Chapter 11: Physical and Chemical Control of Microbes

1. Chapter 11: Physical and Chemical Control of Microbes aseptic Lister • Joseph ______ started _________ techniques with medical applications. By using carbolic acid (_______) -soaked rags and instruments during and after surgery, gangrene and other infections following surgery greatly diminished. • II. Terminology and Methods of Control • A. _____________means COMPLETE destruction of viruses and • microbes (including endospores) so that even if they are placed • in a new growth medium, they will not revive or reproduce. • B. __________means to reduce the number of pathogens (including viruses) until they are not a hazard, usually involving the use of antimicrobial chemicals. • C. _______________ refers to removing toxins. • D. ____________ refers to a substantially • reduced microbial population that meets • accepted health standards. • A clean appearance is expected! phenol Sterilization Disinfection Decontamination Sanitization

2. E. Different situations warrant different levels of microbial control. 1. daily life Simple ____________ with plain soap and water is considered to be the single most important step in preventing the spread of many infectious diseases! handwashing

3. nosocomial weakened pathogenic 2. hospitals Danger of ___________ (hospital acquired) infections because of: a. _________ condition of hospitalized patients b. higher concentration of sick people with ____________ microbes (*many resistant forms!!) c. _______ procedures (such as) d. many health care workers are ______ e. lack of _______ care (handwashing between patients, using gloves, etc.) invasive carriers aseptic 3. microbiology/research/hospital laboratories must use ________ techniques a. Work surfaces should be ______. b. All media and instruments must be ______. c. Used ________ must be properly disposed of. aseptic clean sterile cultures

4. microbe contamination environmental infection endospores III. Selection of an antimicrobial procedure depends on many factors such as the type of _______, the extent of ____________, ____________ conditions, and potential risk of _________. A. types of resistant microbes 1. Bacillus and Clostridium can make ___________. 2. Mycobacterium has ______ cell walls. 3. ____________ is capable of metabolizing unusual substances for food. (Like disinfectants!) waxy Pseudomonas B. the extent of contamination (size of the microbial population) 1. ‘Industry standard’ requires that ____% of the population is killed with every __ minutes of exposure to the treatment a. 100 microbes  10 microbes  1 microbe in __ minutes b. 1010 microbes        would take ___ minutes SO, ________/_________ first helps reduce the population before disinfection or sterilization. C. environmental conditions 1. _____________ ( heat chemical action) 2. _____ 3. ____, _______, _______, ______ can all block chemical action 90 2 4 20 washing scrubbing temperature pH saliva dirt blood feces

5. Critical D. Potential risk of infection 1. _______ items come into direct contact with body tissues. Semicritical 2. ____________ items come into contact with mucous membranes, but do not penetrate body tissues.

6. Noncritical 3. ____________ items only touch keratinized skin surfaces. IV. Methods of Physical Control A. ______ works by_________ cell proteins /enzymes. It is the most common control method because it is fast, reliable, inexpensive & nontoxic. 1. ______ heat a. _______ 100°C/10 minutes (kills most microbes & inactivates most viruses, but does not destroy __________). denaturing Heat Moist Boiling endospores

7. Pasteurization b. ____________: a brief heat treatment followed by rapid cooling. (Kills pathogens and reduces the number of spoilage organisms in milk, juices, wine, beer: Does not sterilize!) (1). LTLT (Low Temperature Long Term) 63°C/30 minutes *(2). HTST (High Temperature Short Term) 72°C/15 seconds Autoclave c. __________ (steam under pressure) (1). 15-20 psi/15-20 minutes/121°C (2). ________ equipment, media, etc. (3). used in canning procedures to destroy Clostridiumbotulinum __________! Sterilizes endospores

8. Microbiology is Fun! 2. ___ heat sterilizes. a. Hot air ovens (160-170°C/2-3 hours) used when ________ is undesirable. b. ____________ (burning) (1). _________/___________used to destroy disposable items, soiled dressings, tissue specimens etc. @ 800°C to 6500°C c. The hottest part of a Bunsen burner flame reaches 1,870°C for ______ during lab. Dry moisture Incineration furnaces incinerators flaming

9. ultraviolet B. Radiation (waves having energy but no mass) causes lethal changes in DNA, denatures proteins, but doesn’t reliably destroy endospores)! 1. Nonionizing rays = ________ radiation a. can be used to reduce the number of organisms in air and on clean surfaces b. of limited use, cannot penetrate materials like cloth, glass, paper 2. Ionizing rays = ________ or _____________ a. can be used to __________ items that are heat or chemical sensitive, such as plastics b. more effective, penetrates liquids and most solids (used to treat Washington DC mail) c. In the US, radiation is approved to treat pork to prevent ___________, to treat beef for ________ contamination and used to treat chicken for _________ contamination. X-rays Gamma rays sterilize trichinosis E. coli Salmonella

10. 3. microwaves a. do not affect microbes directly, but may kill by _____ they generate b. drawback is that microwave heating is ________ heat uneven C. Filtration (may be used for air, some heat sensitive materials such as serum, vaccines, drugs, IV fluidsbeer/wine) 1. _____ ________ ________ ____ (HEPA) filters remove airborne contaminants; used in operating rooma, for people with allergies, etc. 2. In fluid filtration, _______ are separated from ________ by passing through _______ with extremely fine pores a. Mechanical force or vacuum suction helps fluid through the filter b. does not sterilize unless pore size is small enough to trap everything (smaller pores,  cost) High-Efficiency Particulate Air liquids solids filters

11. Chemical membrane DNA proteins V. Methods of ________ Control (* for heat sensitive items, large surfaces) Destructive actions include injury to the cell _________, denaturation of cell ________, inhibiting replication of _____. A. Disinfectants Vs Antiseptics 1. _____________ are chemicals used on inanimate objects. a. ___________ are chemicals that KILL/DESTROY germs. (examples: fungicides, bactericides, viricides) b. __________ refers to chemicals that do not kill, but prevent the growth of microbes . (examples: bacteriostatic, fungistatic) 2. __________ are disinfectants nontoxic enough to be used on skin. B. Germicides are grouped according to their _______ (strength) 1. __________ destroy everything, including endospores (for sterilizing scalpels, respiratory therapy equipment, proctoscopes, plastic Petri dishes, endoscopes) (ethylene oxide gas, hydrogen peroxide) Disinfectants Germicides Germistatic Antiseptics potency Sterilants

12. High 2. ____ level disinfectants (do not reliably destroy endospores) (used for GI endoscopes) (iodine, phenol, chlorhexidine, heavy metals such as silver nitrate) 3. ___________ level disinfectants (will kill Mycobacterium, but do not destroy all viruses or endospores, even with prolonged exposure) (used for stethoscopes, electrodes, thermometers) (alcohols: ethyl alcohol, isopropyl) 4. _____ level disinfectants (will not kill Mycobacterium) (soaps, detergents) Intermediate Low

13. Phenol coefficient time identical organism concentration less C. ______ _________ (5% Phenol is the standard against which chemical agents are tested and compared) 1. Each chemical is compared for the same length of _____ on the same _________ under ________ conditions 2. IF the chemical being tested requires a greater ____________ or a longer ______ than phenol, its efficiency is _____ than phenol. IF the chemical being tested requires a lower concentration or a shorter time than phenol, its efficiency is _______ than phenol. 3. Ratio of: tested chemical activity phenol activity < 1 means _____ efficient than phenol > 1 means _____ efficient than phenol time greater less more

14. Toxicity material D. Selecting the Appropriate germicidal chemical 1. ________ (the benefit of disinfecting or sterilizing an item or surface must be weighed against the risks associated with the use of that chemical) (hospital Vs home/office) 2. compatibility with the ________ being treated (metal, rubber, glass, plastic) 3. ________ may necessitate rinsing 4. ______ and availability (bleach) 5. _________ and stability (concentrates require less space and store for long periods, but when diluted/mixed, often have limited shelf life) 6. _____________risk (disposal procedures needed) Residue Cost Storage Environmental

15. Chemical acids nitrates VI. Methods used for Preservation (delaying spoilage) of Perishable Products A. ________ preservatives (both nonfood and food) 1. organic ______ lower pH (inactivates enzymes, inhibits growth, but does not always destroy microbes) 2. ________ and _______ inhibit germination of Clostridium botulinum endospores! B. Low Temperatures 1. _____________ a. 0-10° C (___° C average) b. retards but does not prevent growth 2. ________ a. ___° C b. prevents growth but does not kill all organisms C. Increased _______ pressure by adding _____ or ______; causes water to leave the cell, killing it. D. ___________ (dehydration) of the material (natural [sun] or artificial) nitrites refrigerator 4 freezer -20 osmotic sugar salt Desiccation

16. E. ____________ (freeze-drying) 1. materials _______ frozen at temperatures well below 0°C 2. vacuum while frozen to remove ________ (lightweight) 3. biological cultures, medications, foods (expensive) Lyophilization rapidly moisture

17. Chap 12: Elements of Chemotherapy Chemotherapy I. Terminology A. ____________ = use of chemical agents to treat disease B. _________________ agent (CTA) = chemical agent used for treatment of disease (even cancer) C. _____________ agent (AMA) = chemical agent used to treat diseases caused by microbes II. Antimicrobial Agents A. Types of antimicrobial agents 1. ________ agents = metabolic products produced by certain groups of fungi and fungal-like bacteria that are antibacterial in action 2. __________ agents = produced in the laboratory 3. _______________ agents = derivatives of natural agents altered in the laboratory by adding chemical groups to improve effectiveness Chemotherapeutic Antimicrobial Natural Synthetic Semi-synthetic

18. enzymes competitive B. Modes of action 1. interfere with microbe’s chemosynthesis by inhibiting ________ 2. Disruption/interference with a. of an essential metabolite by _________ inhibition (Sulfa drugs mimic PABA, blocking folic acid synthesis) (p. 77) b. by weakening/disrupting the bacterial cell ______ (Penicillin inhibits the enzyme that builds the amino acid cross- linkages of peptidoglycan) (p. 78) c. by damaging the cell ___________ (Polymixin cleaves the layers of the membrane like a knife) (p. 78) d. by inhibiting ________________ at 70s ribosomes (p. 79) (Erythromycin inhibits translocase, freezing the ribosome on the mRNA.) (Tetracycline blocks tRNA attachment to mRNA) (Chloramphenicol inhibits transferase, preventing peptide bond formation between amino acids.) (Streptomycin causes a misreading of mRNA.) e. by inhibiting nucleic acid (______ and/or ____) synthesis (Antiviral: AZT inhibits reverse transcriptase.) (Antibacterial: rifampin inhibits RNA polymerase.) (Antifungal: griseofulvin inhibits RNA polymerase.)

20. Folic acid PABA

21. B. Modes of action 1. interfere with microbe’s chemosynthesis by inhibiting ________ 2. Disruption/interference with a. of an essential metabolite by _________ inhibition (Sulfa drugs mimic PABA, blocking folic acid synthesis) (p. 77) b. by weakening/disrupting the bacterial cell ______ (Penicillin inhibits the enzyme that builds the amino acid cross- linkages of peptidoglycan) (p. 78) c. by damaging the cell ___________ (Polymixin cleaves the layers of the membrane like a knife) (p. 78) d. by inhibiting ________________ at 70s ribosomes (p. 79) (Erythromycin inhibits translocase, freezing the ribosome on the mRNA.) (Tetracycline blocks tRNA attachment to mRNA) (Chloramphenicol inhibits transferase, preventing peptide bond formation between amino acids.) (Streptomycin causes a misreading of mRNA.) e. by inhibiting nucleic acid (______ and/or ____) synthesis (Antiviral: AZT inhibits reverse transcriptase.) (Antibacterial: rifampin inhibits RNA polymerase.) (Antifungal: griseofulvin inhibits RNA polymerase.) enzymes competitive wall

22. Glycan “backbone”

23. B. Modes of action 1. interfere with microbe’s chemosynthesis by inhibiting ________ 2. Disruption/interference with a. of an essential metabolite by _________ inhibition (Sulfa drugs mimic PABA, blocking folic acid synthesis) (p. 77) b. by weakening/disrupting the bacterial cell ______ (Penicillin inhibits the enzyme that builds the amino acid cross- linkages of peptidoglycan) (p. 78) c. by damaging the cell ___________ (Polymixin cleaves the layers of the membrane like a knife) (p. 78) d. by inhibiting ________________ at 70s ribosomes (p. 79) (Erythromycin inhibits translocase, freezing the ribosome on the mRNA.) (Tetracycline blocks tRNA attachment to mRNA) (Chloramphenicol inhibits transferase, preventing peptide bond formation between amino acids.) (Streptomycin causes a misreading of mRNA.) e. by inhibiting nucleic acid (______ and/or ____) synthesis (Antiviral: AZT inhibits reverse transcriptase.) (Antibacterial: rifampin inhibits RNA polymerase.) (Antifungal: griseofulvin inhibits RNA polymerase.) enzymes competitive wall membrane

24. hydrophilic hydrophobic amphipathic

25. B. Modes of action 1. interfere with microbe’s chemosynthesis by inhibiting ________ 2. Disruption/interference with a. of an essential metabolite by _________ inhibition (Sulfa drugs mimic PABA, blocking folic acid synthesis) (p. 77) b. by weakening/disrupting the bacterial cell ______ (Penicillin inhibits the enzyme that builds the amino acid cross- linkages of peptidoglycan) (p. 78) c. by damaging the cell ___________ (Polymixin cleaves the layers of the membrane like a knife) (p. 78) d. by inhibiting ________________ at 70s ribosomes (p. 79) (Erythromycin inhibits translocase, freezing the ribosome on the mRNA.) (Tetracycline blocks tRNA attachment to mRNA) (Chloramphenicol inhibits transferase, preventing peptide bond formation between amino acids.) (Streptomycin causes a misreading of mRNA.) e. by inhibiting nucleic acid (______ and/or ____) synthesis (Antiviral: AZT inhibits reverse transcriptase.) (Antibacterial: rifampin inhibits RNA polymerase.) (Antifungal: griseofulvin inhibits RNA polymerase.) enzymes competitive wall membrane protein synthesis

27. B. Modes of action 1. interfere with microbe’s chemosynthesis by inhibiting ________ 2. Disruption/interference with a. of an essential metabolite by _________ inhibition (Sulfa drugs mimic PABA, blocking folic acid synthesis) (p. 77) b. by weakening/disrupting the bacterial cell ______ (Penicillin inhibits the enzyme that builds the amino acid cross- linkages of peptidoglycan) (p. 78) c. by damaging the cell ___________ (Polymixin cleaves the layers of the membrane like a knife) (p. 78) d. by inhibiting ________________ at 70s ribosomes (p. 79) (Erythromycin inhibits translocase, freezing the ribosome on the mRNA.) (Tetracycline blocks tRNA attachment to mRNA) (Chloramphenicol inhibits transferase, preventing peptide bond formation between amino acids.) (Streptomycin causes a misreading of mRNA.) e. by inhibiting nucleic acid (______ and/or ____) synthesis (Antiviral: AZT inhibits reverse transcriptase.) (Antibacterial: rifampin inhibits RNA polymerase.) (Antifungal: griseofulvin inhibits RNA polymerase.) enzymes competitive wall membrane protein synthesis RNA DNA

28. C. Criteria that determine the effectiveness of antimicrobial agents 1. ________ toxicity = destroys or inhibits microbe without affecting host cells Selective Spectrum Broad 2. __________ of activity = range of microbes inhibited or killed a. ______spectrum usually effective against Gram+ and Gram- bacteria (1). useful when no time to figure out exactly which microbe is causing disease (2). disadvantage is that it disrupts normal flora too (resulting in __________ infections caused by opportunists). b. _______spectrum requires identification of the pathogen 3. Tissue distribution, metabolism & excretion a. ______ in body fluids (to be distributed in the blood) b. _______ in body fluids (so it is not broken down easily) assuring constant and effective levels in the body (pH of stomach may limit ______ administration unless coated) c. must be _________ by body tissues affected d. _________ refers to the elimination rate of a drug (this dictates the ___________ of dosage needed) secondary Narrow Soluble Stable oral absorbed Half-life frequency

29. allergenic mutagenic toxic 4. should be non __________ and not cause adverse reactions 5. should be non __________ to reduce development of resistant strains D. Disadvantages of antimicrobial therapy 1. ______ effects on normal tissues (especially liver &/or kidneys) 2. disturb ____________ 3. ________ reactions 4. development of __________ strains of bacteria, usually by producing _________ that destroy AMA (such as penicillinase) a. _________ occur naturally b. resistance genes on _________ that can be spread from bacterial cells to other bacterial cells by ____________, ______________, or ____________. normal flora Allergic resistant enzymes mutations plasmids Conjugation Transduction Transformation

30. Discriminate identification prescription E. Avoid disadvantages by 1. __________ (careful) use of AMA a. Dr: proper ____________ of disease microbe & proper __________ of AMA b. patient: maintain proper levels by (1). taking medication at prescribed _________ (2). taking medication for prescribed length of _____ 2. _________ effect of combination of 2-more AMA when resistance is likely to develop F. AMA testing = _________________ method (p. 66) 1. procedure a. Inoculate a solid ______ of bacteria on agar b. Place paper disks saturated with various _________ on the surface c. ________ 24 hours and then observe intervals time Synergistic disk-plate diffusion lawn antibiotics Incubate

31. inhibits Zone of inhibition 2. The principle behind this is that during incubation, the antibiotic diffuses into the agar and, if effective, ________ growth of the bacteria in its presence. 3. observations a. _________________ (no growth around the disk means the AMA is effective) b. _________ colonies are isolated colonies in the zone of inhibition They represent ________ cells from the original population! Satellite resistant Overlapping antibiotics (with synergistic effects) may be needed if satellite colonies appear. Location of satellite colonies if present