2: Cultivation of Bacteria Flashcards
(67 cards)
1
Q
raw materials required for metabolism and reproduction
A
nutrients
2
Q
events used in biosynthesis and energy production
A
nutrition
3
Q
2 types of nutritional requirements for living organisms
A
mineral nutrients (micro-nutrients and macro-nutrients)
growth factors
4
Q
give 5 examples of micro-nutrients
A
cobalt
copper
manganese
molybdenum
nickel
selenium
tungsten
vanadium
zinc
5
Q
give 5 examples of macro-nutrients
A
oxygen**
hydrogen**
carbon
nitrogen**
phosphorous
sulfur
potassium
magnesium
sodium
calcium
iron
carbon**
carbon, nitrogen, and water as needed in highest quantities
6
Q
3 groups of growth factors
A
amino acids
purines and pyrimidines
vitamins (riboflavin, biotin, niacin, B6, K, thiamin etc.)
7
Q
2 classifications of microorganisms according to CARBON source
A
autotroph and heterotroph
8
Q
- producers,
- photosynthetic
- use CO2 and H2O
- sunlight as energy
A
autotrophs
9
Q
- require preformed food
- digestive and absorptive
- most microbes
A
heterotrophs
10
Q
2 classifications of microorganisms according to ENERGY source
A
chemotrophs
phototrophs
11
Q
- obtain energy by oxidizing electron donor via respiration (i.e., oxidative phosphorylation and others)
A
chemotrophs
12
Q
- capture protons in order to acquire energy via phototrophy (i.e., photophosphorylation etc.)
A
phototrophs
13
Q
4 types of organism in general (combination of energy and carbon source)
A
photoautotroph
- energy from light
- carbon from CO2
chemoautotroph
- energy from oxidation of inorganics
- carbon from CO2
photoheterotroph
- energy from light
- carbon from preformed organic food
chemoheterotroph
- energy from oxidation of organics
- carbon from preformed organic food
(see module for diagram)
14
Q
2 classification of microorganisms according to ELECTRON source
A
lithotrophs - reduced inorganic compounds as electron donors
organotrophs - organic compounds as electron donors
15
Q
4 physical requirements for microbial growth
A
temperature
pH
oxygen and carbon dioxide
hydrostatic pressure
16
Q
- most important factor that determines the rate of microbial growth, multiplication,
survival and death of all microorganisms - directly proportional to rate of enzyme reaction
A
temperature
17
Q
3 cardinal temperatures
A
minimum - lowest temp. at which organisms grow
optimum - highest growth rate
maximum - highest temp. at which growth occurs
18
Q
effects of temperature on the following levels:
- very low temperature
- lower than optimum temperature
- higher than optimum temperature
- very high temperature
A
very low temperature - very slow metabolic rates, cells survive longer
lower than optimum temperature - faster enzymatic rates and growth
higher than optimum temperature - DNA, RNA, and other cell components are irreversibly denatured, growth rate drops to zero
very high temperature - microorganism dies
19
Q
4 classifications of microorganisms according to TEMPERATURE
A
psychrophiles/cryophiles
mesophiles
thermophiles
hyperthermophiles
20
Q
- microorganisms that are able to grow at -20° C to 10° C
- e.g., Pseudomonas, Flavobacterium, and Alcaligenes
A
psychrophiles/cryophiles
21
Q
- grows best in moderate temperatures
- from 20° C to 45° C
- most microorganisms
- e.g., E. coli, Streptococcus pneumoniae, etc.
A
mesophiles
22
Q
body temperature pathogenic to humans
A
37° C
23
Q
- heat-loving microorganisms
- min: 45° C
- optimum: 50° C to 80° C
- most prokaryotes in volcanic areas
- e.g., Thermus aquaticus, Geogemma
barrosii, etc.
A
thermophiles
24
Q
- thrives in extremely hot environments
- from 80° C to 113° C
- CM has high levels of saturated fatty acids to retain shape
- e.g., Sulfolobus solfataricus, Methanococcus jannaschii, Thermotoga, etc.
A
hyperthermophiles
25
- the negative logarithm of Hydrogen ion concentration
pH
drastic variations of cytoplasmic pH disrupts enzyme activities and membrane transport protein activities (leading to inhibition)
26
4 classifications of microorganisms according to pH
acidophiles: 0 - 5.5
neutrophiles: 5.5-8.0
alkalophiles: 8-14
27
identify the pH range for:
- optimum microbial growth
- most bacteria
- molds and yeasts
optimum microbial growth: 6-8
most bacteria: 4-9
molds and yeasts: 5-6
28
2 principal gases that affect the growth of microbial cells
oxygen and carbon dioxide
29
4 classifications of microorganisms according to OXYGEN requirement
**aerobes - survive and grow in an
oxygenated environment (standard air: 21%)
**anaerobes - poisoned by and do not use oxygen
- types:
a. tolerant anaerobes - tolerant to low oxygen concentrations (can be highly or moderately oxygen tolerant)
b. Strictly anaerobes - killed by brief exposure to oxygen
** facilitative anaerobes - do not require oxygen for growth although may use it for energy production if available (e.g., via fermentation)
- e.g., E. coli and yeast
**microaerophiles - grow best at 1-15% oxygen levels (intolerant to normal air with 21% oxygen)
- high susceptibility to superoxide radicals and hydrogen peroxide
30
pressure exerted on the cells by the
movement of water resting on top of them
hydrostatic pressure
31
- pressure-dependent microbes that need a high-pressure environment in order to grow
- e.g., piezophiles
barophiles
32
method of multiplying microorganisms by letting them grow and reproduce in artificial
culture media under controlled laboratory conditions or set-up
bacterial cultivation
33
inoculating the bacteria in a
pre-determined culture medium and
growing in a laboratory controlled
environment
in vitro culture
34
nutrient material prepared for the growth of microorganisms in a laboratory
culture media
35
when microbes are introduced into a culture medium to initiate growth
inoculum
36
microbes that grow and multiply in or on a culture medium
culture
37
3 main purposes of bacterial cultivation
To grow and isolate all bacteria in an infection.
To identify cause of infection and contaminants or colonizers.
To allow identification and characterization of bacteria.
38
7 basic resources in culture media
- distilled water (70-85%)
- carbon
- energy (0.5%) (in form of nitrate, sulfide, or glucose)
- nitrogen
- mineral
- growth factors (AAs and vitamins)
- Oxygen, moisture, pH, and temperature
39
5 components of culture media
1. agar (or agar-agar) - long chain polysaccharide
2. peptone - complex mixture of partially digested proteins
3. yeast extract - source of growth factors
4. malt extract - maltose, starch, dextrins, glucose
5. blood and serum - from human or animal for enriching (10% concentration)
40
3 types of culture media according to PHYSICAL STATE or CONSISTENCY
a. liquid media - peptone and NaCl as main ingredients e.g., broths
b. solid media - agar into liquid media
- for isolating bacteria and determining characteristics of colonies
c. semi-solid media - 0.5% or less agar for cultivation of microaerophilic bacteria and determining their motility
41
3 types of culture media according to their chemical composition
simple media - general purpose; basic nutrients
complex media - common ingredient: peptone (some ingredients with unknown composition)
synthetic or chemically-defined media - from pure chemical substances with known composition
- for special studies such as metabolic requirements of microorganisms
42
8 types of culture media according to FUNCTION
basal media
enriched media
enrichment media
selective media
differential media
indicator media
transport media
anaerobic media
43
- simplest and most common medium in diagnostic laboratories
- for non-fastidious bacteria
- e.g., nutrient agar, nutrient broth, peptone water
basal media
44
- addition of substances like blood, chocolate, or serum to a basal medium
- for fastidious bacteria
- e.g., blood agar, chocolate agar, Lowenstein-Jensen agar, Bordet-Gengou agar, Loeffler's serum slope, and Dorset's egg medium
enriched media
45
- usually liquid medium that inhibits
the growth of unwanted bacteria and favors the growth of wanted bacteria
- e.g., tetrathionate broth, selenite F (feces) broth, and alkaline peptone water
enrichment media
46
- solid medium which inhibits the growth of unwanted bacteria but favors the growth of wanted bacteria
- e.g., DCA agar, TCBS agar MacConkey, Tellurite
selective media (isolating from mixed culture)
47
- reagents or supplements which when incorporated into culture media may allow differentiation of various kinds of bacteria
- e.g.,
blood agar (both differential and enriched) - differentiates hemolytic and nonhemolytic bacteria
Nagler's medium - for lecithinase activity
MacConkey agar (differential and selective) - for inhibiting growth of gram-positive bacteria
differential media
48
- media with indicator which changes color
- e.g., Wilson-Blair medium, potassium tellurite, and MacConkey agar
indicator media
49
- holding medium designed to preserve the
viability of microorganisms in the specimen but not allow multiplication
- e.g., Cary-Blair media, Amies media, Stuart media
transport medium
50
- media are used to grow anaerobic
organisms and contain reducing substances
- e.g., Thioglycollate broth and Cooked meat broth or RCM broth
anaerobic media
51
6 types of aerobic culture methods
Streak Plate Culture
Lawn Culture
Stroke Culture
Stab Culture
Pour Plate Culture
Liquid Culture
52
- aerobic culture method routinely employed for the isolation of bacteria in pure culture from clinical specimens
quadrant streak plate method
53
- aerobic culture prepared by flooding the surface of the plate with a liquid culture or suspension of the bacterium, pipetting off the excess inoculum and incubating
the plate
lawn culture method
54
- aerobic method: slopes are
seeded by lightly smearing the surface of agar with loop in a zigzag pattern taking care not to cut the agar
stroke culture method (same as our first inoculation in vmcb lab)
55
- aerobic method: nutrient gelatin or glucose agar is punctured with a long, straight, charged wire into the center of the medium and withdrawing it in the same line to avoid splitting the medium
stab culture method
56
- aerobic method: sample is spread evenly over surface of agar using sterile glass spreader
- results to surface colonies only
spread-plate method
57
- aerobic method: sterile medium is added and mixed with inoculum
- results to both subsurface and surface colonies
pour-plate method
58
- aerobic method: inoculated
by touching with a charged loop or by adding the inoculum with pipettes
or syringes
liquid culture method
59
2 disadvantages of the liquid culture method
does not provide a pure culture from mixed inocula—the major disadvantage
identification of bacteria is not possible
60
these microorganisms will not grow from small inocula unless oxygen is absent and the pH of the medium is low
obligate anaerobes
61
6 methods of anaerobiosis
- Production of a vacuum
- Displacement of oxygen by other gases
- By absorption of oxygen by chemical or biological methods
- By displacement and combustion of oxygen
- By reducing agents
- Other anaerobic culture systems
62
anaerobic method: attempted by incubating cultures in a vacuum desiccator, but proved to be unsatisfactory because some oxygen is always left behind
production of a vacuum
63
anaerobic method: inoculated plates are placed inside a large airtight container and a lighted candle kept in it before the lid is sealed
Displacement of oxygen by other gases (via candle jar)
64
- anaerobic method: the method of choice for preparing anaerobic jars
- in the form of disposable packet of aluminum foil containing pellets of sodium borohydride and cobalt chloride and of citric acid and sodium bicarbonate
Gaspak (absorption of oxygen by chemical or biological methods)
65
- anaerobic method: jar used to displace and combust oxygen
Mcintosh-Filde's anaerobic jar
66
- anaerobic method: use of ***** agents whose effectiveness can be increased by making a liquid medium semisolid with agar 0.05 to 0.1 percent
- e.g., thioglycollate broth, cooked meat broth (CMB) or Robertson's cooked meat (RCM)
by reducing agents
67
5 ways to preserve bacterial cultures
refrigeration - short-term
deep freezing - quick-frozen, last several years
lyophilization (freeze drying) - quick-frozen then vacuumed to remove water
cold storage
drying methods
