BIOS 111 Exam 3 Study guide
BIOS 111 Exam 3 Study guide BIOS 111
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This 11 page Study Guide was uploaded by Sierra Mongeon on Thursday March 31, 2016. The Study Guide belongs to BIOS 111 at University of Nebraska Lincoln taught by Dr. Kenneth Nickerson in Spring 2016. Since its upload, it has received 133 views. For similar materials see Microbiology in Biology at University of Nebraska Lincoln.
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Date Created: 03/31/16
Exam 3 Study Guide Chapter 6: Microbial Growth Growth factors: 1. Oxygen a. 2. Temperature a. Optimum, lowest and highest i. Autoclaving temps., pasteurization kill w/ high heat b. Psychrophilesàmesophilesàthermophiles i. Most human pathogens = mesophiles c. Refrigeration of food: cold temp slows growth (except Listeria, discussed below) i. Smaller containers of food best, allow cold air to penetrate faster (rice example) 3. pH a. acido-, neutro-, alkalo- (philes) b. most human pathogens=neutrophiles 4. Osmotic pressure a. High salt/sugar used to preserve food: hypertonic soln makes water flow out of cellà plasmolysis, loss of turgor pressure. b. Halophiles= like salt, more internal solutes to prevent plasmolysis Extremophiles= extreme loving • thermo-, halo-, baro- (high pressure), acido-, alkalo-, detergent and solvent resistant Growth Cycle • Reproduce by binary fission o DNA replicates, cell elongates, wall formed between two halves, produces genetic clones • Phases: lag, log, stationary, death. Physiological differences btwn steps. o Lag=getting used to environment o Log=exponential growth, **most Abx sensitive**. o Stationary= same rate of growth and death, constant #. Cells pollute their own environment, make toxins, use up nutrients so there can be no more growth. **During this phase, more Abx resistant!** (most Abx target growing cells) o Death=numbers begin to decline, weakened cell walls. Don’t use to inoculate for new growth/culture, Gram Stain won’t work Counting bacteria in a sample 1. Direct count = Petroff-Hausser a. Hemocytometer= actually counting cells under a microscope using special viewing tool. i. Problem: can’t distinguish between dead or alive ii. Requires at least 10^6 bact. per sample (need large sample size 2. Viable count method = CFU method: serial dilutions, spread plate or pour plate i. Count colonies on agar, extrapolate back using dilutions to determine final number of viable cells per mL. 3. MPN (most probable number) i. Used for testing drinking water for presence of fecal coliforms (indicator of fecal pollution) ii. Add varying amts. water to TSB, observe growth iii. Pre-set statistical chart, gives estimate of bacteria/100mL 4. Indirect count a. Turbidity = using spectrophotometer. i. Shine light through sample, scattered light doesn’t reach detector on other side. Decrease in light reaching detector=more bacteria. ii. Requires at least 10^6 bact. per sample (need large sample size) Growth Media 1. Chemically defined: you know exactly what you put in, predetermined amounts of exact chemicals a. Minimal defined media: only adding the very basics, used to observe biosynthesis b. Also used for fastidious (finicky) microbes 2. Complex media: things like beef and yeast extract, don’t know exact amounts of elements, rough estimate. a. Ex = nutrient agar used in lab 3. Differential: used to characterize bacteria by a specific property a. Ex: hemolysis or not with blood agar 4. Selective: allows only certain type of bacteria to grow. Many media are both selective and differential. a. MacConkey agar (MAC): bile salts, crystal violet i. Selects for Gram – only ii. Bile salt = detergent, crystal violet inhibits growth of Gram + iii. Can organism metabolize lactose? Lactose or sorbitol added to MAC with pH indicator) 1. Sorbitol added = SMAC plates. Used to tell if you have E. coli O157:H7 ß causes UTIs, other disease. 2. Red and pink colonies = able to ferment sorbitol (characteristic of E. coli, but NOT O157:H7) Light and microbial growth • Light either scattered or absorbed. o Only a certain wavelength absorbed, due to chemical structure of molecule o Absorbed light excites e- to higher energy level which reflects the energy (E) in wavelength of light • Light is an energy source for phototrophs (photosynthesis) o Light (energy ) + CO2 (Carbon source) o Oxygenic § Produces oxygen, aerobic § Cyanobacteria • Also fix N from air, need S and P in water to grow. P found in dishwasher detergent, excess runoff leads to “algal blooms” • Produce toxins o Anoxygenic § No O2, anaerobic § Green/ purple pigment § Found in depths of lakes (no O2 but still light penetration) o Photosynthetic microbes found only in water • Photosystems I and II o Photosystem I: light activates ATP production only o Photosystem II: splits H20 to produce O2 § Oxygenic = both photosystems, Anoxygenic=only photosystem I • Cyanobacteria (CB) toxins o Increased risk of ALS, Alzheimers, Parkinson’s (neurodegenerative diseases) § Guam experiment and eating flying fox bats o Do not drink or boil the water (boiling doesn’t eliminate toxin) o BMAA (amino acid derivative), made by 96% CB § Able to cross BBB, enter nervous system o L-serine (amino acid) protects from BMAA toxin. Currently being tested by FDA (phase 3 trial-see trial phases in notes) Chapter 7: Controlling Microbial Growth -cidal = kills -static = inhibits growth Sterile= nothing left alive, destruction/removal of bacteria and spore Commercial sterilization= food industry, get rid of C. botulinum spores Disinfection= kill living microbes on inanimate (non-living) surface Antisepsis= kill living microbes on living tissue Degerming= local antisepsis ( Ex. Cleaning injection site) Santitization= lower microbe count for public health (Ex. Washing dishes in restaurant Plotting the death curve **D value = DRT (decimal reduction time) = how long it takes to kill 90% of population** On a graph: Straight line = single target Sterilization Techniques 1. Filter sterilization: used for heat intolerant liquids (vaccines, enzymes, vitamins a. Liquid on top of filter, vacuum tube pulls liquid through, filter catches bacteria and sterile liquid falls through. (See diagram in lecture notes or book) b. Pore size= .2 or .45 microns (bacteria too large) i. Millipore=cloth like, spun fiber ii. Nucleopore= solid with holes punched in it by gamma rays, uniform hole size c. Filtration used for air = HEPA filter (high efficiency particulate air filter) i. Microbe free air for use in hospitals, military, pharmaceuticals ii. Also used for testing amt. of microbes in air iii. Also used in BSL4 labs d. Filtration used in water quality tests: pass certain volume through filter, place filter on agar plate and observe growth. Can test for fecal coliforms in water. 2. Light (UV radiation) a. Pigments absorb light; pigmented bacteria are rare but pigments may be important in pathogenesis (S. aureus for example) i. Used to protect DNA b. Bactericidal = below 280 nm (absorbed by DNA, damages DNA, kills) i. Germicidal lamp = 254 nm c. DNA damage: thymine dimers à problems with replication. i. Can be repaired, but damage can occur faster than can be repaired 3. Temperature a. High i. Autoclave (pressure cooker, high pressure steam) 1. Can’t be used for everything 2. Size of container affects time needed 3. Relationship between temp, pressure, and altitude ii. Pasteurization 1. Kill pathogens in milk while keeping it drinkable 2. LTLT = low temp long time 3. HTST= high temp short time 4. UHT = Ultra high temp a. High for brief time, used to make sterile milk (so it can be kept for long times without refrigeration, like little coffee creamer tubs at restaurants) iii. Dry Heat Oven b. Microwaves don’t kill microbes! (The heat does, but they heat unevenly) 4. pH a. Stomach = pH 2 (acid barrier, protects from many pathogens) i. Shigella, salmonella = high acid tolerance. b. Weak acids as food preservatives i. Sorbic acid/sorbitol, propionic acid/ propionate 1. Lipophilic: change internal pH of cell 5. ROS (Reactive Oxygen Species) a. Used by macrophage to kill i. Engulfs, lysosome with ROS inside, pathogen killed and digested ii. Some pathogens survive! (S. aureus gold pigment = protection) b. Oxidative stress i. To protect, bact. need to remove oxidant, repair cell damage ii. Enzymes in bacteria neutralize ROS 1. Superoxide dismutase (superoxide to hydrogen peroxide H2O2) 2. Catalase (H2O2àhydroxyl OH) 3. Alkyl hydroperoxide reductase (OHà water) c. Nitric oxide (RNS= reactive nitrogen) also used i. People with infection à high nitrate in urine 6. High Pressure a. Food pressurized @ 6000 atm i. Denatures proteins, maintains flavors, colors and nutrients ii. Called “pascalation” 7. Electronic Pasteurization (electron beams) a. high energy electron beams kill microbes i. Controversial (people think food radioactive, but its not) b. Causes irreversible double strand DNA breaks c. “cold” sterilization d. used for spices, petri dishes, pharmaceuticals, U.S mail (for spores) 8. Chemicals Antibiotics Antiseptics/Disinfectants Specific, internal use Non-specific, external use low conc. , complex chem structure High conc, simple Natural (made my other microbes) Created by chemists a. Antiseptics b. Disinfectants i. Phenol and its derivatives 1. Lysol, hexachlorophene (for acne, impetigo) 2. Triclosan (anti bact. soap) c. Detergents i. Part hydrophobic, part hydrophilic ii. Can be + charged (cationic) or – charged (anionic) iii. Example: ammonia iv. Soaps: physically remove microbes (don’t kill, but wash away) d. Alcohols i. Ethanol 1. Tincture = something mixed with aqueous ethanol 2. 70% ethanol most effective (not 100%!!) 9. Gas – ethylene oxide a. Penetrate into structure, sanitize hospital linens/mattresses b. Modifies proteins, can cause blindness 10.Heavy metals a. Silver (Ag), Copper (Cu) b. Burn ward sponges (black sponge) = silver c. Solid Cu surfaces = registered antimicrobial agent i. Also have liquid Cu containing solns. Chemical biocides: effectiveness Least Difficult to kill-------------------------------------------------àMost Gram + viruses Mycobacteria endospores prions • Spores killed by: autoclave, electron beams, formaldehyde, ethylene oxide Chapter 20 – Antibiotics • See table above for characteristics • Goal: Kill pathogens without harming host • Pharmacokinetics: how long drug stays in blood and at what concentration o Don’t stop taking Abx because you feel better; B in body à resistance • Produced commonly by soil microbes, especially Streptomyces family o Filamentous bacteria, many species o Different types Streptomyces produce different Abx àcan be antibacterial, antifungal, kill R/C (rickettsias/chlamydias) Targets of Abx • Cell wall o Penicillins, cephalosporins, bacitracin, vancomycin • Protein synthesis (ribosomes) o Erythromycin, tetracycline, streptomycin • Nucleic acid replication/transcription o Quinolones • Synthesis of metabolites o Sulfa drugs Penicillin • History o Alexander Fleming discovered o Large scale production: finding right strain, mutating it to produce more • Targets GROWING cell wall o Can only kill growing cells!!! o Inhibits PG cross link • Beta lactam ring = functional unit • Penicillin G o Susceptible to many things. Needed drug with same action, more resistance • Semi - synthetics= keeping B-lactam ring structure but adding different side chains to produce desired properties o Acid/ penicillinase resistance, reach CNS o Amidase used to isolate common penicillin skeleton (the B lactam ring plus some),other things can be added Penicillinase= bacterial enzyme that makes penicillin non-functional by destroying B-lactam ring -Methicillin = resistant to this -suffix “-cillin” used for penicillin drugs Cephalosporins • Prefix “Ceph” or “cef” used for drugs • Generations, each more complex st o 1 gen – kill Gram + o 2 gen- Gram – o 3 gen- Cross BBB, reach CNS (cure meningitis) Tetracyclines, Erythro/Streptomycin • Inhibit protein synthesis, bind to bacterial ribosome (remember they’re different than human ribosomes) o Tetracycline = attachment of tRNA § First broad spectrum Abx o Chloramphenicol = inhibits peptide bond o Streptomycin = read RNA incorrectly à incorrect proteins o Erythromycin = also messes with protein synth. Quinolones • Affect nucleotide synthesis (DNA) • DNA gyrase= makes bact have long, supercoiled DNA o Not present in Euk. o Target of these drugs Fermented foods (picnic): • Lactic acid bacteria help make foods like sauerkraut, sausage, cheese, yogurt, buttermilk, soy sauce. o Captain cook and sauerkraut o Lactic acid benefits for food: lower pH § Adds flavor, preservative (prevent spoilage) • Probiotics= good bacteria in fermented foods. Replace gut flora killed when you’re taking antibiotics. o Eli Metchnikoff • Yeasts: bread, beer, wine Cheese production: • Milk + renin + lactic acid àcurdling • Curds and whey (whey disposed of) o Curds used to make cheese • Curds drained, cooled, salted, pressed • Ripening o Unripe=cottage chz, cream chz. o Ripened= soft and hard chz. • 400 gal milk = 150 lbs chz • Hard chz= wrapped air tight, no limit on size, anaerobic bacteria inside. 25- 35% of casein (milk protein) digested. o Ex: parmesan, romano • Soft chz = surface fungi, aerobic, small size, 100% casein digested o Ex: bleu chz, brie • Starter cultures o Cultures of selected bacteria added to milk Diseases of the Day Meningitis (general term for infection in CSF of central nervous system) • CSF= cerebrospinal fluid, the fluid that bathes brain and spinal cord. Usually sterile • BBB=blood brain barrier, prevents everything from blood from getting into CNS. Lipophilic things can cross barrier! • Brain surrounded by meninges (coverings) and blood vessels. -Caused by virus, bacteria, fungus, protozoa -Many of these causative agents found in the throat, may bacteria enter through respiratory tract. (Bacteria that cause pneumonia also can cause meningitis) • Bacteria= Hemophilus influenza o “heme”=blood o Needs 2 things to grow: heme in human blood, NAD o Gram-, non motile, has capsule. 6 different types of capsules § Type b = most virulent! § Vaccine = Hib (H. influenzae b) rd o Treat with Abx (3 gen. Cephalosporin) -Diagnosed: take CSF sample; needle into subdural space (spinal tap), plate CSF sample Listeria Listeria monocyotogenes • Food-bourne, especially dairy products • Can grow at refrigeration temps and survive stomach acid o Temp range from 4-45°C, survives low pH of stomach (ph 2) • Able to cross BBB and placenta • Can survive in phagocytes (macrophages) o Survives oxidative stress. • Uses actin polymerization as motility mechanism o Actin=long protein found in animal cells o Polymerizes protein into long “pole” it uses to push itself from one cell to the next o Pushes itself out of cell forming pseudopod, this goes inside another cell and pinches off. • Ubiquitous (everywhere!) in soil and water o Soilàanimalsàmilk or feces (which gets into water, and waste products from milk prod. also get into water. o If milk pasteurized, Bact is killed. Some cheese made with unpasteurized milk, cause disease. Serratia marcesens • Blood red pigment = prodigiosin o Pigment only produced at 20-35° • Used in study (like B. globigii in anthrax study), sprayed over land o Pneumonia in elderly people § Opportunistic pathogen only! (Only pathogen if person is already weakened) § Normally an insect pathogen • Cause of “Bleeding host” – result of large pogroms (mass killings) of Jews, thought they had defiled host Irish potato blight • Caused by a water mold (spores with flagella) o Move through water • First disease recognized as being caused by microbe o Still not believed microbes could cause disease in people, only plants Mycoplasmas • Smaller genomes than other free-living bacteria o Degenerative evolution § Free livingàmycoplasmaàrickettsiasàtheoretical minimum # of genes • No cell wall, no defined shape (pleomorphic) • Small, can go through some filters • Low % GC base pairs • Sterols in cell membrane (most prok. don’t use these!!) • Fried egg colonies • Cause pneumonia (primary cause of pneumonia not caused by S. pneumonia) • Tetracycline, erythromycin o Can’t use Abx that attack cell walls because they don’t have them! Rickettsias/Chlamydias • OBLIGATE INTRACELLULAR pathogens – can’t live outside of host cell o Live off host metabolism, steal ATP and other things • Still are considered bacteria -sensitive to Abx Typhus • Rickettsia prowazekii o Gram – o OBLIGATE INTRACELLULAR • Feces of biting lice o Also, rocky mountain spotted fever o 2 hosts: insect and human • Fever, rash, bact multiplies in cells lining cardiovascular system o Vessels collapse, blocked o 30% mortality if untreated • treat by targeting insect (with DDT ) or bacteria (tetracycline) • History o Military: Napoleon’s army, Crimean war o More men died of typhus than of actual war wounds! o Basic sanitation saves lives o Discovered by Howard Ricketts Chlamydia • OBLIGATE INTRACELLULAR; energy parasite • Susceptible to tetracycline • Chlamydia trachomatis o Leading cause of infectious blindess § Tetracycline drops in eyes of newborns § Can get it from swimming pools, towels, flies around the eyes § Trachoma = blindness § Most cases of blindness in children o NGU = non-gonococcal urethritis § GU tract infection not caused by gonorrhea § More common in women • Often a co-infection!! o Gonorrhea and chlamydia: if you have one, you probably have the other • Consequences as an STD o Sterility, damages to GU tract cells à more susceptible to virus o Cervical cancer o Passing on to baby • Often no symptoms • Rapid detetion : PCR tests • Chlamydial pneumonias = mild infection, 50% U.S population has Ab against • “Parrot fever” = type of chlamydia in parrots, can be transferred to humans • Infection cycle: o Elementary bodies – like spores, taken into cell o Reticulate bodies- growing cell. Convert to elementary bodies as host cell destroyed
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