BMSC 210 Microbiology Lecture Notes Fall 2024 PDF

BMSC 210 – Microbiology Lecture 22, and 23 Module 8 – control of microbes October 28 and 30 2024 Review of muddiest point(s) Review of muddiest point(s) Introduction: definition and principles How do we control microbes? I. Kill II. Remove III. Prevent growth Control does not mean...

BMSC 210 – Microbiology Lecture 22, and 23 Module 8 – control of microbes October 28 and 30 2024 Review of muddiest point(s) Review of muddiest point(s) Introduction: definition and principles How do we control microbes? I. Kill II. Remove III. Prevent growth Control does not mean sterility (i.e., free of microbes) Different methods of controls 1. Physical: heat, radiation, filtration 2. Chemical: antiseptics, disinfectants, preservative, chemotherapy The different methods of control Physical Mechanical removal Chemical Heat Radiation Filters Gases Liquids Living Non-Living Dry Moist Chemotherapy Antisepsis Ionizing Disinfection Nonionizing Sterilization Types or level of controls Inanimate items Sterilization - Sterilant Completely eliminate all vegetative cells, endospores and viruses Disinfection - Disinfectant Reduces or destroys microbial load using heat or chemicals Sanitization - Sanitizer Reduces microbial loads to a safe public health level using heat or chemicals Living tissues Antisepsis - Antiseptic Reduces microbial load using an antimicrobial chemicals Degerming Reduce microbial load using scrubbing and mild chemicals Level of controls Purpose of protocols Purpose of the control will define the protocol Protocols are affected by: 1. Time of exposure 2. Temperature/Concentration 3. Microbial load 4. Type of microbe Resistant level for different microbial types https://doi.org/10.1016/j.scitotenv.2023.162976 Poll Everywhere Question Physical methods of control Physical Heat Radiation Dry Moist Nonionizing Ionizing Dry Oven Boiling water, pasteurization X-ray Incineration Steam + pressure UV Autoclave Sterilization Disinfection Sterilization Physical methods: heat Heat kills by denaturing protein and DNA, and melting lipids Economical and easy to control and monitor Moist heat (pressure cooker) is better than dry heat (oven) Faster heat penetration/distribution, better at protein denaturation 20 min at 121°C moist heat vs 16 hours at 121°C dry heat Heat treatment Boiling, dry heat oven, incinerator Flames - Sterilizing loops Pasteurization Reduces bacterial loads but eliminate heat- sensitive pathogens Not sterile Boiling water 10 minutes at 100°C kills bacteria and viruses but not spores Sterility not guarantee Autoclaves Sterilizes 121°C for 15 minutes or more Pasteurization Autoclave: how do they work? Checking for proper autoclaving Quality control of the autoclaving process: spore strip https://microbeonline.com/autoclave-principle-procedure-types-and-uses/ Physical methods: radiation 1. Ultraviolet light - Cross links nucleotide bases in DNA - Stop replication, cause mutations - 45 seconds can kill most microbes (dependent on UV light) But: - Spore require longer exposure - Damaging to skin, eye, etc. - Does not penetrate solids Only use for surface/air sterilization Physical methods: radiation 2. Ionizing radiation (X-ray, gamma rays) Disrupt chemical bonds, break DNA Greater penetrating power than UV Mainly used in industry Cold sterilization Sterilized heat-sensitive products Food irradiation Decrease microbial load Extent shelf life The mechanical removal Mechanical removal Filters Filtration Physical removal of microbes – not killing Create filters with pores smaller than most microbes Cellulose fibers with 0.2-0.5 μM pore size Bacteria, fungi, etc. are trapped but viruses pass For heat sensitive liquids or gasses Injectable drugs, surgical gasses, etc. Summary of physical methods Summary of physical methods Reduce population by killing Summary of physical methods Control growth Poll Everywhere Question Chemical methods: some definitions Germicides: chemical used to kill microbes on surfaces Antiseptics: biological surfaces Disinfectant: inanimate surfaces Note: Disinfectants and antiseptics are not interchangeable Chemotherapy: chemical used as therapeutic to kill or prevent growth of microbes Preservatives: chemical that kill microbes or inhibits microbial growth by creating an unfavorable environment. Note: The concept of “Selective toxicity” does not apply to germicide or preservative (i.e. Germicides do not discriminate between cell type) The chemical methods of control Chemical Gases Liquids Sterilization Disinfection Living Non-Living Chemotherapy Antisepsis - Chemical - Antibiotics - Temperature - Antivirals - Exposure - Antiparasitic time - Antifungal Disinfection Sterilization Chemical methods: germicidal chemicals > 300 different chemicals have anti-microbial properties Different chemicals for different uses No single “perfect” agent Mechanism of action : Denaturing and coagulating cellular proteins and/or Dissolving lipids (e.g., in cell membranes) and/or Oxidizing cellular macro-molecules When used as directed, germicides will: - Always – greatly reduce the microbial “load” on a surface - Sometimes – create a true sterile surface (depends on the agent) Activity of antimicrobial agents Targets bacteria Bacteriostatic Stop growth, does not kill Bactericidal Kills bacteria Bacteriolytic Lyse Fungistatic/cidal Targets fungi Viricistatic/dal Targets viruses Sporocidal – kills spores Poll Everywhere Question Germicidal chemicals: what affects their activity Time of exposure Temperature of environment (less effective at lower temps) Concentration of germicide (higher is usually better) Presence of organic matter Important: Soil, blood, pus form a protective layer around microbes and will reduce the effectiveness of germicides Golden Rule: “Clean before disinfecting” Number, type, and special characteristics of the microbes High microbial increase time Biofilms Some microbes (e.g. spores, Mycobacteria) are harder to kill Assessing susceptibility – disc diffusion Testing the efficacy of germicides Poll Everywhere Question Poll Everywhere Question Poll Everywhere Question A very brief history of antimicrobial agents Pre-Modern age: traditional medicine 1909: Paul Ehrlich (Germany) → concept of selective toxicity (“magic bullet”) 1925 - 35: Screening of many other synthetic chemical compounds 1928: A. Fleming isolates the first naturally-produced antibiotic (penicillin) 1940 - 1970: Most of the currently-used antibiotics are identified Disinfectants vs Antiseptics vs Chemotherapy Toxicity towards the host: dosage and route of administration Selective toxicity for chemotherapy Kill or inhibit pathogen without damaging the host Drugs target unique structures or metabolism of the pathogen Easier for bacteria, harder for virus and very hard for eukaryotic pathogen Spectrum of activity and combination Broad spectrum – kills many different group, indiscriminate Gram-positive and Gram-negative bacteria Narrow spectrum – kills specific group Mycobacterium “Antimicrobial cocktails”: combination Antagonism: negates the effect Additive or synergistic: adds or multiplies the effect Before we start, some definition What is an antibiotic? Chemical substances that kill bacteria or prevent bacterial growth Familiar/current definition is an umbrella term for antibacterial compounds but classically: 1. Antibacterial 2. Produced naturally by microorganism 3. A chemical not a protein (i.e. bacteriocin) General characteristics of antibiotics 1. Sources of Antibiotics a) Produced naturally by soil microbes b) Some are completely synthetic chemicals c) Some are “semi-synthetic” chemicals 2. Selective Toxicity 3. Spectrum of Activity 4. “Cidal” vs. “Static” Targets of antibiotics How do antibiotics work? Principle: Try to achieve “selective toxicity” by targeting a process or structure that is unique to bacteria 1. Block DNA or RNA synthesis a) E.g., DNA gyrase and RNA polymerase 2. Block protein synthesis (16S rRNA vs 18S rRNA) 3. Inhibit bacterial metabolic pathways a) E.g., folic acid and mycolic acids 4. Disrupt bacterial cell membranes 5. Block peptidoglycan synthesis Poll Everywhere Question Antibiotics: the good, the bad and the ugly Adverse Reactions Associated with Antibiotic 1. Toxicity / Intolerance GI upset, nausea, neurotoxicity, tooth discoloration, etc. Varies depending on dose / duration / type of antibiotic, etc. 2. Allergic reactions True allergy (anaphylaxis) is probably rare (< 0.1% for penicillin) Confusion between “allergy” and “intolerance” https://www.todaysrdh.com/tooth-staining-awareness-of-oral-health-effects-of-tetracycline-and-minocycline/ Antibiotics: the good, the bad and the ugly Adverse Reactions Associated with Antibiotic 3. Disruption of “normal flora” Antibiotics (esp. broad spectrum) kill the “normal” microbiota ⇒ Opportunistic infection or “Superinfections” Poll Everywhere Question Anti-fungal agents Fungi are eukaryotes, “selective toxicity” can be difficult Agents can only be used topically Limited number of agents First effective anti-fungal drug was introduced in 1958 Various modes of action Summary: Mechanism of anti-fungal agents Mechanism of anti-fungal agents Poll Everywhere Question Antiviral agents and therapy Viral life cycle is closely linked to host cell processes Problem for selective toxicity Relatively few agents Few agents are truly “broad-spectrum” Most work only against a specific virus, or a small group of closely related viruses Antiviral agents prevent completion of viral life cycle “Static” not “cidal” (i.e. kills virus outside the cell) Many viruses are not treatable Challenges of antiviral therapy and discovery 1. Toxicity in human hosts 2. Do not work against latent viruses Require active viral replication 3. Treatment is more effective if given early Ineffective at later viral stages Requires rapid and accurate diagnosis 4. Resistance can develop quickly 5. Developing new agents is difficult Few animal models for human viruses (how to test new agents?) Summary of anti-viral strategies Examples of antiviral agents and therapy 1. Drugs that prevent viral adsorption or penetration 2. Drugs that prevent un-coating of viral NA 3. Drugs that block viral gene expression & replication a) Viral polymerases or synthases 4. Drugs that block final viral assembly &/or release a) Protease b) Neuraminidase 5. Interferon a) Stimulate anti-viral immune response Examples of antiviral agents and therapy Poll Everywhere Question What is antimicrobial resistance? Reduced ability of an antimicrobial agent to neutralize a microbe that was previously sensitive The antimicrobial agent cannot be used in a clinical setting The microbe is resistant not the host (i.e., individual/person) Subsets: Bacteria = Antibiotics Virus = Antivirals Fungi = Antifungals Parasites = Antiparasitics Biological Mechanisms of Antibiotic Resistance https://bmcbiol.biomedcentral.com/articles/10.1186/1741-7007-8-123 Biological Mechanisms of Antibiotic Resistance 1. Direct breakdown of antibiotic by bacterial enzymes e.g., “β-lactamase” enzymes → break the “β-lactam” ring of penicillin 2. Pump out the antibiotic, or prevent it from entering the cell e.g., Tetracycline - rapidly excreted from the cell after entry (↑ “efflux”) e.g., Gentamycin - altered bacterial cell wall prevents entry (↓ “influx”) 3. Modify the target on which the antibiotic acts e.g., Streptomycin - ribosome is mutated 4. Target amplification: more copies, need more antibiotics 5. Target mimicry: act as a decoy to bind antibotics 6. Enzymatic bypass: redundancy in processes Poll Everywhere Question Antimicrobial Susceptibility Testing Principle: Determine the lowest concentration needed to kill or inhibit growth of a bacteria This is called the “MIC” (or Minimal Inhibitory Concentration) Antimicrobial Susceptibility Testing Interpreting MIC values: High MIC value = a lot of antibiotic needed → High MICs suggest resistance and low MICs suggest susceptibility Antimicrobial susceptivity test Concentration of antimicrobial agent Antimicrobial susceptivity test Concentration of antimicrobial agent Poll Everywhere Question For next class: Friday: Collaborative Midterm #1 Bring an electronic device to take a Canvas Bring pens or pencil Wednesday: Midterm #2 Next 3 Lectures are on the immune system Read And/or Chapter 18, 19, and 20 https://ecampusontario.pressbooks.pub/microbio/chapter/introduction-18/ https://ecampusontario.pressbooks.pub/microbio/chapter/introduction-17/ https://ecampusontario.pressbooks.pub/microbio/chapter/introduction-19/