Term
Microbial Nutrition and Metabolism Mixed Population definition |
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Definition
A population that contins two or more different microorganisms |
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Term
Microbial Nutrition and Metabolism Isolation definition |
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Definition
| the separation of a given microbe from all others in a mixed population |
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Term
Microbial Nutrition and Metabolism Cultivation definition |
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Definition
| the growth of microbes under well-defined laboratory conditions --> results in a culture |
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Term
Microbial Nutrition and Metabolism Medium definition |
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Definition
| A solution that contains, at the very least, those nutrients required for the growth of a microbe |
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Term
Microbial Nutrition and Metabolism Nutrient definition |
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Definition
any substance that is: (i) absolutely required for the growth of a microbe Or (ii) enhances growth of a microbe |
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Term
Microbial Nutrition and Metabolism Inoculation definition |
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Definition
| Seeding of sterile media in the laboratory |
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Term
Microbial Nutrition and Metabolism Solid medium definition |
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Definition
| One that contains a gelling agent, typically agar |
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Term
Microbial Nutrition and Metabolism Liquid Medium definition |
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Definition
| One that contians no gelling agent |
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Term
Microbial Nutrition and Metabolism Colony definition |
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Definition
| a macroscopic manifestation of microbial growth on solid medium |
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Term
Microbial Nutrition and Metabolism Why is agar the most ideal medium gelling agent? |
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Definition
1. Agar is nontoxic only serves as an inert substrate upon which growth occurs 2. Agar is nonnutritive for most microbes --> complex polysaccharide; monosaccharides could serves as carbon/energy souces for most microbes -Vast majority of microbes can't hydolyze agar -If they are able to degrade it, then other agents need to be used 3. Agar gels are stable over entire range of temperature at which microbial growth occurs |
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Term
Microbial Nutrition and Metabolism States of agar at different temperatures Why is this important? |
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Definition
temp above 44C --> liquid temp @ 44 or below --> solid Once solid --> stable until 100C Allows for different types of microbes to grow microbial growth range: -10C to 113C |
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Term
Isolation of Microorganisms What are the three major techniques What type of media(s) is(are) used |
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Definition
Major techniques: 1. Streak plate 2. Spread plate 3. Pour plate Media used: solid - liquid not used because procedures are too "iffy" |
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Term
Isolation of Microorganisms Streak Plate Technique |
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Definition
- acts to dilute the inoculum - growth becomes progessively less dense as distance from the start point increases - Direction 1 (20-30 streaks) - Direction 2 --> rotated 90 -->extended from stop point of direction 1 [image] |
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Term
Isolation of Microorganisms Spread Plate Technique |
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Definition
-suspension of bacteria is diluted in liquid media -100uL sample is spotted at the center of the agar -spread over the surface using glass "hockey stick" --> sterilized by 1/3L in EtOH, bunsen burner |
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Term
Isolation of Microorganisms Pour Plate Technique |
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Definition
- not used very often anymore - flask w/ 30mL of agar maintained at 45C - diluted sample of bacteria is aseptically added - swirled and then emptied into a Petrie Dis - bacteria grow w/ different morphologies at different depths <-- NOT GOOD! |
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Term
Isolation of Microorganisms Shake culture |
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Definition
inoculated medium is shaken to facilitate gaseous exchange between the medium and the atmosphere - done w/ only liquid media |
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Term
Isolation of Microorganisms Test tube Cultures |
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Definition
often referred to as "deeps" done in a test tube use liquid/solid media two types which use solid: 1) stab culture 2) slant culture |
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Term
Isolation of Microorganisms Static Culture |
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Definition
inoculated medium not shaken Done w/ solid media but can be employed with liquid as well |
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Term
Isolation of Microorganisms Stab culture |
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Definition
| culture is inoculated by stabbing down through the solidified medium |
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Term
Isolation of Microorganisms Slant culture |
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Definition
roughly 10mL of agar, slanted on tube "deep" does not apply here even though it is a test tube culture |
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Term
Sterilization Aseptic Technique definition |
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Definition
| Media, glassware, and platicware must be sterile until purposefully brought into contact witht eh microbes being propagated |
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Term
Sterilization Aspects to Aseptic Techniques |
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Definition
1. glass pipettes are sterilized in metal canisters --> bottom 1/2 is passed through bunsen burner --> plastic: sterile and used once 2. Pipettes never handled by tips 3. Inoculating wires/loops heated to red 4. Inoculate media in Petrie Dish, lid is raised as little as possible |
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Term
Sterilization Sterilization Techniques How many What are they |
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Definition
There are four basic types of sterilization procedures 1. Heat 2. Certain chemicals 3. Cold filtration * 4. Radiation *method of choice for liquid media |
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Term
Sterilization Heat definition |
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Definition
two types commonly used 1) Dry Heat: (a) oven T≥125°C ; 90-180 minutes (glassware and heat-resistant plastic) (b) Flaming pipettes and inoculating loops 2) Wet Heat: autoclave (large pressure cooker) (a) Pressure: 15psi (greater pressure, keep liquid from boiling) (b) Temp = 121°C; 20-30 minutes **Drawback: thermolabile components --> must be sterilized separately then added later
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Term
Sterilization Thermolabile components definition how to sterilize |
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Definition
components which are prone to denaturing/destruction w/ high heat Must be sterilized separately then added later (1) Autoclave a concentration solution of the component ie. carbohydrates and polyalcohols rate of thermal decomposition is inversely proportional to concentration (2) Cold-filter sterilize the thermolabile component ie. amino acid solutions |
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Term
Sterilization Chemical Sterilizants |
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Definition
chemical used to sterilize equipment; 4-18 hours; kills microbes/spores by protein denaturation
Ethylene oxide --> most common[image] need special equip. & technical skills limited to industry how they sterilize syringes, petri dish, pipettes White patch on sleeve allows C2H4O to enter, w/ O2 becomes CO2 + H2O |
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Term
Sterilization Cold Filtration |
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Definition
*preferred method for sterilizing liquid (no need to worry about thermolability) - Carried out at fridge temperature (4°C) but RT(23°C) can work too Sterilize liquid food/beverage: milk, beer, fruit juice (need not be refrigerated!) Sterilize air: HEPA filters Membrane filters, w/ pore diamters of 0.35-0.5μm to trap living life forms; 0.01μm to trap viruses [image] |
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Term
Sterilization Nonionizing Radiation |
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Definition
energetic to make molecules more reactive but not enough to ionize molecules ie. microwaves (kill by thermal effects on moisture of cell) ie. UV radiation (damage DNA --> can repair but introduced too rapidly w/ germicidal lamps) --introduces thymine dymers |
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Term
Sterilization Ionizing radiation |
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Definition
render target molecule more reactive by making it ionized γ-ray or β-ray Sources: X-ray machine, cathode ray tubes, radioactive nuclides ionized water formed hydroxyl radicals which damage organic molecules (very reactive) γ-ray emitters: 60Co and 137Cs γ-ray used to sterilize: Tissue grafts: cartilage,tendon, skin, heart valve Drugs: chloramphenicol, ampicillin, tetracycline, atropine, vaccines, ointments Medical & lab supplies Foodstuffs Mail |
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Term
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Definition
The time required for a 10-fold reduction in the population density Higher temperatures reduce population numbers faster Moist heat better than dry heat Spores may survive even though vegetative cells die Thermal death time Time it takes to kill all cells at a given temperature Thermal death point
Temperature at which all cells die in a 10 minute period Heat: measured in minutes Radiation: measured in grays (1 gray = 100 rad = 10,000 erg/g) |
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Term
Medium construction Defined medium |
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Definition
exact chemical composition of each and every ingredient is known Most complicated ingredient: casamino acids - mixture of a.a. obtained by acid/enzymatic hydrolysis of milk protein casien
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Term
Medium construction Undefined medium |
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Definition
precise chemical composition of at least one ingredient is unknown - yeast extract (varies and is unknown) - infusions (brain/heart) * not based on # of ingredients but precision of known composition |
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Term
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Definition
absolutely required for growth, medium must provide all essential nutrients all need: water, carbon source, energy source, nitrogen source Things bacteria can not make for themselves ie. L. mesenteroides can't make 19 of the 20 common a.a so 19 a.a. have to be in the medium |
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Term
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Definition
support growth but not required for growth compounds the cell has the ability to synthesize for itself i.e. e.coli can make all 20 of the a.a., extra a.a in media would be for support only |
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Term
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Definition
| contains only essential nutrients - bare necessities for growth and division |
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Term
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Definition
contains essential and nonessential nutrients Microbes can grow faster - supplies many biosynthetic end products - energy required to make biosynthetic end products - cellular energy can be devoted to growth instead microbes will grow auxotrophically |
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Term
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Definition
wide biosynthetic capacity - require only a few simple nutrients in order to make all the monomers necessary to assemble cellular components |
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Term
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Definition
lost greater/lesser biosynthetic capacity have more complex nutritional requirements |
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Term
Essential Nutrients Water |
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Definition
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Term
Essential Nutrients Source of carbon |
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Definition
two sources: Inorganic: CO2/CO3-2 Autotrophy: use of inorganic source of carbon photoautotroph, photolithotroph, chemoautotroph, chemolithotroph Organic: every major family of organochemicals, including many xenobiotics (chemical found in organism not usually present), can be used as a carbon source. Organotrophy: use of an organic source of carbon photoorganotroph, chemoorganotroph |
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Term
Essential Nutrients Source of energy |
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Definition
Generate energy by: Phototrophy: conversion of light energy into usable energy photoautotroph (photolithotroph); photoorganotroph Chemotrophy: conversion of E released by chemical oxidation into biologically-useful energy chemoautotroph (chemolithotroph); chemoorganotroph |
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Term
Essential Nutrients Source of Nitrogen form bacteria needs form that exists in environment ways to assimilate results storage needed |
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Definition
Needs reduced form: NH3, NH4+1 Exists in environment as oxidized form: NO3-1, N2 Cell needs to reduce nitrogen to use/assimilate it WAYS: (1) direct utilization of reduced nitrogen NH3 --> NH3 (2) Assimilatory nitrate reduction NO3-1 --> NH3 (3) Assimilatory diatomic nitrogen reduction (N2fixation) N2 --> NH3 RESULT: changed to ammonia to use --> VOLATILE must change to nonvolatile and store STORAGE: α-amino group of L-glutamic acid Υ-amide group of L-glutamine |
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Term
Direct utilization of reduced nitrogen |
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Definition
reduced forms produced in most environments: (1) ammonia (2) organic amines (3) organic amides PREFERRED nitrogen sources Ammonia: -large amounts produced by decomposition of dead organic matter - still not a major source of nitrogen (1) VOLATILE (decomposes and is lost in air) (2) competition is great (all bacteria can use) Organic amines and amides -products of decomposition -nonvolatile [image] |
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Term
Assimilatory nitrate reduction |
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Definition
Nitrate -nonvolatile -major source of nitrogen Enzymes (cytosolic/soluble) Nitrate Reductase NO3-1 + 2e- + 2H+ --> NO2-1 + H2O Nitrite reductase NO2-1 + 6e- + 7H+ --> NH3 + 2H2O Cytosolic enzymes convert nitrate to nitrite and then nitrite to ammonia inside the cell |
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Term
Assimilatory diatomic nitrogen reduction |
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Definition
- N2 fixation - most are bacteria but some archaea can fix N2 Enzyme: nitrogenase -rapid and irreversible inactivation when exposed to O2 --> must have mech. to protect or grows anoxically N2 + 8e- + 8H+ + 16ATP + 16H20 --> 2NH3 + H2 + 16ADP + 16Pi
FREE LIVING NITROGEN FIXERS
SYMBIOTIC NITROGEN FIXERS |
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Term
Free living nitrogen fixers |
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Definition
*use N2 when no other sources of nitrogen are avail. *grow at only 5-10% of rate observed w/ others *diverts lots of ATP --> less to use for cellular growth and division |
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Term
Symbiotic nitrogen fixers |
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Definition
only fix N2 when establish a symbiotic relationship w/ a plant (legume) |
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Term
Ammonia acceptors + Ammonia carriers (donors) |
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Definition
- volatile --> must convert Acceptors: picks up ammonia α-ketoglutaric acid L-glutamic acid Carriers: can serve as donor in biosynthetic rxn
L-glutamic acid L-glutamine Ammonia acceptor + NH3 = ammonia carrier [image] |
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Term
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Definition
Oxidized sulfur exists in environment, must be reduced to use Inorganic forms
Sulfate
Thiosulfate
Atomic sulfur
Iron sulfide
Organic forms
Sulfur containing amino acids
H2S -used directly as a source of energy
-Volatile so joins with L-serine (sulfide acceptor) to become L-cysteine (sulfide donor) [image]
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Term
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Definition
- PHOSPHATE: major intracellular and extracellular form of phosphorus - only Pi is transportable by cells Enzyme: Phophatase
- recover phosphate from organophosphates |
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Term
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Definition
- limited solubility in water, but require in high amts Ferrous (Fe+2) --> used by cell
(1) oxic, ph≤2 (2) anoxic, rgrdlss of pH Ferric (Fe+3) --> in envrnmnt more plentiful (1) oxic, ph≥2 - very insoluble so must release iron siderophores (chelating agents) to obtain ferric iron from envrnmnt Ferrous will spontaneously oxidize to Ferric iron when oxic and ph≥2, then precipitates as ferric hydroxide
Fe+2 + 1/4O2 + H+ --> Fe+3 + 1/2H2O Fe+3 + 3OH- --> Fe(OH)3 Sum: Fe+2 + 1/4O2 + 2OH- + 1/2H2O --> Fe(OH)3 |
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Term
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Definition
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Term
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Definition
- essential nutrients - required in high amts - consciously added to the media - water, carbon, sulfur, iron, phosphorous K+, Zn+2, Ca+2, Mg+2, Mn+2 |
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Term
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Definition
- essentail nutrients - required in trace amts - not consciously added to teh media Co+2 and Mo+2 |
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Term
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Definition
- bacteria organism can adapt use light for energy when anoxic when oxic, uses chemotrophic means |
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Term
Enrichment (selective) media |
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Definition
designed to expand the population of a bacteria you are trying to isolate which usually may not be abundant (may be grown over by other more prominent bacteria) |
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Term
Enrichment (selective) media Penecillin or cycloserine |
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Definition
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Term
Enrichment (selective) media High dye concentrations |
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Definition
selects for gram negative |
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Term
Enrichment (selective) media Low concentration of phenethyl alcohol |
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Definition
selects for gram positive (too high concentration inhibits both - and +) |
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Term
Enrichment (selective) media Heating to 80-90ºC for 15-20 minutes |
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Definition
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Term
Enrichment (selective) media No organic source of carbon |
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Definition
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Term
Enrichment (selective) media No source of nitrogen |
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Definition
selects for nitrogen fixers |
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Term
Enrichment (selective) media Bile salts (sodium deoxycholate)
and/or Lactose as sole energy source |
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Definition
selects for enteric (intestinal) bacteria |
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Term
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Definition
prokaryotes divide this way Turns one parental cells into two identical daughter cells (1) CELL GROWTH 95% doubles in size and mass (2) POPULATION GROWTH 5% crosswall forms single cell splits into two |
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Term
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Definition
| each doubling of a parental cell |
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Term
Generation time Doubling time |
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Definition
time it takes for one generation time it takes for parental cell to double g = t/d N=N02d logN = logN0 + 0.3d d = 3.3 log (N/N0) g = t/[3.3log(N/N0)] |
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Term
Fundamental Growth Equation |
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Definition
N = N02d log(N) = log(N0) + dlog(2) log(N) = log(N0) + 0.3d d= # of doublings can be computed but not measured |
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Term
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Definition
measure of the # of generations that occur per unit time in an exponentially growing culture k=1/g d=kt k = growth rate constant t = time d = t/g or g = t/d g = generation time |
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Term
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Definition
mass in culture attributed to living things (cell #) * (cell mass) (cell/mL) * (mg/cell) = mg/mL METHODS: Dry weight: not often used (destructive and large sample) heated to drive off water, residue is weighed Total cell nitrogen: not often used (destructive and large sample) needs special equipment Total cell DNA, RNA, and protein not often used (destructive, small sample) takes too long Absorbance (optical density) most often used nondestructive, small sample, accurate, easy A660nm = Cell Size = Cell Mass |
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Term
Absorbance (Optical Density) |
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Definition
[image] I0=incident light I=tramitted light T=(I/I0) x 100 A = 2 - logT A = 2 - log (I/I0 x 100)
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Term
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Definition
bacteria/mL = AxB A = total bacteria/total large squares counted
X B = total large squares/well volume (mL) Disadvantages: - pricey - tedious --> error - high cell desnity (10e6 cells/mL) - can't be motile - dead and alive look the same |
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Term
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Definition
viable cell = colony forming unit (cfu) N = C/DV N = cfu/mL C = cfu/plate D = cululative tube dilution factor V = volume plated only used for 30 to 300 colonies/plate --> low C = large systematic error --> high C = colony sizes become smaller, could have confluent colonies [image] |
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Term
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Definition
adjustment little to no cell division When cells taken from: (and placed in same conditions) Exponential growth no lag phase when no adaptability is needed Stationary phase lag still exists, lag longer the longer the cells were in stationary phase |
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Term
Growth Curve Log (exponential) Phase |
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Definition
cells obey fundamental growth equation sharp increase in cell # over time balanced growth: cellular components increase at the same relative rate that the population increases |
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Term
Growth Curve Stationary Phase |
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Definition
cells metabolically active but do no divide no increase in cell numbers Reasons depletion of essential nutrients essential gasses (limited to top layer) accumulation of metabolic waste |
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Term
Growth Curve Death Decline Phase |
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Definition
- exponential decrease in cell # - in many cases cell lysis - Autolytic (self lysing) most gram + few gram - |
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Term
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Definition
Tmin = lowest temperature w/ growth observed Tmax = highest temp w/ growth observed Topt = best T for growth (growth rate highest) Topt is closer to Tmax |
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Term
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Definition
| capable of growth over a narrow range of T |
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Term
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Definition
| capabale of growth over a wide range of T |
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Term
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Definition
cold temperatures Tmin≤0ºC Tmin≤0ºC, 7ºC, 12ºC |
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Term
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Definition
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Term
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Definition
moderate T most known bacteria 10ºC, 37ºC, 45ºC Gi tract of warm blooded animals partially shaded soil in temperate regions tropical oceans and soils |
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Term
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Definition
35ºC, 55ºC, 60ºC heat loving bacteria wood chip and compost piles hot water effluents |
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Term
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Definition
65ºC, ~90ºC, ~100ºC heat loving Archaea thermal vents |
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Term
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Definition
Freezer -20ºC many bacteria killed (ice crystals) some just not able to divide not effective as sterilizant |
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Term
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Definition
Sterilization dry heat and wet heat (autoclave) kill/remove all living organisms, viruses, and endospores Pasteurization Regular: 63-66ºC for 30 minutes Flash: 71ºC for 15 seconds Not sterile |
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Term
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Definition
O2 + e- --> O2- Superoxide least reactive most lethal stays to damage cell longer O2- + e- +2H+ --> H2O2 Hydrogen peroxide H2O2 + e- + H+ --> H2O + [OH-] Hydroxyl Radical most reactive least lethal self destructs to water |
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Term
Elimination of Biocidal Oxygen |
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Definition
Catalase 2H2O2 --> 2H2O + O2 or Peroxidase H2O2 + NADH + H+ --> 2H2O + NAD+ All: Superoxide Dismutase 2O2- + 2H+ --> H2O2 + O2 |
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Term
Strict Aerobes what enzymes are needed |
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Definition
- Must have O2 to make ATP - Contain superoxide dismutase - containe either peroxidase or catalase |
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Term
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Definition
- require O2 but at reduced levels - contain superoxide dismutase - contain either peroxidase or catalase |
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Term
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Definition
- can grow equally well w/ or w/out oxygen - contain superoxide dismutase - peroxidase |
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Term
Facultative aerobes (anaerobes)
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Definition
- grow better when oxygen is present but still grow in its absence - contains superoxide dismutase - contain either peroxidase or catalase |
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Term
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Definition
- killed in presence of O2 - no enzymes to deal w/ biocidal forms of oxygens (don't need to deal with oxygen) |
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Term
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Definition
| killed by even low external salt concentrations |
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Term
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Definition
| tolerate but do not require high external salt concentrations |
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Term
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Definition
require moderately high external salt concentrations marine and esturine environments |
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Term
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Definition
need extremely high external salt concentration Archaeons |
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Term
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Definition
raises internal concentration in order to become more saturated reduces effects of a halophilic environment (reduces dehydration) |
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Term
Antimicrobial Agent -cide |
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Definition
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Term
Antimicrobial Agent -lytic |
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Definition
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Term
Antimicrobial Agent - stat |
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Definition
inhibits growth w/out killing agent removed microbe can grow again |
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Term
Minimum Inhibitory Concentration (MIC) |
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Definition
| lowest concentration of agent that allows no measured growth |
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Term
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Definition
- plate inoculated w/ bacteria (1-2 10e6 cfu) spread plate technique used antimicrobial agent disks added to surface of agar incubate and observe Size of zone of inhibition - how sensitive it is (bigger = more sensitive) - diffusion rate of agent through agar (bigger = better diffused) |
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Term
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Definition
antimicrobial used on external body parts Organic mercurials (mercurochrome) silver nitrate iodine 70% isopropyl alcohol hexachlorophene |
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Term
Organic Mercurials (mercurochrome) |
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Definition
antiseptic used on skin denatures enzymes |
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Term
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Definition
antiseptic used on eyes of neonates to prevent Neisseria gonorrhoeae blindness denatures enzymes |
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Term
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Definition
antiseptic used on skin inhibits protein functioning strong oxidizing agent |
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Term
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Definition
antiseptic added to soaps, lotions, deodorants plasma membrane disruption |
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Term
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Definition
used to disinfect inanimate objects, some disinfectants can be used as antiseptics (70% EtOH) sodium hypochlorite mercuric dichloride copper sulfate ethylene oxide 70% Isopropyl alcohol |
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Term
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Definition
disinfectant table tops, floors, etc. Strong oxidizing agent |
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Term
Mercuric dichloride (HgCl2)
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Definition
disinfectant table tops, bench tops, floors denaturation of proteins and enzymes |
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Term
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Definition
algaecide denaturation of proteins and enzymes |
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Term
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Definition
disinfectant sterilize heat-labile materials denaturation of proteins and enzymes |
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Term
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Definition
disinfectant table tops, thermometers, etc. dissolves lipids, protein denaturation OR antiseptic degerming skin prior to injection protein denaturation; dissolve lipids |
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Term
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Definition
- antimicrobial used to combat infectious disease antibiotics - Selective toxicity (should only harm microbe w/out harming own body cells) - Therapeutic treatment given after symptoms appear - Prophylactic treatment given before not usually --> bacteria may become tolerant use when going into high risk areas |
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