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Foundations in Microbiology Chapter 3 PowerPoint to accompany Fifth Edition Talaro Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction.

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Presentation on theme: "Foundations in Microbiology Chapter 3 PowerPoint to accompany Fifth Edition Talaro Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction."— Presentation transcript:

1 Foundations in Microbiology Chapter 3 PowerPoint to accompany Fifth Edition Talaro Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

2 Tools of the Laboratory: The Methods for Studying Microorganisms Chapter 3

3 3 The 5 I’s of culturing microbes 1.Inoculation – introduction of a sample into a container of media 2.Incubation – under conditions that allow growth 3.Isolation –separating one species from another 4.Inspection 5.Identification

4 4 Isolation If an individual bacterial cell is separated from other cells & has space on a nutrient surface, it will grow into a mound of cells- a colony A colony consists of one species

5 5 Isolation technique

6 6 Media – providing nutrients in the laboratory Most commonly used: –nutrient broth – liquid medium containing beef extract & peptone –nutrient agar – solid media containing beef extract, peptone & agar agar is a complex polysaccharide isolated from red algae –solid at room temp, liquefies at boiling (100 o C), does not resolidify until it cools to 42 o C –provides framework to hold moisture & nutrients –not digestible for most microbes

7 7 Types of media synthetic – contains pure organic & inorganic compounds in an exact chemical formula complex or nonsynthetic – contains at least one ingredient that is not chemically definable general purpose media- grows a broad range of microbes, usually nonsynthetic enriched media- contains complex organic substances such as blood, serum, hemoglobin or special growth factors required by fastidious microbes

8 8 Enriched media

9 9 selective media- contains one or more agents that inhibit growth of some microbes and encourage growth of the desired microbes differential media – allows growth of several types of microbes and displays visible differences among desired and undesired microbes

10 10 selective & differential media

11 11 Selective media

12 12 Differential media

13 13 Miscellaneous media reducing medium – contains a substance that absorbs oxygen or slows penetration of oxygen into medium; used for growing anaerobic bacteria carbohydrate fermentation medium- contains sugars that can be fermented, converted to acids, and a pH indicator to show the reaction; basis for identifying bacteria and fungi

14 14 Carbohydrate fermentation media

15 15 magnification – ability to enlarge objects resolving power – ability to show detail

16 16 compound light microscope

17 17 Pathway of light

18 18 Effect of wavelength on resolution

19 19 Oil immersion lens

20 20 Effect of magnification

21 21 Types of light microscopes Bright-field – most widely used, specimen is darker than surrounding field Dark-field – brightly illuminated specimens surrounded by dark field Phase-contrast – transforms subtle changes in light waves passing through the specimen into differences in light intensity, best for observing intracellular structures

22 22 3 views of a cell

23 23 Fluorescence Microscope Modified compound microscope with an ultraviolet radiation source and a filter that protects the viewer’s eye Uses dyes that emit visible light when bombarded with shorter uv rays. Useful in diagnosing infections

24 24

25 25 Electron microscopy Forms an image with a beam of electrons that can be made to travel in wavelike patterns when accelerated to high speeds. Electron waves are 100,000X shorter than the waves of visible light. Electrons have tremendous power to resolve minute structures because resolving power is a function of wavelength. Magnification between 5,000X and 1,000,000X

26 26

27 27 2 types of electron microscopes Transmission electron microscopes (TEM) – transmits electrons through the specimen; darker areas represent thicker, denser parts and lighter areas indicate more transparent, less dense parts Scanning electron microscopes (SEM)– provides detailed three-dimensional view. SEM bombards surface of a whole, metal-coated specimen with electrons while scanning back and forth over it.

28 28 Transmission Electron Micrograph

29 29 Scanning Electron Micrograph

30 30 Specimen preparation wet mounts & hanging drop mounts – allow examination of characteristics of live cells: motility, shape, & arrangement fixed mounts are made by drying & heating a film of specimen. This smear is stained using dyes to permit visualization of cells or cell parts.

31 31 Staining cationic dyes - basic, with positive charges on the chromophore anionic dyes - acidic, with negative charges on the chromophore surfaces of microbes are negatively charged and attract basic dyes – positive staining. negative staining – microbe repels dye & it stains the background

32 32 Staining simple stains –one dye is used differential stains – use a primary stain and a counterstain to distinguish cell types or parts. examples: Gram stain, acid-fast stain and endospore stain special stains: capsule and flagellar stains

33 33 Types of stains


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