Chapter 2 Flashcards
1
Q
What are the 5 techniques used to manipulate, grow, examine, and characterize microorganisms in the lab?
A
- Inoculation
- Incubation
- Isolation
- Inspection
- Identification
2
Q
What is inoculation?
A
Implantation of microorganisms into or onto culture media
3
Q
What is an inoculum?
A
Sample of microorganism
4
Q
Instruments used for inoculation must be ________ to avoid contamination
A
Sterile
5
Q
What are 2 definitions of a culture?
A
- Visible accumulation of microorganisms in or on a nutrient medium
- Propagation of microorganisms with various media
6
Q
What are types of clinical samples that may be submitted for culture?
A
Body fluids, discharges, anatomical sites, diseased tissue
7
Q
What are examples of body fluids (clinical sample)?
A
Blood, cerebrospinal fluid, peritoneal fluid
8
Q
What are examples of discharges (clinical sample)?
A
Sputum, urine, feces
9
Q
What are examples of anatomical sites (clinical sample)?
A
Throat, nose, ear eye, genital tract
10
Q
What are examples of diseased tissue (clinical sample)?
A
Abscess and wound
11
Q
Besides clinical samples, what other samples can be tested?
A
Soil, water, sewage, foods, air, inanimate objects
12
Q
What is incubation?
A
Isolation of a sample growth in a temperature-controlled environment to encourage growth
13
Q
What range of incubator temperature may you find in a medical lab?
A
20-45 C
14
Q
What range of incubation may be needed for macroscopic growth?
A
1 day to several weeks
15
Q
What is media?
A
Nutrient used to grow organisms (namely bacteria and fungi) outside their natural habitats
16
Q
What kind of compounds may be required for organism growth?
A
Inorganic and/or organic
17
Q
Can all microbes be grown in a lab?
A
No
18
Q
Media is classified by what 3 properties?
A
- Physical state
- Chemical composition
- Functional type (purpose)
19
Q
What are the 4 media types?
A
- Liquid
- Semisolid
- Solid (can be converted to liquid)
- Solid (cannot be converted to liquid)
20
Q
What is agar?
A
Complex polysaccharide isolated from the alga Gelidium
21
Q
At what temperature is agar solid?
A
Room temperature
22
Q
At what temperature is agar liquid?
A
100 C
23
Q
What are key characteristics of agar?
A
It is flexible, moldable, and not digestible to most microbes
24
Q
What are the 2 types of chemical composition?
A
- Synthetic and chemically defined
- Complex and not chemically defined
25
What is synthetic media?
Contains pure organic and inorganic compounds that vary little from one source to another and has a molecular content specified by an exact formula; standardized and reproducible!
26
What is complex media?
Contains rich supply of nutrients from extracts from animals, plants, yeasts (can be sourced from ground-up cells, tissues, and secretions); type and amounts of nutrients vary from batch to batch, but is not a major problem
27
What are examples of extracts/bodily fluids used for complex media?
Blood, serum, meat extracts/infusions, milk, yeast extract, soybean digests, peptone
28
What are examples of complex media?
Nutrient broth, blood agar, MacConkey agar
29
What is the purpose of general purpose media?
Allow growth of a broad spectrum of microbes
30
What are examples of general purpose media?
Nutrient agar and broth, brain-heart infusion (BHI) agar, trypticase soy agar (TSA)
31
What is the purpose of enriched media?
Contains complex organic substances such as blood, serum, hemoglobin, or special growth factors (specific vitamins, amino acid) that certain species must have in order to grow
32
If a bacteria is considered fastidious, what does it mean?
Requires complex nutrients and growth factors to grow (found in enriched media)
33
What are examples of enriched media?
Blood agar (addition of sterile sheep, or rabbit blood to a sterile agar base), Thayer-Martin agar, chocolate agar (blood agar with hemin and nicotinamide adenine dinucleotide)
34
What do halos around colonies on blood agar indicate?
Blood hemolysis
35
Thayer-Martin agar can be utilized to grow what organism?
Neisseria
36
What is the purpose of selective media?
Contains one or more agents that inhibit the growth of certain types of microbe or microbes, but not others
37
What is selective media crucial for?
Isolation of a specific organism from a sample containing many different species (i.e., feces, saliva, skin, water, soil)
38
What are examples of agents used in selective media?
Bile salts (in media for isolating intestinal pathogens), dyes (methylene blue and crystal violet), and antimicrobial drugs
39
What is the purpose of differential media?
Contains chemicals that microbes react to, producing variation in colony size, colony color, media color, gas bubble and/or precipitate formation for differentiation, but does not inhibit growth
40
What are examples of agents used in differential media?
Dyes (pH indictors that change color when an acid or base is produced by the microbe)
41
Can media be selective and differential?
Yes!
42
What is the purpose of reducing media?
Contains a substance (sodium thioglycollate or cystine) that absorbs oxygen or slows the penetration of oxygen in a medium to reduce oxygen availability
43
What is reducing media helpful for?
Growing anaerobic bacteria (that don't require oxygen for growth) and determining oxygen requirements of isolates
44
What is the purpose of specimen transport media?
Maintain and preserve specimens that have to be held for a period of time before clinical analysis, or to sustain delicate species that die rapidly if not held under stable conditions
45
What is the purpose of assay media?
To test the effectiveness of antimicrobial drugs in clinical labs, and used to assess the effect of disinfectants, antiseptics, cosmetics, and preservatives on the growth of microorganisms by drug manufacturers
46
What is a pure culture?
Growth medium containing a single known species or type of microorganism
47
What is a mixed culture?
Growth medium containing two or more identified and easily differentiated species of microorganisms
48
What is a contaminated culture?
Growth medium once pure or mixed with known species, but has since had unwanted microbes if uncertain identity introduced
49
How may contamination occur?
If the lids of tubes or petri dishes are removed for too long or if the loop is not sterilized or handled properly
50
What is isolation?
The separation of microbial cells by serial dilution or mechanical dispersion on solid media to create discrete colonies
51
What is a colony?
Macroscopic cluster of cells appearing on a solid medium, each arising from the multiplication of a single cell
52
What are 3 main techniques used for isolation?
1. Streak plate
2. Pour plate
3. Spread plate
53
What is inspection?
Observation of macroscopic and microscopic morphology of growth in broth cultures or colonies on agar
54
Which level is usually determined and reported in the clinical lab?
Species
55
What methods may be utilized to identify microorganisms in the clinical lab?
Macroscopic/microscopic morphology, biochemical tests, immunological tests, and/or genetic analysis
56
What is the size range of macroscopic organisms?
Centimeters to meters
57
What is the size range of microscopic organisms?
Micrometers to millimeters
58
What are 3 important qualities of the microscope?
1. Magnification
2. Resolution
3. Contrast
59
Where are objective lens located?
Closest to the specimen
60
What kind of image does the objective lens form?
Real image
61
What is the typical range power of objective lens?
4x-100x
62
Where are the ocular lens located?
Closest to the eye
63
What kind of image does the ocular lens form?
Virtual image (resulting from the projection through the microscope)
64
What is the typical power of the ocular lens?
10x
65
The _________ image is received by the eye and converted to a _________ image.
Virtual, retinal/visual
66
How is the total magnification calculated?
Objective multiplied by ocular
67
What is resolution (resolving power)?
Capacity of an optical system to distinguish two adjacent objects or points from one another
68
What is the resolution of the human eye?
0.2 mm
69
What is the resolution of the oil immersion (100x objective) lens?
0.2 um
70
How does the oil immersion lens work?
Oil captures light that would otherwise scatter, increasing resolution
71
What is refractive index?
The measure of the degree of light that is bent/refracted as it passes from one medium (i.e., water or glass) to another medium (i.e., bacterial cells)
72
The higher the difference in refractive index, the more _______ of light and _______ the contrast that is registered by the microscope and eye
Bending, sharper
73
What is the iris diaphragm?
Adjustable control that controls the amount of light entering the condenser
74
What may too much light do to contrast?
Reduce it
75
What is the function of the condenser?
Gathers light from the light source into a cone
76
What can the lack of contrast of cell components be compensated for by?
Special lenses or adding dyes
77
What is the microscope field?
Circular area viewed through the ocular lens
78
How is light transmitted in a bright-field microscope?
Through the specimen, shows a dark specimen surrounded by an illuminated field
79
How is light transmitted in a dark-field microscope?
Around the specimen since a disc is added to the condenser and blocks the light, shows an illuminated specimen surrounded by a dark field
80
How does a phase-contrast microscope work?
Devices transform changes in light waves passing through the specimen into differences in light intensity, shows details of differing cell structures that have varying densities and thus alter the light passing through
81
What is the source of a fluorescence microscope?
Ultraviolet radiation
82
How does fluorescence microscopy work?
Fluorescent dyes and minerals are added to the specimen and the specimen is placed in contact with a source of fluorescence and UV radiation, shows image formed by light given off by specimen
83
What is the source of a confocal microscope?
Laser beam
84
What kind of samples can be observed using confocal microscopy?
Fluorescently stained or live unstained cells and tissue
85
How does confocal microscopy work?
Scans various depths in the specimen, shows sharp image focusing on a single plane at any level
86
What is transmission electron microscopy (TEM) used for?
Viewing detailed structures of cells and viruses
87
How does transmission electron microscopy (TEM) work?
Transmission of electrons through the specimen
88
What is scanning electron microscopy (SEM) used for?
3-D view of specimen
89
How does scanning electron microscopy (SEM) work?
Transmission of electrons onto the specimen, scanning back and forth
90
What are 3 considerations when preparing a specimen for microscopy?
1. Condition of specimen
2. Aims of the examiner
3. Type of microscopy available
91
What are examples of fresh, living specimen preparations?
Wet mounts and hanging drops
92
What does a wet mount and hanging drop allow the visualization of and why?
Movement; a suitable fluid is used to temporarily maintain viability and allow space for movement
93
How does one prepare a fixed, stained specimen smear?
Place a thin film of liquid suspension of specimen on a glass slide and let air dry prior to heat fixation
94
What does heat fixation allow?
Kill the specimen and adhere it to the slide
95
What is staining?
Procedure that applies colored chemicals (dyes) to specimens
96
How do dyes work?
They become affixed to cells through chemical reactions, giving them color
97
What is a basic (cationic) dye?
Positively charged dye that attracts negatively charged cell parts and more readily stains bacterial cells since they possess more negatively charged acidic substances on their surfaces
98
What is an acidic (anionic) dye?
Negatively charged dye that attracts positively charged cell parts, is generally repelled by cells and thus more suitable for negative staining
99
What is the difference between the background of positive and negative stains?
Background is not stained in positive stains, background is stained in negative stains
100
What is the difference between the appearance of specimens stained using positive and negative stains?
Specimen is colored by dye in positive stains, and specimen is colorless in negative stains
101
What are examples of basic dyes?
Crystal violet, methylene blue, safranin, malachite green
102
What are examples of acidic dyes?
Nigrosin, India ink
103
What are 3 subtypes of positive stains?
1. Simple stains
2. Differential stains
3. Special stains
104
What are 2 subtypes of negative stains?
Capsule and endospore
105
What are simple stains?
Single dye that stains all cells
106
What are differential stains?
Two different colored dyes called primary stain and counterstain that stain cells based on differing properties, and may require additional chemical reagents or techniques
107
What are 3 types of differential stains?
1. Gram stain
2. Acid-fast stain
3. Endospore stain
108
Who invented the Gram stain?
Hans Christian Gram
109
What color are Gram-positive cells?
Purple
110
What color are Gram-negative cells?
Pink/red
111
What color are Gram-variable cells?
Pink/red and purple
112
In what way are Gram-stains useful?
They can allow a headstart on antimicrobial treatment prior to obtaining susceptibility results
113
What color are acid-fast positive cells?
Pink
114
What color are acid-fast negative cells?
Blue
115
What was the acid-fast stain originally utilized for?
Detection of Mycobacterium tuberculosis
116
What does the endospore stain allow the the differentation of?
Vegetative cells and endospores
117
Members of what genera may form endospores?
Bacillus and Clostridium
118
What are special stains?
Stains that emphasize certain cell parts that are not revealed by conventional staining methods
119
What are 2 types of special stains?
1. Capsule
2. Flagella
120
What is the capsule stain utilized for?
Observing the microbial capsule that is a protective layer surrounding the cells of some bacteria and fungi which repel most stains
121
What is the flagella stain used for?
Observing flagella after deposition of coating on the outside for enlargement so they may be visible after staining to determine the presence, number and arrangement for bacterial identification
