 3. purpose (functional type) |
| what are the four physical states of media? | 1. liquid- broths, milks, and infusions 2. semisolid- clumpy liquid used to check for motility 3. solid- provide surfaces for growth of colonies |
| what are the two chemical states of media? | 1. synthetic- chemically defined media; put together by following a formula 2. complex (non-synthetic)- partially digested meats with unknown exact composition |
| what are the four types of media? | 1. enriched media 2. selective media 3. differential media 4. reducing media |
| enriched media | media that meets special growth needs of certain bacteria |
| selective media | media that inhibits growth of most bacteria, except for the desired type (game of elimination) |
| differential media | media that allows several bacteria to grow, each with a unique indicator (colors, gas bubbles, precipitates, etc.) |
| reducing media | media that contains chemicals that absorb oxygen to help anaerobic microbes grow |
| what are the two necessary aspects of microscopy? | 1. magnification 2. resolution |
| magnification | creating an enlarged view by way of two phases: 1. first, the objective lens forms a real image 2. then the ocular lens forms the virtual image |
| resolution | the ability to reveal detail and see two close objects as distinctively separate objects; anything close than 0.2 micrometers cannot be resolved with the microscopes we use in lab |
| numerical aperture | cone of light that enters the objective lens after passing through the specimen; numerical value is printed on the side of the lens |
| what is the purpose of immersion oil? | the oil immersion lens is the highest power objective lens; the oil used prevents the loss of light due to refraction, effectively increasing the numerical aperture for that objective |
| refractive index | the ability of a substance to bend light |
| bright-field microscopy | the basic light microscope; the image is formed as light passes through the specimen |
| when is the appropriate time to use dark field and phase contrast microscopes? | when staining is NOT appropriate in observing living organisms |
| fluorescence microscopy | the use of fluorochromes (fluorescent dyes) to highlight organisms |
| electron microscopy | consists of two types: 1. transmission microscopy 2. scanning microscopy |
| transmission electron microscopes | transmission of electrons through thin slices of specimens coated with metals for contrast |
| what are the three questions that should be answered when preparing specimens for observation? | 1. Are we observing living or prepared specimens? 2. What are we trying to do (observe, morphology, motility, or identify)? 3. What type of microscope is available? |
| scanning electron microscopes | images of the surfaces of specimens coated with metal for contrast (electrons are deflected back and read by special detectors from which an image is formed) |
| smear | thin film of material is spread over the slide surface; air-dried |
| fixing | attach film to slide prior to staining either chemically or by heating |
| staining | coloring the specimen with a dye; changes the refractive index of specimens so it will contrast with the background |
| simple stains | stain that requires a single dye to highlight the entire organism (shows morphology) |
| differential stains | stain that distinguishes between bacterial types (shows morphology) |
| special stains | stains that show specific parts of the microbes (endospores, flagella, or capsules) |
| gram stains | classifies bacteria into two groups: - gram positive (dyes purple) - gram negative (dyes red/pink) |
| what is the four step process of gram staining? | 1. apply the primary stain (crystal violet) 2. add the mordant (iodine) 3. apply the decolorizing agent (alcohol/acetone) 4. apply the counterstain (safranin) |
| acid fast stains | stains that bind to bacterial cells with waxy material in cell walls; these cells retain red dyes after an acid-alcohol wash and are used to identify bacteria in the genus group, Mycobacterium |
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