Presentation on theme: "Foundations in Microbiology"— Presentation transcript:
1Foundations in Microbiology PowerPoint to accompanyFoundations in MicrobiologyFifth EditionTalaroChapter3Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
2Tools of the Laboratory: The Methods for Studying MicroorganismsChapter 3
3The 5 I’s of culturing microbes Inoculation – introduction of a sample into a container of mediaIncubation – under conditions that allow growthIsolation –separating one species from anotherInspectionIdentification
4IsolationIf an individual bacterial cell is separated from other cells & has space on a nutrient surface, it will grow into a mound of cells- a colonyA colony consists of one species
6Media – providing nutrients in the laboratory Most commonly used:nutrient broth – liquid medium containing beef extract & peptonenutrient agar – solid media containing beef extract, peptone & agaragar is a complex polysaccharide isolated from red algaesolid at room temp, liquefies at boiling (100oC), does not resolidify until it cools to 42oCprovides framework to hold moisture & nutrientsnot digestible for most microbes
7Types of mediasynthetic – contains pure organic & inorganic compounds in an exact chemical formulacomplex or nonsynthetic – contains at least one ingredient that is not chemically definablegeneral purpose media- grows a broad range of microbes, usually nonsyntheticenriched media- contains complex organic substances such as blood, serum, hemoglobin or special growth factors required by fastidious microbes
9selective media- contains one or more agents that inhibit growth of some microbes and encourage growth of the desired microbesdifferential media – allows growth of several types of microbes and displays visible differences among desired and undesired microbes
13Miscellaneous mediareducing medium – contains a substance that absorbs oxygen or slows penetration of oxygen into medium; used for growing anaerobic bacteriacarbohydrate 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
21Types of light microscopes Bright-field – most widely used, specimen is darker than surrounding fieldDark-field – brightly illuminated specimens surrounded by dark fieldPhase-contrast – transforms subtle changes in light waves passing through the specimen into differences in light intensity, best for observing intracellular structures
23Fluorescence Microscope Modified compound microscope with an ultraviolet radiation source and a filter that protects the viewer’s eyeUses dyes that emit visible light when bombarded with shorter uv rays.Useful in diagnosing infections
25Electron microscopyForms 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
272 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 partsScanning 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.
30Specimen preparationwet mounts & hanging drop mounts – allow examination of characteristics of live cells: motility, shape, & arrangementfixed mounts are made by drying & heating a film of specimen. This smear is stained using dyes to permit visualization of cells or cell parts.
31Stainingcationic dyes - basic, with positive charges on the chromophoreanionic dyes - acidic, with negative charges on the chromophoresurfaces of microbes are negatively charged and attract basic dyes – positive staining.negative staining – microbe repels dye & it stains the background
32Staining 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 stainspecial stains: capsule and flagellar stains