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MicroBiology Mid term-Lab
Terms in this set (139)
What is a smear?
A thin film of material containing microorganisms spread over the surface of a clean microscope slide
Sucess at bacterial staining depends on what?
preperation of a suitable smear of the organism
What is heat-fixing?
using a flame; the heating of a smear on a slide briefly and gently. partially melts the cell walls of bacteria
What are the three things Heat-fixing does?
1: causes the cells to adhere to the slide so it wont wash off
2:Kills the microorganisms
3:Causes changes in the bacterial cells which causes them to stain better
Hows does Heat-Fixing work?
It removes water from the bacterial cell, pores open in the cell so more stain can enter the cell wall.
A good heat-fixes smear should be able to do what 2 things?
1. Must be able to withstand ane or more washings w/out the loss of microorganisms
2.Does not result in excessive cell distortion ***heat-fixing always results in some degree of cell distortion,
Name 7 things that may go wrong in heat-fixing.
1.Over heat; incincerated cells will not stain
2.Under heat;if bacteria cells aren't killed, their ability to accept stain is altered
3.Must be Air-Dried; if not organism could be boiled and rupured and will not adhere to slidw
4.Too thick; light cannot pass, you cannot see individual cells
5.Too Thin; not enough cells to stain or see
6.Liguid broth;may be too little cells;use 2-3 loopfuls to compansate
7.Solid media (Agar);colonies are here and may have too many;dilute with 1-2 looful of distilled water.
It is an absolute value : 100%
-The complete destruction or removal of all microbes, including viruses(nonliving) & endospores(Resting)
How are the organisms killed?
Steam; dry heat; incineration
A process in which vegetative microorganisms are destroyed on non-living objects and surfaces
What does vegetative mean?
What is the number 1 agent for disinfection?
What are the 3 definitions of Sepsis?
1:The growth of microorganisms in normally sterile tissues of the body
3:The presence of bacteria in blood that ate ectively multiplying (Serious)
A medical tem, any procedure that prevents the entrance of infectious agents into sterile tissue, thus preventing infection.
What is Aseptic Techniques?
Practices used by micobiologists to exclude all organisms from contaminating media or contacting living tissues
Chemical agents ( often dilute disinfectants) that can be safely applied, externally, yo living tissues (skin, mucous membranes) to destroy or inhibit vegetative bacteria
What is the #1 Antiseptic
List 7 facts about Bacteria.
1. Most numerous and widely distributed organism
2.Found eveywhere in nature
3.Where microbes are normally found(like staph on skin) is presumably where they are adapted to
4.They are not confined to to those areas and can easily move to other environments
5. They can be transfered easily
6.They can tolerate and take advantage of their new environment
7.Need Aseptic techniques for handling micobes
What is Media?
Any nutrient substance ( solid or liquid) that can support microbial growth
What is Inoculate?
To introduce microorganisms into normally sterile media
What is a Fomite?
Any non-living, inanimte abject that can carry (transmit) micobes
(and and ourselves are not fomites)
Smallest living organisms & most are transparent & colorless
What is ultrastructure?
The fine (microscopic) details of organisms, tissue, cell.
*can only see w/ microscope
An optical instrumeny consisting of a lens or a combo of lenses for making enlarged images of minute objects
What are the 2 abilities of a microscope?
Is an infinite power
1: the ability of a lens to make an image larger
2. To have the power of causing abjects to appear larger than they are
Focue to See
1. The ability to see the fine details of an image in Clear, Sharp focus
2.The process or capaility of making distinguishable the individual parts of an object
Name the 2 types of microscopes
Define Simple Microscope
AKA: Magnifying glass
-Only has 1 lens
Define Compund Microscope
-consisting of 2 lenses
Objective lens & Ocular Lens
Define Ocular Lens
AKA; Eye Piece
-you look through it
-magnifies the image made by the objective lens
Define Objective Lens
-Does the intial magnification of the specimen on the slide
- The longer the lens=the more powerful
What is the Condenser?
-3rd lesn system in a compound light microscope
-it collects & directs the light from the lamp to the slide being studied
What is Parfocalized?
-Only minor focusing adjustments are needed whan changing from low to high powers
-only on good quality microscopes
What is the power on the Red ring ocular lens?
-Total magnification 40X
What is the power on the Yellow ring ocular lens?
-Total magnification 100X
What is the power on the Blue Ring Ocular lens?
-Total magnification 400X
What is the power on the White Ring ocular lens?
-Total magnification 1000X
What is Oil Immersion technique?
-Special Mineral Oil
-is used because it has same refractive index as glass
-Prevents the loss of light(resolution) due to pending of rays as they pass through air
-Enhances the resolving power of the microscope
The study of the form & structure of organisms & their specific structural features
What are the 4 different properties of Moprphology?
2.Shape (3 of them)
3.Cell arrangement:(genetically determined)
4.Gram Staining reaction (Purple/pink +/-)
What do almost all bacteria cells have?
Characteristic and genetically determined shape
-1 permanent shape of bacteria, (shape dictates function) due to its rigid external cell wall called Peptidoglycan
Makes up the bacteria cell wall
composed of unique complex carbohydrate
Name the 3 Classification of Bacteria Shapes
-Bacilli is plural
-"staff" rod-shaped bacteria
-Cocci is plural
-"Berry" sphere or round-shaped bacteris
-tiny little balls
-helical or curved shape
-3 types of spiral
What are the 3 types of spiral shape bactera?
Define Vibrio shape
Define Spirchetes shape
Rigid cork-screw shaped
List the 6 classification by arrangement?
2.diplo- a pair of cells
3.Tetrad- group of 4
4.Sarcinae- group of 8
5.Strepto- chains of cells
6.Staphylo- Grape like clusters
Gram Negative turns what color?
Pink after staining
Gram Positive turns what color?
Purple after staining
Small G- bacilli are always in what arrangement?
Large G+ Bacilli are always what arrangement?
Spiral bacteria are always in what arrangement?
G+ Cocci can be in 4 arrangements, name them.
Tetrads, sarcinae,Strepto, Staphylo
Stepto arrangement is the only shared by who?
G+ cocci and G+ bacilli
colorless(Most of them)
a color dye used to visualize specific structures or cells on a smear when veiwing thru a microscope
Negative (background) stain
a staining procedure that results in color-less (clear, unstained) bacteria against a dark stained background
-the color-bearing ion in a stain
-the molecule that colors
What is used for negative staining?
Nigrosine and India ink
In the negative stain methods, why doesnt the bacteria stain?
If the Chromophore is negatively charged(anionic), it is repelled by the bacteria which is also negatively charged due to its nucleic acid(20%of its volume)
What is negative staining useful for?
Determining a cell's true morphology
organisms are allowed to dry on a glass microscope slide which is then passed thru a Bunsen burner flame
-this partially melts the cell walls
What are the 3 functions of heat fixing?
1.the cell sticks, "Fixed" to the side of the slide and will not wash off
2.The organism is killed
3.The organism stains better
What does heat fixing do to the cell?
-removes water from the cell
-it shrinks and becomes distorted
-doesn't allow one to visualize the cell in their natural state
Why is negative staining the most accurate method?
Because there is no-heat-fixing. no distortion or shrinkage occurs.
What is bacteria's morphology?
-single dye which is added to a smear
-a one-step staining process
-the use of a single stain to color a bacterial organism
The chromophore is postively charged (Cationic), it is attracted to and will only stain the slightly negative charged bacteria
What charge is the cell walls of bacteria?
Negatively charged and are stained by basic stains
What is commonly used for a simple stain?
Methylene blue-a basic stain
When do you use a basic stain?
is used when information about cell size,shape and arrangement (Morphology) of the bacteria is wanted
Microorganisms that grow and multiply in or on a culture medium.(solid or liquid)
any nutrient substance that can support microbial growth (solid or liquid)
Many different microorganisms growing together on a culture medium
-commonly found in nature
-especially on the body,soil water and food
only that one specific species (1 type of mcroorganism) rowing on the culture meium
-rare in nature
-usually occuring under extreme environmental conditions
Where does bacteria usually exist?
-In mixed populations
-rare situations that bacteria occur as a single species (a harsh environmental niche)
the process of separating an individual microoranism from a mixed poulation
Why is isolation important?
To be able to study the:
characteristics of an individual species
List the 6 reasons why it is necessary to have a Pure culture
1.The germ theory specifically calls for a pure culture and Koch's postulates were done to create them
2.To identify your unknown bacteria you need to have and maintain a pure culture
3.Needed for identifying specific spoilage and pathogenic microorganisms
4. Pasteur needed them to explain fermentation
5.Needed to test for water quality and safety
6.Needed to determine the quality and safety of food and milk
A visible mass of bacterial cells on the surface of solid media
only on solid media
-unique for individual species
-colonies produced by different bacterial species vary in their characteristics (size,shape color texture of the colony)
How is a pure colony made?
-made by aseptically transferring a small inoculum to a fresh medium and using 1 or 2 common methods
The microorganisms that are introduced to a sterile medium
What are the 2 commonly used methods to obtain a pure colony by isolation?
1.Streak Plate Method
2.Pout plate Method
Streak Plate Method
Physically spreading out cells over the surface of solid media far enough so that individual cells can grow into pure colonies
Pout Plate method
The demonstation of pH lowering becomes evidence of susceptibility to dental caries
List the facts that make the isolation of a pure culture possible.
-as bacteria grow and multiply on the surface of solid media they form clumps of bacteria called colonies
-each colony consists of a mass of cells that arise from a single bacteria cell = Binary fision
-Colony Morphology occurs
Snyder Test Agar
Used in the pour plate method
-relies on the rapidity of microorganisms in the salvia to lower a pH in a medium
-contains: 2% dextrose(sugar), Bromcresol green (pH Indicator)
-Decalcification of enamel begins at pH 5.5 and progresses rapidly as th pH is lowered to 4.4 and less
- Results: Green=negative (pH 4.8)
Yellow=positive (pH 4.4 and below)
The degradation of tooth enamel and dentin in the formation of tooth decay
How are dental caries formed?
Occurs as a result of the production of lactid acid by bacteria in the prence of high levels of sucrose
Information about dental caries
-considered a bacterial infection and a chronic disease
-the most widespread category of infection worlwidw
-microbes present in salvia and tooth plaque produce acid which can disolve teeth and lead to tooth decay
-The acidogenic potential of these microbes is greatly enhanced by sucrose
What are the 2 main bacterias that cause tooth caries?
-is the microorganism most frequently isolated from the surface of decayed teeth
-they not only produce acid but also have a high tolerence for acid accumulation in their environment
are also commonly involved in tooth decay
-they not only produce acid but also have a high tolerance for acid accumulation in their environment
What are the 2 populations of bacteria on the skin called?
1.Transient (contaminant) Microorganisms
2.Resident (Normal microbiota) microorganisms
-Not permanent skin residents
-7-8 minutes of hand washing will remove them
-easily removed from skin and transmitted to fomites
-only present on skin for a few hours to a few days
-sensitive to antiseptics
-only slowly removed by hand washing
-permanent skin residents, fimly entrenches in the skin
-less sensitive than the transient microbes to antiseptics
-low pathogenic staphylococcus species
a habitat supplying the factors necessary for the existence of an organism or species
Why is the skin an excellent ecological niche?
1.Sweat (sudoriferous) glands produce persiperation which contains nitrogeneous wastes (urea, uric acid amonia) and nutrients (lactic acid) and salts
2.Oil (Sebaceous) glands produce sebum, an oily subs. composed of lipids and fattey acids
3.pH of the skin is normally between pH 4-6. this is inhibitory to some micobes. (fatty acids are toxic
What degree does the agar soldify?
Agents that physically remove microorganisms from a limited area of the skin.
ex: Alocohol swabs, water, soap & water combo
-cocci round in grape like clusters
What are the 2most important Staphylococcus
-a normal skin bacteria
-(white dots on an agar plate= normal ones)
-MOST SIGNIFICANT PATHOGEN OF THE STAPH. SPECIES
-The medically important staph
-normally found on the skin and nasal membranes
-causes skin infections, throat infections, Stapylococcal food poisoning and toxic shock syndrome
-(Yellow color on agar plate)
List the mixed infections that Strptococci and Staphylococci cause
(Per professor only need to know names not definitions)
G+ pus-forming cocci
Collectively Staphylococci and Streptococci are refereed to as th pyogenic bacteria
Staphylococci are usually found where?
All 19 species of Staphylococci have what in common?
-irregular , grape like cluster
What 3 things do Streptococci have with Staphylococci
What are the 4 unique characteristics of Staph. Aureus that can be used to isolate and identify them
1.on the Solid Media they are YELLOW
2.They can grow on a selective media containing 10% salt NaCl=salt tolerant
-The ONLY one to ferment the sugar mannitol to produce acid
-Cause Beta-hemolysis on a blood agar plate
The complete destruction of red blood cells on a blood agar plate
-appears as a clear zone on the blood agar plate
-Bacteria produce the enzyme, hemolysin
Partial destruction of red blood cells on the bloos agar plate
-Appears as a brownish-green zone on a blood agar plate
Staph. Epidermis does this
Staph Aureus bacteria can coagulate rabbit plasma ( turns it to a clear gel
Staph. Epidermis cannot turn the rabbit plasma clear
-a microbiological growth media that contains chemicals (slat) that prevent the growth of unwanted bacteria
-but allows for the growth of the wanted species of bacteria
-it is used when isolating a specific microorganism for a mixed culture
-A microbiological growth media that indictaes the presence of a specific microrganism usually by a COLOR CHANDE due to some unique product or metobolic step of that 1 paticular species of bacteria
A postive reaction in a Differential media is?
A color change in the medium after inoculation and incubation
Mannitol Salt Agar
-is a selective and differential media
What does Mannitl salt agar contain
-Mannitol- the sugar that only the Staph Aureus can ferment to produce acid (Differental)
-7.5% NaCl-this salt concentration inhibits other microbes except the salt tolerant Stapylococci (Selective)
-Phenol Red- A pH indicator-has a range of 6.8 to 8.4
**basic=red=pH above 7
**neutral=red= pH 7
**acid =yellow=Below pH7
ONLY STAPH. AUREUS DOES THIS
-While there are other bacterias besides staph that ferments Mannitol, they wont on an MSA plate because of the Salt concentration
-Yellow color means acids were formed and ONLY by the salt tolerant mannitol fermenting STAPG AUREUS
How do Streptococci and Staphylococci differ?
The Strepto: are arranged in a chain-like pattern
List the 6 things Steptococci alone causes
1.pneumonia:disease of the lungs
2.meningtis:bacterial disease inflammation of the meninges
3.endocarditis:inflam. of the lining of the heart & valves
5.erysipelas:acute disease, fever,swellig, inflamation of skin
6.glomerulonephritis;inflamation of the capillaries in the renal area
Where is Streptococci normally found on the body?
-surface of the teeth
**The medically important species of Strepococci
**this one only causes Beta-Hemolysis(clear zone on blood agar plate= +
As a group Strptococci are responsible for?
-More illnesses (total amount)
-causing a greater variety of disease than any ohter group of Bacteria
Safe for drinking
Bacteria in water:
-water is seldom pure in nature
-water that contains large numbers of bacteria may still be safe to drink
-it the specific Human Pathogens present in water Not the number of microorganisms that determine its quality
Dysentary, typhoid fever cholera; spread by the Fecal0Oral route through contaminated water
-not normally found in water; unless contaminated by sewage
from the word Colon. man made group
Main bacteria in this group is E. coli
-a sewage indicator organism
What do all the bacterial spieces of the Coliform group have in common?
3.Non -endospore formers
4.Bacilli (rod shape) & randon arrangement
5. Ferments lactose sugar to produce gas***important
-A postive presumption test tubes
-a small inverted gas collecting tube in lactose broth
-it indicates the presence of gas by a small air bubble (+) trapped within it
Why is E. Coli the standard sewage indicator? this is a must know question per professor
1.It is not normally present in water or soil
2.it is only found in the large intestines of warm-blooded animals and humans
3.it is easy to identify by using the slective/differental media (lactose broth)
4.it survives longer in water than many enteric pathogens so it makes them easy to identify and not to be missed
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