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Lecture 31 PMB
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- 1Physical methods for microbial growth control--> temperature
--> temperature and pressure
--> UV/etc
--> filters - 2Chemical methods for microbial growth control--> disinfectants
--> antiseptics
--> antibiotics - 3sterilizationREMOVAL of ALL living cells, spores, and viruses on an object
the only method effective against bacterial endospores! - 4disinfectionREMOVAL of PATHOGENS from INANIMATE surfaces
chemical agents: 70% EtOH, bleach, strong ionic detergents
do not put any of these in our body!! - 5antiseptisREMOVAL of PATHOGENS from surface of LIVING TISSUE (ie. skin)
chemical agents: 70% EtOH wipe, hydrogen peroxide, Neosporin, iodine containing compounds - 6sanitationREDUCING microbial population to safe levels
- 7autoclaveused for heat/pressure sterilization
--> steam achieves high T, rapidly kills everything
--> takes about 20-40 minutes for required T to be reached by required time - 8Pasteurizationprecisely controlled heat REDUCES microbial load, pathogen spread prevented
--> reduced spoilage, longer shelf life
--> does NOT kill all bacteria, just pathogens - 9Flash Pasteurizationliquids flowing through heat exchanger raised to 71ºC for 15 sec --> rapid cooling
--> milk has refrigerated shelf life of ~2 weeks after treatment - 10Ultra-high temperature Pasteurization (UHT)liquid is heated to 135ºC for 1-2 sec (using continuous flow heat exchangers) --> also kills endospores --> products with an unrefrigerated shelf life of 6-9 months, if unopened
- 11What reduces the effectiveness of disinfectants/antiseptics?Organic material (dirt, biofilm, etc) can bind to/react with these to reduce their effectiveness
- 12Antimicrobial drugs/Antibioticschemicals used in humans that kill or inhibit the growth of microorganisms
--> can be natural or synthetic
--> must have SELECTIVE TOXICITY so the host is not harmed - 13bacteriostatic growthgrowth is inhibited, but bacteria are NOT killed
OD/direct visual count and viable cell count stay constant
inhibitory molecules are at reversible binding equilibrium with a cellular protein, must keep agent at right concentration to constantly prevent growth!! - 14bactericidal growthcells are killed, but not lysed
OD/direct visual count constant, viable cell count decrease
some irreversible damage to a protein or other component --> if agent is washed out/decreased, bacteria will still be dead - 15bacteriolytic growthcells are killed and lysed
OD/direct visual count decrease, viable cell count decrease
usually do damage to the cell wall (PG) or cytoplasmic membrane --> if agent is washed out/decreased, bacteria will still be dead - 16Minimal Inhibitory Concentration (MIC)lowest concentration of an agent that completely inhibits growth
measured by a series of dilutions within culture medium, turbidity occurs in tubes with concentrations below the MIC
compares the effects of different agents (as long as everything is standardized) - 17Streptomyces antibiotic production--> production regulated by quorum sensing (G+ peptide system)
--> occurs around time of sporulation, triggered by nutrient deprivation
--> gives a competitive advantage against soil microbes
--> resistant to own antibiotics - 18spectrum of actiontells us which bacteria are affected
narrow spectrum: very specific to harming only a certain species (ie. Isoniazid)
broad spectrum : affects many types of microbes (Tetracycline, Sulfonamides, Cephalosporins, Quinolones) - 19Mechanism of Action (MOA)tells us the specific molecular target that is
bound/the process that is inhibited by the drug
typical targets: cell wall, cytoplasmic membrane, DNA synthesis, RNA synthesis (tx), protein synthesis (tl), metabolism - 20Examples of drugs that target well studied areasCell wall synthesis: penicillins
Folic acid metabolism: sulfonamides
Protein synthesis (30S): Tetracycline
DNA gyrase: Cycloflaxin - 21Target: cell wall biosynthesisMOA: B-lactam ring of Penicillin is suicide inhibitor of PBPs (perform transpeptidation in PG synthesis) --> BACTERIOLYTIC
Spectrum: most effective agains G+ bacteria since they don't always cross OM
-can have altered R groups: molecules more acid-stable, or β-lactamase resistant, or better at crossing OM of G- cells - 22how B-lactam ring binds to PBPB-lactam ring is cysteine + valine, resembles D-ala-D-ala piece of PG --> PBP transpeptidase domain binds to this COVALENTLY, enzyme is inactivated
peptide x-linking during PG synthesis
blocked - 23different steps of cell wall biosynthesis to be blockedD-Ala ligase - Cycloserine
bactoprenol recycling - bacitracin
transpeptidation - Penicillins,
cephalosporins (D-Ala mimics) and Vancomycin (binds D-Ala-D-Ala) - 24target: DNA supercoiling/replication/integrityMOA: Quinolones interact w/ bacterial DNA gyrase --> stabilize covalent intermediate between enzyme and DNA --> gyrase-DNA complex is roadblock for replication + creates broken ends --> irreversible damage to DNA, BACTERICIDAL
Spectrum: both G+ and G-
Ex: Ciprofloxacin, more soluble in fluorinated form - 25DNA gyrasea topoisomerase II, makes ds breaks and introduces negative supercoiling
found in ALL bacteria
quinolone stabilizes covalent intermediate between DNA and gyrase --> blocks DNA replication - 26target: MetabolismMOA: Growth factor analog, resembles PABA and blocks synthesis of folic acid; binds reversibly --> BACTERIOSTATIC
Spectrum: G+ and G- bacteria
Ex: Sulfanilamide - 27folic acid and PABACells need folic acid as cofactor for single-carbon reduction rxns (used in nucleotide
synthesis), and can't import it through bacterial cell membrane --> synthesize it from PABA, pteridine, and glutamic acid
Sulfanilamide binds enzyme and attaches to pteridine, acts as a competitive inhibitor - 28target: protein synthesisMOA: binds reversibly to 30S subunit of ribosome at A site --> prevents charged tRNA from entering --> BACTERIOSTATIC
Spectrum: G+ and G-
Ex: Tetracycline - 29Napthacene ring system and protein synthesisBinds reversibly to 30S subunit of ribosome at the A site
Tetracycline resembles this with two different R groups --> spectrum of activity (slightly diff drugs bind ribosomes
of diff bacteria + diff drugs can taken up into cytoplasm of diffbacteria) - 30all steps of protein synthesis and corresponding drugspremature peptide termination: Puromycin (mimics tRNA)
peptide bond formation: Chloramphenicol
(binds to 50s RNA)
movement along mRNA: Erythromycin
(binds to 50S rRNA)
incoming tRNA: Tetracycline (binds to A site)
mRNA reading accurately: Streptomycin (changes shape of 16S RNA) - 31antibiotic resistance mechanisms1) Slightly alter the target so that the drug no longer binds
(EX: point mutation in rpsL for protein S12)
2) Modify the antibiotic so that it no longer binds the target
(EX: nptI phosphorylates kanamycin, cat acetylates chloramphenicol, B-lactamases cleave B-lactam ring)
3) Pump the drug out of the cell
(EX: tatA catalyzes exchange of tetracycline out for protein in) - 32Triclosanthought to be non-specific, disrupting membrane structure/function
used in toothpaste, soaps, scrubs
low water solubility but can be used in high concentrations in hydrophobic substances - 33Triclosan resistance: experimental methodsperform genetic selections to identify mutant genes: make a genomic library from each resistant mutant --> select for plasmids that would confer increased resistance on WT strain
- 34results of Triclosan experimentsresistant mutants had pt. mutations --> a.a. subs in FabI (carrier protein reductase ENR involved in fatty acid (FA) biosynthesis that makes new cell membranes)
MOA: Triclosan binds active site of FabI --> blocks FA biosynthesis --> arrests growth at LOW concentrations (0.1-20 μg/ml) - 35Problems with Triclosan todayTriclosan is used at HIGH 2-20 mg/ml conc. in products and toothpaste + it’s a stable compound --> it’s accumulating in the environment + could disrupt the endocrine systems of fish and mammals + *some bacteria could become resistant by upregulating the expression of efflux pumps*
- 36Triclosan effect on efflux pumpsefflux pumps that have relaxed specificity could help bacteria pump out OTHER antimicrobials used to treat disease
--> really bad
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- Microbial Growth Control and Antibiotics
- Creado por
- Candice Young
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