Lecture 13, 10/5; Lab Media

Here is the audio for today's lecture.

• Monday October 15 – exam 2
• No lab practical until he gets back.
• Handout from 10-5 (3 articles inside will be on exam 2)
• Study up on legionair’s disease in your book. He said it will probably appear on exam 2.

• • Metabolism in a nutshell (handout from 10/5/07)
• Glycolosis
• The Krebs cycle
• Electron Transport Chain or System
• Enzymes – biocatylists of the cell
• Fermentation – anaerobic metabolism of sugars
• • Natural fermentations: metabiosis → silage, sourkraut
• Various foods highlighted on page 7 of the handout from 10/5
• • Bacterial Nutrition and Growth Media (handout from 10/1)
• M-9 Important as it gives us an idea of the minimal nutrients for growth in our world.
• • E. coli can make everything it needs on m-9. It can even grow on sugar salts.
• • We would need many more nutrients (for our 20 amino acids) than m-9 has to offer.
• Complex media – don’t know what/how much is in it
• • Nutrient broth – little peptone, beef extract and water
• Peptone: It is a enzymatic digest of protein
• Beef extract: boil beef and extract nutrients
• • To make a nutrient agar add 1.5% agar
• • Nutritional groups
• Autotrophs – use CO2
• • Photo – gets energy from light (plants)
• • Chemo – get energy from oxidation of inorganic substances
• Heterotrophs – use organic carbon
• • Photo – energy from light
• • Chemo – use oxidized organic material for energy (that’s us). Classical bacteria, molds, yeasts, protozoa and E. coli too.
• Hypertrophs – “below metabolism” (don’t have their own metabolism) the category for those that don’t fit in any other groups
• • Viruses, OIP, rickettsia, chlamydia
• • Complex media (continued)
• Peptone – pepsin found in our stomach (enzymatic digest)
• Tryptone – a triptic digest of protein
• Gelatin – a protein
• Beef extract - protein
• Yeast extract – got b vitamins
• Glucose – carbon source, energy
• Agar – complex polysaccharide. Very few organisms can brake it down. It is such a good solidifing agent because it is metabolically inert. Helps it to stay solid. Stable over a wide temperature range – high boiling point and low solidifying point. Agar is hard to melt
• pH indicator – phenol red. Tells us pH of solution
• • pH 7 beautiful red
• • Acid turns it yellow
• • Basic turns it hot pink
• Basic dyes – inhibits gram (+)
• Animal blood – very nutritious. Organisms that grow on this are called – fastidious
• Trace elements – comes from tap water
• • Types of media - Selective and differential media
• Differential – both grow but behave differently therefore you can differentiate them. Best example of differential media is TSI → Triple sugar iron agar.
• Selective – only certain organisms grow. We have seen sab-dex which has a low pH.
• Selective Differential (S/D) media. EMB (eosin methalene blue agar).
• • Selective for gram (-) because of the dye
• • Differential because it has lactose. Organism can either break down lactose or not → lac positive or lac negative.
• • E. coli – lac positive. It produces so much acid from lactose that the colony is dark (metallic green sheen). No breakdown of lactose leaves it clear.
• Entrobacter pyogenes and Klebsiella are also lac positive but don’t form as much acid. Entrobacter has flat fish eye (dark center light perimeter). Klebsiella has gooey fish eye (dark center light perimeter).
• Proteus – lac (-) → clear colony
• MSA mannitol salt agar – also selective and differential. Selective because of salt, differential because of the mannitol and phenol red
• • 7.5% salt media → halo tolerant organisms grow in this media. The staphs are halo (salt) tolerant (halophilic love salt)
• • has mannitol (a sugar alchol) and has phenol red – a pH indicator. These organisms break it down to acid → yellow color.
• Staphylococcus aeureus is mannitol fermenter → changes color
• Staphylococcus epidermidis is a non mannitol fermenter → no color change
• (M-ent) M – enterococcus medium. Highly selective.
• • Highly selective for Enterococcus faecalis – found in poop.
• • In its niche it is non pathogenic. Out of its niche it is!!!
• • Some strains are resistant – (VRE) Vanocomycin resistant enterococci. We would grow MRSA in MSA.
• Enriched Media
• • Blood agar - When in doubt put it on blood agar so that we don’t miss fastidious organisms. Like TSA+5%sheep blood. Human blood is not good to grow because we have lots of antibodies. Sheep blood does not have as many.
• Fastidious: streps, pyogenes, get a sputim for strep pnemonea and grow it on blood,
• • Chocolate agar (looks chocalate)
• If it is real fastidious you grow it here. Made same but agar is at higher temperature when you add the blood which bursts the blood cells and denatures the hemoglobin.
• Extremely fastidious: neisseria gonorrhoeae, neisseria meningitides (sample taken from cerebral spinal fluid), Haemophilus (blood lover) influenza – organism that follows the flu.
• • Considerations in order to grow an unknown organism.
• Nutritional requirements
• • Autotrph – use simple medium because they use CO2 from the air
• • Heterotophs
• • Hypertrphs
• Temperature requirements
• • Thermophiles grow from about 50degrees and up
• Streptococcus thermophiles.
• • Mesophiles – most of the organisms we see are in this category. 15-50degreesC. (we are mesophiles). From the soil we would grow it at 25. If we got it from us we would grow it at 35.
• • Psychorphiles – 1-15C. Optimum growing temp is 10º (that is why stuff still grows in fridge)
• Bacillus psychrophylis
• Oxygen requirements
• • Obligate Aerobes – no oxygen no growth. Ex. Micrococcus luteus, Pseudomonas aeruginosa.
• • Microaerophiles
• • Facultative anaerobes
• • Anaerobes
• pH requirements
• self requirements
• • high level halophiles
• • low level halophiles