Chapter 7 Microbial Growth Flashcards
Macronutrients
(C,H,O,P,N,S)
Required in large amounts
Metabolism and cell structure
K+,Ca++,Mg++,Fe+++ moderate quantities
Micronutrients
(Mn,Zn,Ni,Co,Cu,Mo)
Small amounts - difficult to determine exact requirements (tapwater sufficient)
usually enzymatic function
Chemical requirements for growth
- Carbon - CO2 or organic
- N, S, P - DNA & RNA, protein, some lipids
- Trace Elements - Metal ions - only certain cells and in tiny small amounts
- Growth Factors - not made by organism
•purines and pyrimidines: required for synthesis of nucleic acids (DNA and RNA)
•essential amino acids: required for the synthesis of proteins
•vitamins: needed as coenzymes and functional groups of certain enzymes
Media formulations
Chemically defined media - exact amounts of pure chemicals (research)
Complex media - composition varies (enzymatic)
Specialty medias
Selective - inhibit growth of certain species.
Differential - most things will grow- but will look different
Selective and differential - utilizes both aspect
Photoautotroph - Oxygenic
Produces oxygen (O2)
(Inorganic)CO2 + H2O + light = (CH2O)n + O2
cyanobacteria, algae, green plants
Photoautotroph - Anoxygenic
Does NOT produce oxygen
use inorganic material like hydrogen sulfide (to sulfur) or hydrogen gas (to water)
are obligate anaerobes (O2 toxic)
bacteriochlorophylls - green and purple sulfur bacteria
Photoheterotroph
Use light for energy
Use organic compounds like alcohols and fatty acids for carbon source
obligate anaerobes
Green and purple non-sulfur bacteria
Found in bogs, moist soil, paddy fields (light but no O2)
Chemoautotroph
Electrons from reduced inorganic compounds used for energy
CO2 used for carbon
remove electrons from H2, H2S, S, or Fe and combine them with CO2, O and H
deep-sea vent bacteria; hot springs
Chemoheterotrophs
Use organic molecules for both energy and carbon
Aerobic: uses oxygen as final electron acceptor
Anaerobic: uses sulfate, nitrate, etc. as final electron acceptor
most bacteria, all fungi, all protozoa and all animals, pathogens
Inorganoheterotrophs
Use reduced inorganic molecules for energy
Organic molecules for carbon source
some bacteria
Parasites and Saprobes
Parasites: derive nutrients from host •Pathogens •Some are obligate parasites Saprobes: free-living microorganisms that feed on organic waste from dead organisms •Opportunistic pathogen •Facultative parasite
Gas Requirements
Oxygen: O2
•Powerful oxidizing agent
•Must be “detoxified”
Carbon dioxide: CO2
•Essential for autotrophs
Oxygen Radicals (Must be neautralized even if organism doesnt use oxygen)
Toxic by-products of O2 use and exposure:
•Superoxide O2- superoxide dismutase
•H2O2
catalase, peroxidase
•Singlet Oxygen
photosynthetic organisms have carotenoids which scavenge the singlet oxygen and render it nontoxic
Oxygen Requirements- Obligates
Aerobes-
require O2 for growth (21% O2 in air)
they use O2 as a final electron acceptor in aerobic respiration
Anaerobes-
do not need or use O2 (toxic)
anaerobic respiration, anoxygenic bacterial photosynthesis, methanogenesis
Oxygen requirements- Others - Facultative anaerobes
Facultative anaerobes- Can switch between aerobic and anaerobic types of metabolism (O2 prefered, fermentation or ana-respiration if not)
Capnophiles
require a high level of CO2 and are grown in candle jars
Oxygen requirements- Others - Aerotolerant anaerobes
Exclusively anaerobic type of metabolism Insensitive to the presence of O2
Produces superoxide dismutase and peroxidase enzymes
Ex: Streptococci
Oxygen requirements- Others - Microaerophiles
Are aerobic, but need low concentrations of O2
<15% O2
Ex: Helicobacter pylori
Anaerobic growth media:Fluid Thioglycollate Medium
Anaerobes are killed by exposure to oxygen
Use a reducing medium to deplete available oxygen
Heating the media also drives off oxygen
pH Requirements
Neutrophiles: Most bacteria grow between pH 6.5 to 7.5
Acidophiles: grow below pH 4.0
Lactobacillus acidophilus (yogurt, ~pH 4)
Some molds and bacteria can tolerate acid (food spoilage)
Alkalinophiles: grow best under basic conditions (not many)
Transport: Movement of Chemicals Across the Cell Membrane - Passive transport
does not require energy
substances exist in a gradient and move from areas of higher concentration toward areas of lower concentration
Types:
•Simple diffusion – small things slip between the phospholipids (CO2, O2, N2, water)
•Osmosis – diffusion of water
•Facilitated diffusion – requires a carrier
Transport: Movement of Chemicals Across the Cell Membrane - Active transport
requires energy and carrier proteins
gradient independent
Types:
•Active transport
•Group translocation – transported molecule chemically altered
•Bulk transport – endocytosis, exocytosis, pinocytosis
Diffusion
Net Movement of Molecules Down Their Concentration Gradient Passive Transport
No addition energy required
Osmotic Solutions
Isotonic - w or w/o cell wall equilibrium
Hypotonic
Cell wall - wall prevents bursting
w/o cell wall - swelling then osmolysis
Hypertonic
cell wall - membrane shrinks inside wall -plasmolysis
w/o cell wall - shrinking and distortion
Facilitated Diffusion
Monomer sized molecules need a transport protein to cross the membrane
•Carrier protein
•No additional input of energy required
Endocytosis
Bringing substances into the cell through a vesicle or phagosome
•Phagocytosis ingests substances or cells
•Pinocytosis ingests liquids
Symbiotic Relationships
mutualism – obligatory, dependent; both members benefit
commensalism – commensal member benefits, other member not harmed
parasitic – parasite is dependent and benefits; host is harmed
Quorum sensing
a group of microorganisms coordinate functions (to release digestive enzyme, toxins, transfer DNA, etc.)
Quorum: critical number of cells
Inducer molecules: coordinate some response
Counting Bacterial Growth
2n, where n is the number of generations
exponential growth
5-10-20-40-80-160-320
Phases of Growth: Lag
no or slow growth, yet
•culture “waking up”- synthesizing DNA, enzymes, ribosomes, etc.
•or culture so dilute it is below limit of detection
Phases of Growth: Log
- Cell division as fast as possible (“balanced growth”)
- Adequate nutrients and favorable environment
- Appears linear on a log scale
- Cells are dividing and growing in geometric progression (2,4,8,16,32,64,128,etc.)
- Phase used to calculate generation time
Phases of Growth: Stationary
- Growth slows such that new cells = dying cells
- Growth is limited by space, supply of nutrients
- Depleted nutrients, and waste is accumulating
Phases of Growth: Death
- Number of cells dying increases exponentially due to build up of waste products.
- Some remain viable
Direct Measures of Growth - plate counts
Plate Counts
•enumeration of bacteria: Assumption that every live bacterium will produce a colony on a plate
•Viable counts
Direct Measures of Growth - filtration
- Used when quantity of bacteria is low
- Need to concentrate bacteria onto filter
- Transfer bacteria from the filter to an agar dish
- Touch the filter to an agar dish, then incubate to grow colonies
Direct Measures of Growth - Direct Cell Counts
- Petroff-Hausser cell counter
- Use very accurate counting chamber under microscope (like a hemocytometer)
- Use grid to establish number of organisms
Direct Measures of Growth - Automated Direct Counts
Coulter Counter •Separate based on charge •Automatic •Expensive •Breaks easily
Flow cytometry•Can count cells based on fluorescent labels•Separates live from dead•Must be able to manipulate organisms DNA
Direct Measures of Growth - Automated Direct Counts
Coulter Counter •Separate based on charge •Automatic •Expensive •Breaks easily
Flow cytometry
•Can count cells based on fluorescent labels
•Separates live from dead
•Must be able to manipulate organisms DNA
Indirect Measures of Bacteria
Turbidity: Use spectrophotometer to measure amount of light scattered by a culture
Metabolic activity: measure amount of product made or substrate used up
Dry Weight: organisms are filtered and then weighed to determine the starting material
Preserving Bacterial Cultures
- Refrigeration: for short term storage – what we use in laboratory
- Deep freezing (-70C): long term (years)
- Lyophilization (freeze drying): quick frozen cultures then placed under vacuum and sealed (decades)
Motility Medium
- Soft agar
- Inoculate straight in with needle
- Incubate at room temp
If agar cloudy then microorganisms motile but if they are near injection point non-motile
