Chapter 3

The 3 Classifications of Media: physical state, chemical composition, functional type

Liquid Media: water-based solutions that do not solidify above freezing; broths; Bacteria growth is seem as cloudiness/particles

Semisolid media: clot-like consistency at room temp.; determine motility and localize reactions

Solid media: a firm surface on which cells can form discrete colonies; a firm surface on which cells can form discrete colonies

Synthetic Media and example: compositions are precisely chemically defined; Glucose Salt Mediums

Complex (non-synthetic) media and examples: not chemically defined; nutrient broth, yeast extract broth, glucose broth

General purpose media and examples: to grow as broad a spectrum of microbes; usually non-synthetic; nutrient agar and broth, TSA

Enriched media and examples: contain complex organic substances to support fastidious bacteria; blood agar, thayer-martin medium

Selective media and examples: contains one or more agents that inhibit the growth of certain microbes; MSA, MacConkey agar

Differential media and example: allow multiple types of microorganisms to grow but display visible differences; MacConkey agar

Reducing Media: absorbs oxygen or slows its penetration in the medium; used for growing anaerobes or for determining oxygen requirements

Carbohydrate fermentation media: contain sugars that can be fermented and a pH indicator

Transport Media: used to maintain and preserve specimens

Assay Media: test effectiveness of antibiotics, antiseptics, etc.

Enumeration media: count number of organisms in sample

The Five I’s of Culturing Microbes: inoculation, incubation, isolation, inspection, identification

Inoculation: producing a culture

Isolation: Separating species in a culture

Streak Plate Method: small droplet of culture or sample spread over surface of the medium with an inoculating loop

Loop dilation/pour plate method: sample inoculated serially in to a series of liquid agar tubes to dilute the number of cells in each successive tubes

Spread plate method: small volume of liquid, diluted sample pipette on to surface of the medium and spread around evenly by a sterile spreading tool

Incubation: usually around 20-40 Celsius; can control atmosphere as well

Pure culture (axenic): growth of only a single known species

Mixed culture: holds two or more identified species

Inspection and Identification: using appearance as well as metabolism (biochemical tests) and sometimes genetic analysis or immunologic testing to identify the organisms in a culture

Magnification: An image is formed by the refracted light when an object is placed a certain distance from the lens and is illuminated with light. The image is enlarged to a particular degree- the power of magnification

Magnification occurs in two phases: objective lens forms the real image and ocular lens forms the virtual image which equals the total power

Resolution: ability to distinguish two adjacent objects or points from one another

Resolving power equation: wavelength of light in nanometers/ 2 times objective lens

Shorter wavelengths of light equal: better resolution

Increased magnification decreases: the resolution

Visible light microscopes: bright-field, dark-field, phase contrast, and interference

Other light microscopes: Fluorescence microscopes and confocal microscopes

Phase-contrast microscope: allows differentiations of internal components of live unstained cells

Fluorescence Microscopes are commonly used to: diagnose infections caused by microbes using fluorescent antibodies

Confocal Microscopy: Uses a laser beam of light to view cells at higher magnifications and creates 3D model

Electron Microscopy: forms an image with a beam of electrons; increased resolving power and magnification

Transmission electron microscope: Electrons transmitted through specimen; the dark areas represent denser areas

Scanning Electron Microscope: Creates 3D view; Electrons bombard the surface of metal coated specimen

Hanging drop mount: uses depression slide, Vaseline, and coverslip; the sample is suspended from the coverslip

Smear technique: spread thin film made from liquid suspension of cell and airdrying it; heat fixate

Positive staining: dye sticks to specimen to give color

Negative Staining: dye does not stick to specimen and settles around the boundaries; india ink

Simple stain: require only dingle dye

Differential stains: use two different color dyes; primary dye and counter stain

Gram-staining: differentiates gram-positive and gram-negative bacteria

Acid-fast staining: differentiates acid-fast bacteria (pink) and non-acid fast (blue); used to diagnose TB and leprosy

Endospore Stain: dye is forced into endospores by heat

Special Stains: Used to emphasize certain parts are that aren’t emphasized certain cell parts that aren’t revealed by conventional staining