Lec 14: Bacterial nutrition, culture media & cultivation Flashcards
Nutrition in bacteria influenced by..
Sources of carbon, e- & external energy for growth
Diversity of chemicals needed to make bacterial cell components
The need for growth factors
Carbon
The backbone of organic molecules
Electrons
Electron flow in bio reactions -> energy
Needed to reduce molecules during biosynthesis
External energy source
Necessary to power cell
Make ATP
Autotroph
Carbon source: CO2
Heterotroph
Carbon source: organic molecules made by other molecules
Phototroph
Energy source: light
Chemotroph
Energy source: Exergonic (heat or energy is released) chemical reactions (oxidation of organic or inorganic compounds)
Lithotroph
Electron source: reduced inorganic molecules
Organotroph
Electron source: organic molecules
Carbon
Found in: most cell structure (e.g cell wall) & functional chemicals (e.g DNA)
External sources (come from): CO2 (inorganic), simple & complex sugars (organic)
Nitrogen
Found in: proteins, nucleic acids, some polysaccharides & lipids
External sources: NH4+, NO3, N2 (inorganic). Proteins, AA, urea (organic)
Hydrogen
Found in: organic compounds
Come from (external sources): organic compounds, H2O
Oxygen
Found in: organic compounds and electron transport chain (respiration)
Come from (external source): H2O, CO2, O2
Sulphur
Found in: S-AA (methionine, cysteine) & vitamins (biotin, thiamine)
Come from: SO4 (inorganic). Cysteine, methionine (organic)
Phosphorus
Found in: nucleic acids, phospholipids, ATP
Come from PO4
K, Ca, Mg, Fe
Found in enzymes & ribosomes
External sources: as ions in solutions (e.g K+)
Micronutrients or trace elements
Mn, Zn, Co, Ni, Cu
Found in: cofactors (helper molecules; bind to & assist enzymes to function properly)
Come from: as ions in solution
Problems for cell in uptake of nutrients by bacteria
Microbes can only take in dissolved molecules (must take in nutrients from dilute solutions against conc gradient)
Cytoplasmic membrane prevents passage of most substances
Enormous variety of nutrients
Nutrient uptake mechanisms should be specific
Bacteria mechanisms for nutrients uptake across membrane
Passive diffusion
Facilitated diffusion (ATP not required)
Active transport (ATP required)
Group translocation (ATP required). Transport of solute molecules to higher conc in which molecule is chemically modified during transport
Facilitated diffusion
Diffusion across membrane involving carrier protein
Carrier protein in outward facing conformation binds to solute -> changes conformation to inward facing conformation -> releases solute into cell
Active transport
Transport of solute molecules to higher conc (against conc) using transport proteins
ATP energy required
After binding solute -> solute binding protein approaches ATP binding cassette (ABC) transporter -> protein attaches to transporter and releases solute -> energy released by hydrolysis of ATP -> movement of solute across membrane
Effect of environmental factors on microbial growth when culturing: oxygen
Some bacteria & most multicellular organisms are aerobic, some are anaerobes
Aerotolerant anaerobes
grow equally well in presence or absence of oxygen
Obligate anaerobes
grown only when oxygen is absent
Facultative anaerobes
do not require oxygen. Can grow with or without oxygen. Most grow better aerobically (energy production better in oxygen)
Microaerophiles
damaged by atmospheric O2 and require O2 for growth
Effect of environmental factors on microbial growth when culturing: Temperature
Microbes cant regulate their internal temp -> affected by external temps
Enzymes vary in temp sensitivity
High temp -> membrane degrades
Low temp -> membrane becomes solid
Psychrophile
Cold environments
Grow well at 0 degrees, optimum ~15 degrees
Psychrotolerant
Cold environments
Grow at 0 degrees
Optimum ~25 degrees
Mesophiles
Moderate temps
Optima ~20-45 degrees
Thermophile
High temps
Optima ~55-65 degrees
Hyperthermophile
Very high temps
Optima ~85-113 degrees
Design of culture determined by..
Sources of carbon, electrons & external energy used for growth
Diversity of chemicals needed to make bacterial cell components
The need for growth factors
Agar
A carbohydrate, extracted from
red algae
Usually added to liquid broth medium
Dissolves at 90 degrees
Sets at ~45 degrees, stable from 0-80 degrees when set
Easily sterilised
Not degraded by bacteria
Spread plate, streak plate, or pour plate yields single isolated colonies
Culture media classified based on..
Chemical composition: defined (synthetic), complex
Physical nature: liquid, semisolid, solid
Function: supportive, enriched, selective, differential
Defined or synthetic media
A medium in which all components are known
Complex media
Medium containing some ingredients of unknown chemical composition
Formulated to support growth of wide range of bacteria
Used where nutritional requirements of particular organism are known
Examples: nutrient broth/agar, tryptic soy broth/agar (TSB/TSA), MacConkey agar
Typically made from protein digests & meat extracts
Selective media
Have ingredients/conditions which inhibit growth of unwanted microbes & favour growth of specific organisms
Bacteria may also be selected by incubation with nutrients they specifically use
Differential media
Distinguish between different genera or species based on biological/metabolic properties
Often contain acid-base indicators & selective agents -> limit range of bacteria growing
Example: mannitol salt agar. Staphylococcus aureus ferments mannitol -> produce acids-> pH indicator goes from red to yellow. Non-pathogenic staphylcocci remain red
Supportive media
Support growth of many microorganisms
E.g TSB or TSA
Enriched media
Blood & other nutrients are added to general purpose media -> encourage growth of fastidious microbes
Blood agar: horse/sheep blood. Enriched and differential (alpha, beta, gamma haemolysis)
Chocolate agar (contain lysed blood cells)
MacConkey agar
Example of complex media
Use bile salts & dye crystal violet -> inhibit Gram +ve and not Gram -ve
Example of differential media
Have lactose & neutral red dye: lactose-fermenting (e.coli) colonies: red; non-fernenters (salmonella) are colourless
