Microbiology: Chapter 6
Terms for classifying organisms based on carbon and energy sources.
+Autotrophs: -use inorganic carbon source (CO2) -make organic compounds from CO2 and don't require carbon from organic compounds from organisms +Heterotrophs: -catabolize reduced organic molecules (proteins, cars, amino acids, and fatty acids) -use organic carbon source +Phototrophs: use light as energy source +Chemotrophs: acquire energy from redox reactions involving inorganic and organic chemicals -either an/aerobic or fermentation depending on final acceptor
What is Differential media?
+Differentiates among bacteria +Distinguish among bacteria by observing changes in the media or the colonies
What happens to cell membranes when temperatures are too cold or too hot? Proteins and enzymes?
+Low Temperatures: -the 3-dimesional shape of microbes are more likely to form -membranes become rigid and fragile since lipids are temp sensitive -transport proceess are too slow to support metabolic activity High Temperatures: -3-dimensional shape(determined by temp sensitive hydrogen bonds) is more likely to break which denatures the cell -Lipids become too fluid, and the membrane cannot contain the cell or organelle
What is Selective media?
+Selects for specific bacteria +Contain ingredients that -favor the growth of particular organism OR -inhibit the growth of unwanted ones
Psychrophiles
- grow best about 15 degrees (between -5 - 20 degrees) -live in snowfields, ice, and cold water -do not cause disease in humans but some cause food spoilage in refrigerators
What techniques are used to grow bacteria in the various oxygen classes?
-Anaerobic bacteria growth: are reducing media (like Thioglycolate broth [consumes oxygen before getting to anaerobes) and anaerobic vessels like the Gas-Pack jar(seals bacteria which does not allow oxygen to get in) -Areotolerant/Microareophiles: carbon dioxide incubators or Candle Jars( plate sealed with candle which consumes oxygen and leaves CO2) -Preserve by refrigeration or deep freezing/lyophilization
Aerotolerant (Aerotolerant Anaerobes)
-Can live either in the presence or absence of oxygen +Do not use aerobic metabolism +Have enzymes that neutralize toxic oxygen
Facultative ( Facultative anaerobes)
-Can live either in the presence or absence of oxygen (prefer oxygen) -can use all three types of metabolism(will use aerobic if possible)
Obligate Anaerobes (Strict anaerobes or Anaerobes)
-Cannot survive in the presence of Oxygen +Use anaerobic metabolism +Lack enzymes to neutralize toxic oxygen
Membrane Filtration When would you use technique? Does it count living bacteria?Does it distinguish between life or dead cells?
-Direct Method -large sample (maybe many liters) is poured (or drawn in vacuum) through a membrane filter with pores small enough to trap cells -membrane transferred onto a solid medium, and the colonies present after incubation are counted -USED when population density is very small -ONLY ACCOUNTS LIVING CELLS -REQUIRES incubation
Microscopic Counts When would you use technique? Does it count living bacteria?Does it distinguish between life or dead cells?
-Direct Method -sample is placed on a cell counter(a glass slide composed of an etched grid positioned beneath a glass coverslip -each 1mm^2 portion of grid contains 25 large squares so they can be counted and then calculate mean # per sq -USED when a speedy estimate of population is required -DOES NOT DISTINGUISH BETWEEN LIFE AND DEATH
Viable Plate Counts (and Serial Dilution) When would you use technique? Does it count living bacteria?Does it distinguish between life or dead cells?
-Direct Method -stepwise dilution of a liquid culture in which dilution factor each step is constant -count colonies with 25-250 colonies on plate from each dilution -then multiply the number by reciprocal dilution to estimate # of bacteria per milliliter of original -USED to estimate # of microorganisms when population is large -ONLY ACCOUNTS LIVING CELLS -REQUIRES incubation
What is Complex Media? Ex's?
-Exact chemical composition is unknown EX: Extracts of yeast, organs, organisms, milk
Hydroxyl Radical
-Formed from ionizing radiation and reduction of hydrogen peroxide -most reactive of the four toxic forms of oxygen +Neutralizing: -Peroxidase and/or Catalase Antioxidants
Peroxide Anion
-Formed when superoxide anions are neutralized +Neutralizing: -Peroxidase and/or Catalase (enzymes) -Catalase converts hydrogen peroxide to water and molecular oxygen -Peroxidase breaks down hydrogen peroxide without forming oxygen, using a reducing agent such as coenzyme NADH
Metabolic Activity When would you use technique? Does it count living bacteria?Does it distinguish between life or dead cells?
-Indirect Method -(# of organisms estimated by rate of metabolism) -under standard temp conditions, the rate at which a population of cells utilizes nutrients and produces waste depends on their # -once metabolic rate is established we can estimate # of cells in culture through changes in nutrient utilization, waste production, or pH -USED often when studying environmental water samples -ONLY ACCOUNTS LIVING CELLS
Turbidity When would you use technique? Does it count living bacteria?Does it distinguish between life or dead cells?
-Indirect Method -(# of organsims estimated by measuring turbidity of broth culture) -turbid = cloudy -the greater the bacterial population the more turbid broth will be -measured with spectrophotometer (measures amount of light transmitting through) -USED since it is easy and fast -ESTIMATES BOTH LIVING AND DEAD
Dry Weight When would you use technique? Does it count living bacteria?Does it distinguish between life or dead cells?
-Indirect Method -(Organisms in broth cultures are desiccated[dried] and weighted) -organisms filtered from culture medium, dried, and weighed -suitable for broth cultures, but growth cannot be followed since they're dead -USED since filamentous microorganisms can't be measured directly -ONLY ACCOUNTS FOR DEAD CELLS
Anaerobic Media
-Oxygen-free +may contain reducing compounds such as thioglycollate -Anaerobic culture vessels
Microaerophiles
-Require lower oxygen levels (from 2-10%) +undergo aerobic respiration(must have some oxygen) +limited ability to detoxify hydrogen peroxide and superoxide radicals -if too much oxygen is present they will die (damaged by 21% concentration of oxygen[what humans breath])
Obligate Aerobes (Strict aerobes or Aerobes)
-Require oxygen +undergo aerobic respiration +use oxygen as final electron
Whats is Defined Media?
-The exact chemical composition is known
What is Transport Media?
-Transport clinical specimens from patient to the lab (speed is important so they won't die)
Lag Phase
-adjusting to new environment -actively synthesize enzymes to utilize novel nutrients in the medium (getting ready to reproduce and is making proteins and acids)
Log or Exponential Growth Phase
-after bacteria synthesize necessary chemicals for metabolism for environment, they begin rapid chromosome replication, growth, and reproduction(binary fission) -population increases logarithmically, and the reproductive rate reaches a constant as DNA and protein synthesis are maximized
Psychotroph
-both a Psychrophile and Mesophile since it can spand between -5 - 40 degrees EX: Listeria
Hyperthermophiles
-can grow between 65 - 105 degrees -stabilize proteins with extra hydrogen and covalent bonds between amino acids
Mesophiles
-grow at 37 degrees causing human pathogens -grow best in temperatures ranging between 20-40 degrees
Thermophiles
-grow between 45 - 80 degrees -are in habitats like compost piles and hot springs -stabilize proteins with extra hydrogen and covalent bonds between amino acids
Halophiles
-microbes able to live in high salt concentrations Ex: Obligate halophiles -require salt and grow up to 30% salt (burst if put in fresh water)
Alkalinophiles
-microbes that can live in habitats up to pH 11.5 EX: Vibrio cholerae, causative agent of cholera, grows best outside body in water at pH 9
Neutrophiles
-microbes which can survive at neutral pH levels (6.5-7.5) -most microbes are in this category
Acidophiles
-microbes(bacteria/fungi) in acidic habitats EX: chemoautotrophic prokaryotes that live in mines and in water running in mine waste rock(habitat has 0 pH) -oxidize sulfur to sulfuric acid(lowering pH) EX: obligate acidophiles which will die if pH reaches 7 EX: acid-tolerant microbes survive without preferring it
Singlet Oxygen
-molecular oxygen where electrons are boosted to a higher state, typically during aerobic metabolism(Photosynthesis) -very reactive oxidizing agent -Phagocytic cells( human white blood cells) use it to oxidize pathogens -photochemically produced by reaction of oxygen and light +Neutralizing: -carotenoids neutralize it -phototrophic microorganisms often contain carotenoids in pigment (remove excess energy from singlet Oxy.)
Streak-Plate
-most commonly used isolation technique in microbiological laboratories -sterile loop (or needle) is used to spread an inoculum across the surface of a solid medium in Petri Dishes -loop is lightly streaked a set pattern to dilute the sample to a point that CFU (colony formation unit) are isolated from one another
Death Phase
-nutrients are not added and wastes are not removed, population reaches a point where cells die faster than being produced -a few cells remain alive and continue metabolizing/reproducing -sometimes cells will all die and other cases few survivors may remain indefinitely
How does pH affect bacterial growth?
-organisms are sensitive to changes in acidity because hydrogen ions interfere with hydrogen bonding within proteins and nucleic acids(have ranges of acidity they prefer/tolerate) -most bacteria grow at a neutral pH (6.5 - 7.5)
Capnophiles
-organisms that grow best with a relatively high concentration of carbon dioxide(3-10%) in addition to low oxygen levels EX: Neisseria gonorrhaea
Barophiles
-organisms that live under extreme pressure (membranes and enzymes depend on pressure to maintain their 3-D, functional shape) -can't cause diseases in humans/plants/animals unless they live in depth of ocean
How does osmotic pressure (and salt concentration) affect bacterial growth?
-osmotic pressure of solution is the pressure exerted on a membrane by a solution containing solutes(dissolved material) that cannot freely cross membrane +Hypotonic: cells will gain water (salt concentration higher inside cell) and swells (cells lacking cell wall will burst) +Hypertonic: cell will lose all the water inside(if salt levels are higher outside of cell) and dies from crenation(shriveling of cytoplasm)
Superoxide Anion (or Radical)
-produce by incomplete reduction of oxygen during ET in aerobes and during metabolism by anaerobes in presence of oxygen +Neutralizing: -aerobic organisms must produce enzymes called SUPEROXIDE DISMUTASES -enzymes active sites contain metal ions combine 2 superoxide radicals and 2 protons to form hydrogen peroxide(H2O2) and molecular oxygen(O2)
How does hydrostatic pressure affect bacterial growth?
-some organisms depend on pressure to maintain their 3D shape to stay alive -without structure the proteins would denature
Stationary Phase
-the number of dying cells equals number of cells being produced -size of population remains constant -metabolic rate of surviving cells declines -nutrients are depleting, waste is adding up, and reproduction is decreasing
Generation Time
-time required for a bacterial cell to grow and divide or time required for a population of cells to double in number -most bacteria have generation time of 1 to 3 hours -under optimal conditions some bacteria (E.coli and S.aureus) have generation time of 20 min. -some slow growing species like Mycobacterium leprae require more than 10 days
What are the four main phases of bacterial growth? and what happens in each phase?
1. Lag Phase 2. Log or Exponential Growth Phase 3. Stationary Phase 4. Death Phase
What are the 5 Major Oxygen Classes
1. Obligate Aerobes 2. Obligate Anaerobes 3. Facultative 4. Aerotolerant 5. Microaerophiles
What are the 4 Major Temperature classes of bacteria?
1. Psychrophiles 2. Mesophiles 3. Thermophiles 4. Hyperthermophiles
What are the four main toxic by-products of oxygen?
1. Singlet Oxygen 2. Superoxide Anion 3. Peroxide Anion 4. Hydroxyl Radical
What is the term for the process by which a bacterial cell divides?
Binary fission: process in which a cell grows twice its normal size and divides in half to produce two daughter cells of equal size
What is the main technique used to obtain a pure bacterial culture?
Streak-Plate is the most commonly used isolation technique in microbiological laboratories
Lypholization
method of preserving bacterial cultures that involves freeze drying
What are all the techniques used to quantify the number of bacteria in a sample?
Direct Methods(for mechanically counting colonies): -Viable plate counts -Membrane Filtration -Microscopic counts Indirect Methods(estimating using machines): -Metabolic activity -Dry weight -Turidity
Do bacteria grow exponentially or arithmetically?
Exponential Growth or logarithmic growth: -trough binary fission cell divides in two; then each of the new cell divides in two to make four, and then the four to eight and so on
