Microbiology 1 Flashcards
1
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Microbiology
A
the study of microscopic forms of life
2
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Microscopic groups
A
bacteria, fungi, protazoa, viruses
3
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Pathogenicity
A
certain mircobes cause disease, learning how diseases are transmitted is important to health sciences.
4
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Bioremediation
A
process using naturally occurring or genetically engineered microorganisms to degrade or detoxify environmental hazards (ex. oil spills)
5
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Microbes biological roles
A
are essential to web of life, some are photosynthetic, others decompose dead organisms and some provide nitrogen to plants
6
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Fundamental biology
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microbiology provides insight into life processes of all life forms
7
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Microbes
A
are useful in research, are prokaryotes
8
Q
antibiotics
A
can be isolated from microbes (ex penicillium notatum)
9
Q
genetic engineering
A
microbes can be used to create useful human proteins (ex insulin and HGH)
10
Q
Characteristics of microbes
A
ubiquitous, microscopic, unicellular (some are multicellular), sexual and/or asexual reproduction, growth in number, metabolic activity, free living (some are parasitic), respond to stimuli, mutations, range in size from 20nm to 100um
11
Q
Domains
A
Domain Archaea (extremeophiles), Domain Bacteria, Domain Eukarya
12
Q
Microbe types
A
Bacteria, Algae, Fungi, Viruses, Protazoa
13
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Bacteria
A
are unicellular, microscopic, some are photosynthetic, no nucleus, have 3 types of morphology
14
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Bacteria shapes
A
round, rod, tight spiral, loose spiral, comma shaped
15
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Round bacteria
A
cuccus, cocci
16
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Rod shapes bacteria
A
bacillus, bacilli
17
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Tight spiral shaped bacteria
A
spirochetes
18
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Loose spiral shaped bacteria
A
spirillum
19
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Comma shaped bacteria
A
vibrio
20
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Algae
A
unicellular or multi-cellular, eukaryotic
21
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Fungi
A
kingdom fungi, in domain eukaria, unicellular yeasts, multi-cellular molds, macroscopic (mushrooms), are an important source of antibiotics
22
Q
Mushrooms
A
absorb their nutrients and decompose, no photosynthesis
23
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Viruses
A
not living things, need host cell to reproduce, are more like complex chemicals, have neuclaic acid genome and protein capsid-“neucleoproteins”, obligate intracellular parasites
24
Q
Protazoa
A
Eukaryotic organism, has many vacules and organelles, are motile, ameba
25
Robert Hooke & Anton Van Leeuwenhoek
Invented the first microscope, first to see living organisms under microscope, 1600's. Hooke coined term "cell"
26
Carolus Linneaus
developed binomial nomenclature by using genus and species. Genus is capitalized, species is not, 1735
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Compound light microscope
system of magnifying lenses arranged to produce an enlarged image of an otherwise invisible object, has two lenses (ocular and and objective)
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Theodore Schwann
developed the cell theory
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Cell theory
cells are the fundamental units of life and carry out the functions of living things
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Germ theory of disease
microorganisms can invade other organisms and cause disease
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Spontaneous generation
the belief that life can spontaneously arise from non-living matter
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Redi
1668, disproved the theory that maggots come from meat
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Needham
boiled chicken broth and put it into different containers to prove spontaneous generation for microbes in 1745
34
Spallanzani
repeated Needham's experiment but containers, no bacteria formed, critics said it was because of lack of oxygen
35
Pasteur
1861, disproves spontaneous generation and proves biogenesis, also developed aseptic technique, pasteurization, rabies vaccine, fermentation
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Biogenesis
all life comes from living things
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Robert Koch
1884, developed pure culture technique
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Koch's postulates
1. the specific causative agent must be found in every case of disease
2. the disease organism must be isolated in pure culture
3. inoculation of the culture into healthy susceptible animal must produce the same disease
4. the disease organism must be recovered from the inoculated animal
39
Semmelweiss
1840's, hand washing, pioneer of aseptic technique
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Mortality and handwashing
handwashing use lowered mortality rate fom 10-35% to 1% in maternity ward
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Lister
1860's developed use of chemical disinfectants in surgery
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Alexander Flemming
1928, discovered penecillin
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Chemotherapy
treatment of a disease with a chemical
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Antibiotics
microbe made chemicals
45
Synthetics
man made chemicals
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Microscpoe
a system of magnifying lenses arranged in an order to provide a visible image of an otherwise non-visible object
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Polio Virus size
20nm (0.02um)
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Small Pox Virus size
300nm (0.3um)
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Staphylococcus aureus
0.5um, typical coccus shape (round)
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Pseudomonas aeruginosa
5um long, typical bacillus shape (oblong)
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Treponema pallidum
20um long, very thin, spirochete shape
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Resolution
R= λ / 2 x NA
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NA
numerical aperture, a mathematical expression relating to the extent that light is concentrated by the condenser lens and collected by the objective
54
λ
wavelength, is directly proportional to resoulution, as the wavelength decreases, the resolving power increases
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Important features of a microscope
magnification, resolution
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Resolution
smaller numbers make a clearer image, is a number that indicates the smallest microbe or part of a microbe that can be viewed with clarity
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Light microscopy
uses light as the illuminating source, uses glass lenses, λ=550nm
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Electron microscopy
uses electrons as illuminating source, uses electromagnetic lenses, λ= 0.005nm, has 100,000 fold resolution than light microscopy
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Light properties
light can be reflected, transmitted and absorbed, we want to avoid refraction
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Index of refraction
speed at which light passes through material
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Microscope types
compound bright field (compound light microscope), dark field, phase contrast, fluorescence, electron
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Compound bright field
field of view is brightly lit, magnification=2,000x, resolution with oil objective=0.2um
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Compound bright field uses
clinical, research, fixed and stained microbes
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Compound bright field limitations
poor image production of viable microbes, can not see anything under resolution of 0.2um
65
Dark field microscope
light reflected off surface of specimen to view, background is dark, magnification=2,000x, resolution with oil objective=0.2um
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Dark field uses
image production of viable microbes that are not easily stained or cultured in vitro
Major use: diagnosis of tetranemapalidum (syphilis) which is a clinical use
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Dark field limitations
poor image production of classically fixed and stained microbes, can not see anything under resolution of 0.2um
68
Phase contrast microscope
dual beam-light comes from two sources
magnification=2,000x
resolution with oil objective=0.2um
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Phase contrast uses
highly contrasted images of internal structures of visible microbes including flagella and cilia
70
Phase contrast dual beam
can cause reinforcement or interference because light comes directly from light source and is also refracted off the specemin
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Phase contrast limitations
poor image production of classically fixed and stained slides, can not see anything under resolution of 0.2um
72
Fluorescence microscope
AKA ultraviolet or immunofluorescent, uses ultraviolet light, magnification=2,000x
Wavelength of UV light is between 100-400um making for better resolution
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Fluorescence microscope uses
look at microbes with natural fluorescence or those that selectively bind to flurochains
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Pseudomonas aeruginosa
is naturally fluorescent, best used with fluorescence microscope
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Mycobacterium tuberculosis
binds to auromine fluoricome and appears bright yellow under fluorescence microscope
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Bacillus enthracs
binds tofluorescin isothioyanate (FITC) to make apple green color under fluorescence microscope
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Acridine orange
binds to neulcaic acids like DNA
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Fluorescence microscope major use
immunofluorescence
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Fluorescence microscope clinical use
fluorescent antibody, used to diagnose rabies and syphylus
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Fluorescence microscope research
need to make an antibody and combine it with fluorocrome, can use fluorescent antibody to bind to a sample
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Electron microscopy
Transmission (TEM) and Scanning (SEM)
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Transmission microscopy
transmission electron microscope, magnification=100,000x, resolution=2.5nm (0.0025um)
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Transmission microscopy uses
research for fixed and stained microbes, no size limitation
84
Scanning microscopy
produces 3D image, magnification=10,000x, resolution=10nm
| displaces electrons from the surface of cells, secondary electrons are responsible for creating shape
85
TEM slide prep
takes days to prep slides which keeps use limited
1. grow culture of bacteria overnight
2. fix bacteria in "aldahide" substance
3. sequential dehydration using alcohol
4. embed cells into a resin
5. thin sections of resin using microtome (sections are 10-50nm)
6. float sections onto copper mesh where sections are stained using heavy metal
86
Transmission microscopy limitations
preparation can cause distortion of the sample causing mesosomes which are artifacts
87
Microscope slide techniques
Wet mount
smears
staining
hanging drop-is good for viewing motility
88
Simple stain
used to see morphology and arrangement
| ex: crystal violet
89
differential stain
can differentiate organisms and place microbes into major taxanomic groups
90
gram stain
type of differential stain, can see composition of cell wall, can be gram positive or gram negative
91
acid fast stain
trying to detect certain cell wall component (myloric acid)
| Only myobacterium and nocardia are acid fast bacteria
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Special stain
try to detect presence and/or location of certain structure
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acidic dye
is a special stain, dye molecule is associated with the negatively charged ion. It is a negative stain.
Won't stain bacteria because they already have a net negative charge
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Negative stain
is a special stain, will stain the background and then you have to go back and stain the bacteria
95
Flagellar stain
is a special stain, can see flagella, very difficult stain to achieve, can see presence and location of flagella
96
Endospore stain
is a special stain, used to identify the presence of endospores
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Taxonomy
the classification of bacteria into major groupings based on observable, measurable traits
Similarities are presumably due to relatedness
98
systemats
the study of the evolutionary relationship of organisms, can be proven by DNA sequencing
99
5 kingdom classification
plantae, fungi, animalia, protista, monera
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Domain eukaria
contains, plantae, fungi and animalia
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Kingdom protista
no longer recognized, is being broken down into additional kingdoms using rRNA sequencing
organisms in this kingdom are very nutritionally diverse, contains all unicellular animals and unicellular plants
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Kingdom monera
no longer recognized, contained all bacteria and archea, is divided into domain archea and domain bacteria
103
Prokaryotic species
population of cells with similar characteristics because they are all descendants of a single parental cell
clonal population=pure culture
104
Bacterial strain
ex: E. Coli 0157H7
Numbers and letters indicate strain type
Strains are created by picking up extra genes
105
Growth media function types
Liquid (broths)
solid (agar plates, slants, deeps)
semi-solid (tubes)
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General purpose media
contains nutrients that allow certain types of bacteria to grow
ex: TSA (trypticase soy agar)
107
Selective media
contains a selective agent that only allows certain types of bacteria to grow
ex: MSA, Sabdex
108
MSA
mannitol salt agar, contains a high concentration of salt, is selective for certain organisms
109
Sabdex
high dextrose concentration, low pH which favors the growth of fungi over bacteria making it selective but it is also a general purpose media for growing fungi
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Enriched media
contains specific growth factors and/or vitamins that needed by bacteria that can't grow on general purpose media
ex: blood agar
111
Differential media
contains a constituent that causes an observable change (in color or pH) in the medium when a particular chemical reaction occurs
this makes it possible to distinguish between organisms
112
MSA
type of growth media, contains mannitol, salt and phenol red (pH) indicator
pH>6.8=red
pH<6.8=yellow
113
Magnification
increased size of a virtual image
114
Resolution
increased clarity or definition of detail or virtual image
115
Light microscopy
refers to microscopy that uses visible light to observe specemins
116
Image production pathway
illuminator
condenser (which directs light through the specimen)
objective lens(which magnifies the specimen)
occult lens (magnifies the specimen again x10)
117
Total magnification
Objective lens X ocular lens
118
Working distance
distance between object on the slide and the objective
119
Refractive index
refers to the light bending properties of differential substances
120
Oil immersion
used to reduce refraction, has same refractive index of glass, acts as an extension of the objective lens
121
Smear slide
before we stain any slide we need to prepare the smear
122
Smear slide objective
bind cells to the slide
inactive cells
facilitate stain penetration
123
Measuring microbes
at 100x distance between 2 lines is 1um
| at 40x distance between 2 lines is 2.5um
124
Scientific method
```
observation/research
hypothesis
prediction
experimentation
results
conclusion
```
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Observations
things we see in nature
hear/read about
possibly research the topic
126
Hypothesis
provides a solution to our observations
an"educated guess"
a good hypothesis is testable
127
prediction
hypothesis can take the form of prediction
uses and if-then statement
allows one to specify how the hypothesis will be tested
128
experiment
testing your hypothesis
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Independent variable
intentionally changed
130
dependent variable
the value we measure from the experiment
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Controlled variables
are held constant
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Control treatment
should vary from the experimental group by only one variable or it will be hard to determine what caused results
the independent variable is eliminated or set to a standard value
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Controls
positive control
| negative control
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Positive control
an experiment whose results are known to be positive
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Negative control
an experiment whose result is known to be negative
136
Results
data should be presented in tables and graphs for analysis
| data should be able to "stand alone"
137
conclusions
based on results, either supports or does not support the hypothesis
you don't "prove" anything from just one experiment
138
Mushrooms
are not plants, they do not preform photosynthesis
139
Kingdom plantae
multi-cellular photosynthetic autotrophes (self nourishing)
140
Kingdom anamelia
heterotrophic consumers
141
Domain bacteria
all pathogens, also non pathogenic bacteria from soil and water
142
Domain archea VS domain bacteria
domain archea do not have peptidoglycan in their cell walls and domain bacteria do
143
Domain archea
Have 3 groups
Methanogens
Halophiles
Hyperthermophiles
144
Methanogens
make methane
are strict anerobes(need anerobic environment
CO2 + H2 --> OH4 (methane)
145
Halophiles
require high salt consentration
| obligate/strict halophiles
146
Hyperthermophiles
grow in extremly hot environments
| proteins have adapted and do not denature in heat
147
Taxonomic study steps
establish a pure culture
characterization of a pure culture
DNA hybridization
148
How to establish a pure culture
must use streaking, " streak plate method"
| EX: onto an agar plate
149
Characterization of a pure culture
```
microscopic characteristics
cultural characteristics
biochemical characteristics
chemical characteristics
genetic characteristics
```
150
Microscopic characteristics of a pure culture
use a gram stain to see (AKA differential stain)
151
Cultural characteristics of a pure culture
environmental factors like temperature, O2 required and pH required
colony characteristics like pigment, opacity, margins and elevation
152
Biochemical characteristics of a pure culture
best way to determine unknown
| uses dichotomous key (we will make our own)
153
Chemical characteristics of a pure culture
immunological, use ELIZA test
154
ELIZA test
enzyme linked immunosorbent assay
| used to determine chemical characteristics of microbes
155
Genetic characteristics of a pure culture
use ribosomal gene sequencing
156
Ribotyping ribosomes
composed of protein and ribosomal RNA, highly conservative (very few evolutionary change over time), all members of a certain domain, may have signature sequences
157
Facultative anerobe
can grow in oxygenated or non-oxygentaed environment unlike a strict anerobe
158
Klebsiella
large mucoid colonies
159
Genus protius
has swarming motility
160
Phenol red
if pH is > 6.8, it is a red color
| if bacteria ferments lactose to acid products, pH will drop and color is yellow
161
DNA hybridization
heat DNA to break hydrogen bonds between nucleotides
| only used to exclude relatedness, can't say things are related based on their G+C ratio
162
DNA composition
base composition is a fixed property
G base pairs with C, A base pairs withT
% of G+C gives you, in theory, A's and T's
163
DNA hybridization method
1. purify DNA from unknown
2. heat double strand of DNA until it is a single strand
3. monitor process using spectrophotometer
4. determine melting temperature
5. based on melting temp, we get % of G+C, compare % to other known organisms
164
DNA hydrogen bonds
A's and T's have 2 hydrogen bonds and melt at a lower temperature
G's and C's have 3 hydrogen bonds and melt at a higher temperature
165
Gram positive cell
has thick peptidoglycan layer
physically stronger
has small "pores"
166
Gram negative cell
```
has thin peptidoglycan layer surrounded by outer membrane
chemically stronger (outer membrane resists penetration of chemicals)
large "pores"
```
167
Gram positive cell stain
purple
| CV-I complex is larger than pores, alcohol dehydrates and further shrinks peptidoglycan, trapping purple stain
168
Gram negative cell stain
pink
alcohol dissolves the outer membrane and CV-I complexes wash out of larger pores
counter-stain pink with safranin
169
Gram type
differences in cell's wall
170
Gram reaction
the color it shows when stained
171
Gram-Variable
gram type and gram reaction do not always correlate
| technique and age of the culture can affect the reaction
172
KOH test
can be used to confirm gram stain test
gram-negative bacteria will lyse in the alkaline solution making it's DNA stringy
Gram positive cells will not lyse
