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Terms in this set (56)
The five I's
introduction of microorganisms into or onto a new environment/culture media
culturing of microbes under controlled optimal environmental conditions for a period of time
usual lab propagation temperatures fall between 20-40*C
during incubation, microbes grow and multiply, producing visible growth in the media (colony)
the separation of microbes from a mixed culture into isolated colonies for the purpose of creating a pure culture
a macroscopic cluster of cells appearing in a solid medium arising from the multiplication of a single cell
visible accumulation of microorganisms in or on a medium
macro and microscopic evaluations of microbial growth and cellular characteristics
biochemical and physiological characterization of the microbe of interest.
microbes can be identified through
-characterization of cellular metabolism
-determination of products given off during growth, presence of enzymes, and mechanisms for deriving energy
-genetic and immunological characteristics
how are pure, mixed, and contaminated cultures similar?
all contain live microbes and these microbes can be propagated.
contain only one type of KNOWN microbe
contain more than one type of known microbe but originate from one specimen
contain additional microbes from unknown sources such as air, fomites, or from the microbiologist. UNWANTED microbes.
how to avoid contamination?
use ASEPTIC technique. sterilizing instruments, media, and vessels, as well as keeping cultures closed from the environment are all important.
(singular medium): nutrients used to grow microorganisms outside their natural habitat
media in different physical forms
LIQUID: growth occurs throughout container.
SEMISOLID: do not flow freely and have soft consistency at room temp.
SOLID: 1-5% agar. firm surface for cells to form discrete coloniese.
complex polysaccharide from alga "Gellidium"
liquefies at 100
C and solidifies at 42
C and can be poured in liquid form that will not harm the microbe or handler
flexible and moldable; can hold moisture and nutrients
not a digestible nutrient for microorganisms
three types of functional media
enriched, selective, and differential
encourage growht of microbes w/special growth requirements (e.g. blood added to media enhances growth of fastidious streptococci)
contains one or more agents that inhibit the growth of certain microbes but not others
important in the primary isolation of a specific type of microorganism from samples containing dozens of species
allow multiple types of microorganisms to grow but are designed to display differences among those microorganisms
differentiation shows as variations in colony size or color, media color changes, formation of gas bubbles or precipitates
media can be selective and differential T/F?
what is required for proper isolation? what are three isolation techniques?
to isolate microorganisms from a mixed sample, a SOLID MEDIUM is used along w/DILUTION TECHNIQUES to achieve isolated colonies.
-3 common lab techniques for separating a mixed sample: QUADRANT STREAK, LOOP DILUTION, AND SPREAD PLATE. all achieve dilution!
added notes on isolatation.
an isolated colony is formed from an original microscopic parent cell deposited on the surface of a solid medium that reproduces to form a mass of cells that is macroscopic.
an isolated colony differs from a pure culture in that the isolated colony is used to create a pure culture for purposes of identification.
what are two principal functions of dyes in media?
1. growth inhibitors in selective media
2. pH indicators in differential media
How are observations made in microbiology? what tools are necessary?
MICROSCOPIC OBSERVATIONS: require a microscope to see individual cells and characterize them based upon their size, shape, arrangements, physical features (e.g. flagella) and color, if using a differential stain (e.g. Gram stain).
MACROSCOPIC OBSERVATIONS: require culture media that expand populations to visible masses or colonies. these growths can be characterized based upon growth patterns, color, texture, size, and odor. the media may also contain substances (e.g. differential media) that allow for the characterization of species based upon biochemical or physiologic properties of the microbe.
how do you improve resolution w/a microscope?
-objective lens-- forms initial image called the real image
-ocular lens: forms the second image called the virtual image that will be received by the eye and converted to the retinal and visual image
aka resolving power.
the capacity of an optical system to distinguish or separate two adjacent points or objects from one another
the human eye can resolve two objects that are no closer than 0.2 mm apart.
resolving power formula
resolving power (RP)= wavelength of light in nm/ 2x numerical aperture
ways to improve resolution:
shorter wavelengths provide a better resolution
numerical aperture is how well the lens gathers light
oil immersion lens
-uses oil to capture light that would otherwise be lost to scatter
-reducing scatter increases resolution
-oil immersion lens can resolve images that are at least 0.2 micrometers in diameter and at least 1.2 micrometers apart
uses visible light (tungsten bulb)
bright field scope
is used to observe most microbes
visible light passes through multiple lenses and through the specimen
light microscopes usually have at least 3 lenses: low-power, high-power, and oil-immersion
the lens system must have high resolving power to see the specimen clearly
fixed, stained smears
more permanent mounts used for long-term study
smear technique developed by Robert Koch over 100 years ago
-spread a thin film made from a liquid suspension of cells on a slide
-heat fix: heat gently to kill the specimen and attach to the slide
staining is any procedure that applies colored chemicals (dyes) to specimens
-BASIC dyes have a POSITIVE charge
-ACIDIC dyes have a NEGATIVE charge
bacteria have numerous negatively charged substances and attract basic dyes
acidic dyes are repelled by cells
dye sticks to specimen and gives it color
does not stick to specimen but settles some distance from its outer boundary, forming a silhouette
- - charged cells repel the negatively charged dye and remain unstained
- smear is not heat fixed so there is reduced distortion and shrinkage of cells
- also used to accentuate a capsule
-nigrosin and India ink are used
simple vs differential staining: SIMPLE STAINS
only require a single dye and an uncomplicated procedure
-cause all the cells in smear to appear more or less the same color, regardless or type
-reveal shape, size, and arrangement
simple vs differential staining: DIFFERENTIAL STAINS
use 2 differently colored dyes: the primary dye and counterstain
distinguish cell types or parts
more complex and require additional chemical reagents to produce the desired reaction
types of differential stains
Gram stain and acid-fast stain
developed in 1884 by Hans Christian Gram
consists of sequential applications of crystal violet (primary stain), iodine (mordant), an alcohol rinse (decolorizer), and safranin (teh counterstain)
different results in the Gram stain are due to differences in the structre of the cell wall and hwo it reacts to the series of reagents applied to the cells
remains the universal basis for bacterial classification and identification
differentiates acid-fast bacteria (pink) from non-acid-fast bacteria (blue)
originated as a method to detect Mycobacterium tuberculosis
used to detect Mycobacterium, due to their mycolic acid (waxy) cell wall.
used to emphasize cell parts that are not revealed by conventional staining methods
used to observe the microbial CAPSULE, an unstructured protective layer surrounding the cells of some bacteria and fungi
negatively stained with India ink
used to reveal tiny, slender filaments used by bacteria for locomotion
flagella are enlarged by depositing a coating on the outside of the filament and then staining it.
select methods below that enable isolation of bacteria:
quadrant streak plate
bright field microscopy
use of selective media
quadrant streak plate
use of selective media
examples of positive stains
examples of negative stains
scope of microbiology includes bacteria, several types of eukaryotic microorganisms, and even noncellular, parasitic particles called _____.
major groups of microorganisms include:
_____ term used to describe field of science that employs the human manipulation of microbes for used in industrial processes
all microorganisms are too small to be seen without aid of microscope. t/f?
Best describe Woese-Fox taxonomic system
3 distinct cell lines called domains
a type of complex media, ____ media is supplemented w/organic substances such as blood, serum, or special growth factors and is formulated to support the growth of _____ microorganisms.
a medium that has been designed to support growth of MRSA while inhibiting all species and strains of other bacteria.
reducing the intensity of light using the iris diaphragm or staining a specimen can improve ____.
in positive staining, the organism is stained w/dies that have a ___ charge.
[it will adhere to the negative charge on cell surfaces]
ability to make objects appear larger to view them is known as _____ while being able to see detail and distinguish two separate objects is known as _____.
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