a sample is placed onto a media to culture a microbe
(5 I’s)
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incubation
placing the medium container into a temperature-controlled chamber to encourage growth
(5 I’s)
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pure culture
only one type of species in a culture
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mixed culture
two or more species in a culture
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contaminated culture
a pure or mixed culture plus an unwanted microbe in a culture
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media classification
1. physical state (liquid, semi-solid, solid) 2. chemical composition 3. functional type
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general purpose media
media type that promotes growth in a broad spectrum of microbes
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enriched media
media type that contains complex organic compounds for the growth of fastidious (having complex/precise nutrient requirements) microbes
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selective media
media type that inhibits the growth of certain microbe(s) to isolate the particular microbe of study
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differential media
media type that allows multiple types to grow but displays visible differences in how they grow
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isolation
separating an individual bacterial cell to create a colony
(5 I’s)
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inspection & identification
looking at the microscopic appearance, determining nutrient requirements, products given off during growth, and other characteristics
(5 I’s)
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virus size
20-400 but up to 800-1500 nm
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bacteria size
200 nm-750micro-m
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protozoa size
100-300micro-m
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principles of light microscopy
1. magnification 2. resolution 3. contrast
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objective lens, ocular lens
magnification occurs in 2 phases, first in the ___ and second in the ___
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oil immersion lens (100x)
lens that reduces scatter to increase resolution
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bright-field microscopy
* most widely used type of light microscope * forms image when light is transmitted through specimen * can be used for live, unstained/preserved, and stained materials
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dark-field microscopy
* adapted from bright-field microscope by adding a “stop” to block light from entering the objective lens * used to visualize living cells that would be distorted by drying or heating or cannot be stained by usual methods
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phase-contrast microscopy
* used to observe intracellular structures such as organelles, endospores, etc.
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fluorescence microscopy
* uses a specially modified compound microscope with a UV light * the use of certain dyes show fluorescence * specimen is first coated/placed in contact with fluorescence source * used to diagnose infections and pinpoint particular cellular structures
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confocal microscopy
* uses a laser beam to scan various depths of the specimen * used on fluorescently stained specimens or to visualize live unstained cells/tissues
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transmission electron microscope (TEM)
* transmits electrons through specimen * used to view detail structure of cells and viruses * dark areas = denser areas
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scanning electron microscope (SEM)
* specimens are metal-coated while electrons scan back and forth over it * color is always added after, actual images are black and white
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simple stain
stain that requires only a single dye and caused all the cells to appear the same color which reveals its shape, size, and arrangement
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differential stain
stain type that uses 2 differently colored dyes (primary dye and counterstain) to distinguish cell types or parts
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gram stain
differential stain type
* gram positive = purple * gram negative = pink
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acid-fast stain
differential stain type
* acid-fast = reddish/purple * non-acid-fast = blue
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endospore stain
differential stain type
* endospores = green * vegetative cells = pink
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capsular staining
* used to observe an unstructured protective layer surround the cells of some bacteria and fungi * cell is negatively stained with India ink
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flagellar staining
* used to reveal tiny, slender filaments used by the bacteria for locomotion * these filaments are enlarged by depositing a coating on the outside of the filament and then staining it
external bacterial structures (2) used for motility
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fimbriae
small, bristle-like fibers that sprout off the surface of many bacterial cells and allow for adhesion between cells, allowing bacteria to colonize and infect host tissues
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pili
used in cell-to-cell contact between bacterial cells (usually for the transfer of genetic material)
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capsule
a type of glycocalyx that is formed by many pathogenic bacteria to protect against phagocytosis and produces a sticky character to colonies on agar
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biofilms
groups of bacteria attached to a particular coating
ex: plague on teeth
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cell envelope
part of bacterial cell that lies outside the cytoplasm
composed of: cell wall, cytoplasmic membrane, and (sometimes) outer membrane
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bacterial cell wall
determines the shape of a bacterium, provides support and protection, composed of peptidoglycan (contributes rigidity)
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gram-positive cell wall
cell wall that lacks outer membrane
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crystal violet
1st step of Gram staining
all cells appear purple
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Gram’s iodine
2nd step of Gram staining
the mordant (stabilizer) causes the dye to form large complexes in the cell wall
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alcohol
3rd step of Gram staining
dissolves the lipids in the outer membrane and removes dye from the gram-negative cells
all appear purple (+) or colorless (-)
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safranin (red dye)
4th step of Gram staining
dyes colorless (-) cells to make their presence apparent
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cytoplasm
internal bacterial structure
composed of sugars, amino acids, and salts and is 70-80% water
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ribosomes
sites of protein synthesis
composed of rRNA (60%) and protein (40%)
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endospores
dormant bodies that can resist extremes of heat, drying, freezing, radiation, chemicals, etc.
exist in either a dormant or vegetative stage
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Bergey’s Manual of Systematic Bacteriology
classification system for bacteria and archaea that is based on rRNA sequencing
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Bergey’s Manual of Determinative Bacteriology
classification system for bacteria and archaea based on phenotypic characteristics
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chitin, no cell wall
fungi cell walls are made of ___ while protozoa and helminths have ___
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nucleus
internal eukaryotic structure that contains DNA and is separated from the cytoplasm by an envelope
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endoplasmic reticulum
eukaryotic internal structure that is a series of membrane tunnels used in transport and storage
may be smooth or rough
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Golgi body
eukaryotic internal structure that is the site of protein modification and shipping
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vacuoles
internal eukaryotic structure that are sacs containing fluid or solid particles
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mitochondria
internal eukaryotic structure that generates ATP for the cell
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cytoskeleton
internal eukaryotic structure made of protein strands for structure, movement, and transport
motile feeding stage of protozoa that requires ample food and moisture to stay active
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cyst
dormant, resting stage of protozoa when conditions in the environment become unfavorable
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helminth life cycle
fertilized egg, larva, adult
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human genome
8% of the ___ consists of sequences that come from viruses
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active/inactive
terms used to describe viruses instead of alive/dead
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properties of viruses
* ubiquitous in nature * ultramicroscopic in size * not cells, compact and economical structure * do not independently fulfill characteristics of life * basic structure: protein shell capsid that surrounds nucleic acid core * nucleic acid that is either DNA or RNA (not both) * lacks enzymes for most metabolic processes and lacks machinery for synthesizing proteins
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capsid
a protein shell that surrounds viral nucleic acid
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spikes
projections from the nucleocapsid or envelope of a virus that allows it to dock with host cells
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virion
a fully formed virus that is able to establish an infection