NURS N111F Week 2 Microbiology & Pharmacology (PDF)

NURS N111F Fundamentals of Microbiology and Pharmacology WEEK 2 Interaction between Microbes and Host Learning Outcomes for Week 2 On completion of this lecture, you will be able to: 1. Describe the bacterial growth curve; 2. Explain factors affecting bacterial gr...

NURS N111F Fundamentals of Microbiology and Pharmacology WEEK 2 Interaction between Microbes and Host Learning Outcomes for Week 2 On completion of this lecture, you will be able to: 1. Describe the bacterial growth curve; 2. Explain factors affecting bacterial growth; 3. Describe the terminology of controlling microbial growth; 4. Describe methods for controlling microbial growth; 5. Describe the interactions between microorganisms and human; and 6. Describe the mechanism of pathogenicity. 2 Interaction between Microbes & Host Part 1: Microbial Growth  How microbes grow  Methods measuring bacterial growth  Factors affecting microbial growth [Factors favouring infection] Part 2: Control of microbial growth  Terminologies  Physical methods  Chemical methods Part 3: Interactions between microbes & human  Normal microbiota  Pathogenic microbes  Mechanism of pathogenicity [How bacteria (and other microorganisms) cause diseases] 3 PART 1: Microbial Growth https://www.youtube.com/watch?v=GX55_-IlfyI  How microbes grow  Methods measuring bacterial growth  Factors affecting microbial growth 4  BINARY FISSION IN BACTERIA Bacterial Growth Bacterial growth = increase in number of cell  NOT size Growth by:  Under favourable environmental conditions Mcstrother, CC BY 3.0 https://creativecommons.org/licenses/by/3.0  Binary fission via Wikimedia Commons [bacteria]  Budding [e.g. yeast, less common in bacteria]  Under unfavourable environmental conditions  Sporulation [Forming spores] Created with BioRender.com 5 Binary Fission Process by which 1 cell divides into 2 cells Grow exponentially 1 20  2 21  4 22  8 23  16 24 … … Generation time  Duration required for doubling a population (~10 min to 48 hours)  Different bacteria grow at different rates 6  Steps for Binary Fission ​English Wikipedia user ZabMilenko, CC BY-SA 3.0 http://creativecommons.org/licenses/by-sa/3.0/, via Wikimedia Commons 1. Cell elongates & DNA is replicated 2. Plasma membrane begins to constrict & new wall is made 3. Cross-wall forms, completely separating the 2 DNA copies 4. Cells separate Created with BioRender.com 7 Bacterial Growth Curve Noexchauge rf mateunals wren N suwnawings Bacterial growth in a close system (e.g. a culture flask) by binary fission follows the bacterial growth curve 1. Lag phase 2. Log phase (Exponential growth phase) 3. Stationary phase cronet ) increase 4. Death phase (Decline phase) Created with BioRender.com 8  Bacterial Growth Curve (cont’d) 1. Lag phase ( prebavation prase )  No obvious increase in population size (cell number)  Bacteria are undergoing a period of intense metabolic activity  Synthesize enzyme to prepare for cell division 2. Log phase (Exponential growth phase)  Exponential increase in population size  Number of newly divided cell > Number of dying cell veny  Plenty of nutrient support, with relatively less competition steady  Generation time can be calculated in this phase  Fast grower has a steeper slope 3. Stationary phase 𝐺𝑟𝑜𝑤𝑡ℎ 𝑟𝑎𝑡𝑒 4. Death phase (Decline phase) 𝐶𝑒𝑙𝑙 𝑛𝑢𝑚𝑏𝑒𝑟 = 𝑇𝑖𝑚𝑒 9 「 Bacterial Growth Curve (cont’d) 1. Lag phase 2. Log phase (Exponential growth phase) competefor 3. Stationary phase space nutrents ,  Flat portion of the curve  No net increase in cell number, but cells are still growing  Number of newly divided cell = Number of dying cell metabolzc  A period of equilibrium waste  Approach the carrying capacity with accumulation of waste & space 4. Death phase (Decline phase)  The population drops, with decrease in the number of living cells  Number of newly divided cell < Number of dying cell  Lack of nutrient support & accumulation of waste products 10  cunfovonroble condneion ) Bacteria -- Endospores [Sporulation] Usually in Gram +ve bacteria Form under adverse environmental condition (sporogenesis)  e.g. Nutrient depletion, Bacterial Spore Formation Animation Video extreme temperature or pH https://www.youtube.com/watch?v=NAcowliknPs Can survive under extreme heat, lack of water, exposure to chemical and radiation Dormant state for bacteria Can re-grow under favourable conditions (germination) 11 Created with BioRender.com Method Measuring Bacterial Growth Ho m e Se a rc h Bo oks he lf Se tti Eb ook Cen tr al™ A. Direct measurement methods  Counting the number of actual bacteria cells physically 1. Direct microscopic count (DMC)  Counts bacterial cell number under microscope using cell counter ⑦ Textbook: Microbiology Fig. 6.20 2. Serial dilution and plate count method  Counts the number of bacterial colony grown on agar plate (pronne nuynents)  Calculate the bacterial number by multiplying with dilution factor B. Indirect measurement methods 성 췄쏟 밌포포니 Textbook: Microbiology Fig. 6.16 12 Method Measuring Bacterial Growth (cont’d) Turbrd clear A. Direct measurement methods tzbacterra B. Indirect measurement methods  Not counting the actual number of bacteria, but the effect of bacteria cells on the physical properties of the culture medium (e.g. turbidity)  Turbidity estimation of bacteria number  Bacterial growth in liquid culture broth turns the solution turbid (cloudy) 며 없 □ Textbook: Microbiology Fig. 6.21  Bacteria (live or dead) in the liquid scatter the light passing through  ^ A spectrophotometer is an instrument for measuring turbidity (spectrophotometry) )  Turbidity is measured in terms of absorbance (A) or optical density (OD) measures laght  More cells  More scattering pass through the  Less light being transmitted  High Absorbance specimen 13 Factor Affecting Microbial Growth Physical Requirements: Temperature pH Osmotic pressure Chemical Requirements: Carbon Nitrogen Phosphorus Sulphur Trace elements Oxygen ) fon bacterra togrow Toxncsusstanceinher bacternal groamen 14  Physical Requirement -- Temperature bulish cow meta. Different microorganisms have different requirement for corspeea  Minimum temperature Tnautve o Bortern  Growth no longer occurs below this temperature [min. temp. for bacterial growth]  Optimum temperature  The most rapid growth occurs [bacteria grow best]  Maximum temperature  Growth is not possible above this temperature [kill the bacteria when over the max. temp.] Denatured Lower uppev Temp https://ucanr.edu/blogs/blogcore/postdetail.cfm?postnum=26278 ↓ repyratoy H, w ghe. Bactev 개 Temp resnroty il 15 、 」  Physical Requirement -- Temperature (cont’d) Classification according to temperature requirement: Low temperature/cold  Psychrophiles W  Can grow at 0 – 10oC, optimum temp. at ~10oC  Psychrotrophs  Optimum temp at ~20oC  Mesophiles Middle range  Can grow at 25 – 40oC, optimum temp. at 37oC Textbook: Microbiology Fig. 6.2  Thermophile Hot temperature  Optimum temp. at ~50-60oC  Hyperthermophiles Super hot temperature  Optimum temp. at >80oC Toxin that releases by some toxic bacteria cannot be removed even the toxic bacteria is killed by high temperature Textbook: Microbiology Fig. 6.1 16 Physical Requirement -- pH Different microbes have different requirement for minimal, optimal & maximal pH to grow Grow better in acidic ph Acidophiles  Grow best at low pH (acidic environment) Alkaliphiles  Grow best at high pH (alkaline environment) Extreme pH can kill the microbes by  Inactivating (Denaturing) proteins  Disrupting transport in the plasma membrane  Destroying enzymes in the cytoplasm Bacteria usually grow best at neutral (~pH 6.5 – 7.5)  In general, pathogens do not grow, or grow very slowly, at < pH 4  Molds & yeasts will grow over a greater pH range than bacteria 17 Physical Requirement -- Osmotic Pressure Salt (as solute)  osmolarity of surrounding environment Microorganisms require water for growth  Water account for 80 – 90% of their composition High osmotic pressure has the effect of removing necessary water from a cell (water move out of the cell)  [Salt] in solution > [Salt] in cell (hypertonic solution)  Water moves out  Osmotic loss of water causes plasmolysis or shrinkage of the cell’s cytoplasm Must t sa Obligate halophiles  Grow well in high salt concentration  e.g. Microbes found in Dead Sea Both Textbook: Microbiology Fig. 6.4 1 Facultative halophiles  Do not require high salt concentration, but can grow at moderate salt level (~2%) 18 Chemical Requirement Carbon  Universal source of energy needed by all living organisms  For synthesis of all organic molecules to build cell structures  Carbohydrates (e.g. glucose, starch) are the major energy source Nitrogen  For synthesis of DNA, RNA, proteins, etc. Phosphorous  For synthesis of DNA, RNA, ATP, membrane phospholipids Sulphur  For synthesis of proteins Trace elements  Needed in small amounts  As enzyme cofactors, growth factors  Important for cellular functions, e.g. energy production, metabolism 19 Chemical Requirement (cont’d) Oxygen Aerobes  Require O2 to grow a. Obligate aerobes  O2 must present ratmosphernclevel b. Microaerophiles  Require O2, but lower than atmospheric level Anaerobes  Growth is inhibited by O2 c. Facultative anaerobes  Can grow with / without O2 (grow better with O2) d. Aerotolerant anaerobes  Cannot use O2, but tolerate the presence of O2 e. Obligate anaerobes  Poisoned by O2 (O2 is toxic) 20 PART 2: Control of Microbial Growth  Terminologies  Physical methods  Chemical methods 21 Purpose to Control Microorganism Growth To kill, control, slow down the growth of microorganisms To prevent infections To reduce transmission of diseases To maintain a good health / hygiene To achieve an aseptic environment for clinical / surgical / laboratory practices 22 Terminology for Controlling Microorganism Growth -cide / -cidal (suffix) Methods  = To kill  e.g. Bactericidal = kill bacteria :햇업 Sterilization -static / -stasis (suffix) Reduction in microbes MOsT tA 에  = To stop or steady *311 , ; 로걸 시브 … , ,  Stop the growth of microorganisms, Disinfection NOT kill : FXP 묻 XAIl tRemwreae lowlevel  Growth might resume Sanitization  e.g. Bacteriostatic Cleaning 23  Terminology for Controlling Microorganism Growth (cont’d) Control Method Reduction Level Methods Examples Sterilization o Destruction or elimination o Heat sterilization o Autoclave of ALL living o Using chemical - High temperature microorganisms, sterilizer (121oC) + High pressure incl. endospores (15psi) for 15 minutes Disinfection o Elimination MOST of o Chemical o Using disinfectant to / Antisepsis vegetative microorganisms, disinfection perform decontamination EXCEPT endospores o Bleach vs o Using antiseptic to perform o ≠ Complete sterility 70% Alcohol antisepsis o Disinfection kills microbes on non-living things o Antisepsis kills microbes on living object (non-toxic) Sanitization o Reduce bacteria number o Chemical o Using sanitizer to reduce to a safe level microbial number o Using soap to wash hands 24 「 Methods for Controlling Microbial Growth A. Physical Methods B. Chemical Methods 1. Heat sterilization 1. Low antimicrobial power by thermal destruction a. Soap and detergent  Moist heat b. Heavy metal a. Autoclave b. Pasteurization 2. Intermediate antimicrobial power c. Boiling c. Phenolic compounds  Dry heat d. Halogen d. Direct flame e. Alcohol e. Hot-air oven 3. High antimicrobial power 2. Irradiation f. Oxidizing agents a. X-ray / Gamma ray radiation g. Aldehyde b. UV-light radiation >- Damuye BwA mutation 25  Physical Methods -- Heat Sterilization Heating  A universally well-accepted, cheap & easy method to control microorganism  High temperature can denature protein  -cidal effect cess cess Time I Temp ) less Efforts Moist heat is more effective than dry heat  ∵ Penetrates materials much more rapidly than air + Water molecules conduct heat better than air   Moist heat sterilization requires relatively lower temperatures & shorter exposure time when compared to dry heat sterilization 26 」  ^ Moist Heat -- Boiling 4 Stevy inme hetwod. Boil object in boiling water at 98 – 100 oC Heat inactivates microorganisms by denaturing protein  10 minutes can inactivate most vegetative cells, but NOT heat-resistant forms e.g. endospores  Some fungal spores & viruses may require up to 30 minutes  Bacterial spores often require > 2 hours Common method to “sterilize” drinking water 27 、  Moist Heat -- Autoclave t Autoclave " ☆ 평했습 Global standard for absolute control of microbial growth 니도  Steam under pressure (use moist heat + pressure)  15 pounds / square inch (psi) (~1.1 kg/cm2)  At 121oC for 15 mins  Most common sterilization method for materials not damaged by heat  Examples  Use in hospital  Instruments, glassware, intravenous solutions Textbook: Microbiology Fig. 7.2  Use in laboratories  Instruments, culture medium 28 「 Moist Heat -- Pasteurization Named after the great microbiologist Louise Pasteur Reduce the number of non-spore forming bacteria & heat-sensitive microbes in milk & juice Does NOT kill all microorganisms  Only disinfection (not sterilization) Can keeps quality of the milk and juice, retains flavor Used to control pathogenic bacteria in milk or juice  e.g. Prevent food spoilage caused by Salmonella spp., E. coli O157:H7 High-temperature short-time (HTST) pasteurization  72oC for 15 sec [milk sold at 4oC] Ultra-high-temperature (UHT) pasteurization  135oC for 2-5 sec [milk sold at 25oC] 29 Dry heat -- Direct Flame Kill by burning  Oxidation e.g. To sterilize inoculating loops & the opening of test tube during bacterial culture [Try it in the lab session] Dry heat -- Hot-air Oven Very high dry temperature  e.g. 171 oC for 1 hour or 160oC for >2 hours Used for sterilizing metal instruments, powders materials  Moisture is undesirable 30 Physical Methods -- Irradiation Radiation kills microorganisms by damaging DNA & protein Strength depends on its  Wavelength  Intensity  Duration Textbook: Microbiology Fig. 7.5 31 Physical Methods -- Irradiation (cont’d) mutation i X-ray / Gamma Ray Radiation A T UV-light Radiation X - Higher energy ray Cu kF Lower energy ray Stronger effect Weaker effect Ionizing radiation  Cause DNA breakage 졌챠 to cause microorganism death Non-ionizing radiation  Cause DNA mutation to cause microorganism death t , High penetration power Poor penetration power 어깝 e e. on surface  Only microorganisms on the surface are susceptible to destruction  Only useful for controlling surface contaminants Sterilization of interior structure of Commonly used in instruments, e.g. needles hospital operating rooms & sinks, aseptic filling rooms of pharmaceutical companies, laboratory culture chamber 32  Chemical Methods -- Low Antimicrobial Power Pw Soap & detergent sauntukator phinc , uphobc  Common sanitizer to reduce number of microbes  Physical rubbing off the bacteria 9 999. Heavy metal Otuna nembrane  Interfere with microbial metabolism Q 우 o to inhibit growth & kill microbes  e.g. Sliver, mercury, copper ¢ ㆀ Textbook: Microbiology Fig. 7.8 33 」 Chemical Methods -- Intermediate Antimicrobial Power Phenolic compounds 몹  Disrupt plasma membrane & denature protein to kill microbes  With odour smell  Examples  Hexachlorophene: hand washing agent for surgical uses  Triclosan: active ingredient in toothpaste rNot sood freeradzal Halogen fm크본  Forming radial that can oxidize microbes  Examples  Chlorine (e.g. hypochlorite ion CIO-) in bleach  Iodine as antiseptics for wound cleaning  Fluorine as antiseptics in toothpaste & drinking water 34  Chemical Methods -- Intermediate Antimicrobial Power (cont’d) Alcohol  Disrupt plasma membrane & denature protein to kill microbes  Effective concentration: 60% - 95%  Most effective concentration: 70%  70% alcohol can penetrate bacterial cell wall  Denature proteins & enzymes inside the cell Ifry  100% alcohol has poor penetrating power  Can only denature proteins ¤ on cell wall surface frxny baltenn  -cidal activity will be lost once evaporated Textbook: Microbiology Table 7.6 35 」 B3. High Antimicrobial Power Oxidizing agents  Denature protein by oxidization  Examples  Hydrogen peroxide H2O2  Ozone O3 Aldehyde 쁩  Very high power  Fix protein & nucleic acid  Used in preserving biological specimen, preparing vaccine & histological specimen for medical examination  e.g. 2% - 8% formaldehyde (formalin) [carcinogenic] 36 PART 3: Interactions between Microbes & Human  Normal microbiota  Pathogenic microbes  Mechanism of pathogenicity 37 Relationship between Normal Microbiota & the Host *  Microbiota or normal flora (microflora) Ithaseunes Human body is inhabited by many different bacteria benesthaal Relationship between normal microbiota & the host illustrates symbiosis  One organism is dependent on the other Colonization 춤 Pb  Presence of bacteria on body surface without causing disease  Either  Externally on skin surface; or  Internally on mucosal surface Examples of Microbiota on Human ⑦☆  Skin: S. epidermidis  Eye, mouth, nose: S. epidermidis, S. aureus  Gastrointestinal tract: Various gut microbiota for fermentation e.g. E.coli  Urogenital tract: Lactobacillus (maintain low pH in the vagina) L, sevetl And - cowply s 38 에 Textbook: Microbiology Table 14.1 39 Beneficial Effects of Microbiota 1. As a barrier to inhibit pathogenic bacteria  A phenomenon called competitive exclusion  By pathogen displacement, nutrient and oxygen competition, altering pH  Microbiota secrete harmful substances against the invading microbes 2. Stimulates immune response t 9. Furg ' ? sewets  Induce production of immunoglobulins (Ig) (antibodies) peneuhn  Enhance immune system development in newborn 3. Improves digestion (in gut only)  Gut microbiota acts as probiotics  Bacterial fermentation of plant carbohydrates facilitates digestion, stimulates peristalsis, prevents constipation  Fermentation also produces nutrients  e.g. Short chain fatty acid (SCFA), vitamins B2, B9 (folic acid), B12, K 40  Interaction between Pathogens & Human Pathogens  Microorganisms that can cause diseases in human Infection  The entry & development or multiplication of an infectious agent in the body that causes disease Levels of infection  Subclinical / inapparent infection (asymptomatic) i 뛰 : Latent infection (inactive  active & produce symptoms)  Clinical infection (symptomatic) 41 “Harmfulness” of a Pathogen Pathogenicity 도 춰가는  The inherent ability of a microorganism to cause disease to its host during the host-pathogen interaction  e.g. Normal E.coli without causing diarrhoea vs highly pathogenic O157 E.coli that can cause diarrhoea Virulence 빽  Degree of pathogenicity (disease-producing power) of a microorganism  e.g. Viruses that cause common cold with less severe symptoms vs influenza A with severe symptoms uthe β )  A pathogenic microorganism can be of high or low virulence nfmenra  A non-pathogenic microorganism can never be of high virulence 42 Mechanism of Pathogenicity How is a disease / infection being developed? 1. Portal of entry 2. Adhere 3. Defense / survive in host 4. Invade 5. Damage 6. Portal of exit 43 Textbook: Microbiology Fig. 15.9 Microbial Mechanism of Pathogenicity (cont’d) 1. Portal of entry  Pathogens usually entry host through  Skin Contact with the  Mucosal membrane environment on urogenital tract, digestive tract, respiratory tract, conjunctiva  Different pathogens have different preferred portal of entry  e.g. COVID-19 – respiratory mucosal membrane 2. Adhere  After getting into the host  The pathogen should get adhere onto the cell / tissue  Recall:  Bacteria: pili, capsule  Virus: viral spike, surface protein  Fungi: surface protein  Parasite: hook, sucker 44 고답석[A Textbook: Microbiology Table 15.1 45 Mechanism of Pathogenicity (cont’d) 3. Defense / survive in host  Ability of the pathogen to defense the host’s immune system, & being survived within the host by  Avoiding phagocytosis (chemical nature of the capsule)  Producing pathogen’s molecules to interfere / suppress the host’s immune responses (e.g. Ig A protease – digests the antibody) 4. Invade  Ability of the pathogen to penetrate the tissue & spread in host’s body by  Producing enzymes to break down the integrity of supporting tissues to help pathogen spread  e.g. Invasins – cause rearrangement of nearby cytoskeleton 46 Mechanism of Pathogenicity (cont’d) 5. Damage  Cause damage to the host’s tissue by  Using the host’s nutrient  Helminths absorb host nutrient for survival  Causing direct damage ∞  Virus can kill the host cells or transform normal cells into malignant cells  Produce toxins  Bacteria produce exotoxin & endotoxin ^ …  Inducing hypersensitive reaction ☆면 고 모구종 47 Exotoxin vs Endotoxin abie tosecrete eθ Exotoxin Endotoxin From both gram +ve (more common) LPS on the cell wall of & –ve bacteria gram –ve bacteria Protein produced inside pathogenic bacteria Being secreted out through Released into the host after exocytosis / after cell lysis bacterial cell lysis Can cause serious disease & death Effects  Fever, hypotension, thrombosis, septic shock, etc. Most of them can be inactivated by heat Examples 좁어* 부에를  Diphtheria toxin, botulism toxin, 함 tetanus toxin, various food poisoning toxins (cholera toxin, Shiga toxin, Staphylococcus enterotoxin, etc.) 48 Textbook: Microbiology Table 15.3 49  Microbial Mechanism of Pathogenicity (cont’d) f t×75t ) 6. Portal of Exit  Ways by which bacteria leave the host  Allow pathogen to spread in a population  Usually same portal of entry & exit  Examples  Respiratory tract: droplet, expelled during coughing or sneezing  GI tract: Faeces  Chicken pox: discharge from blister 50 「 Recap Part 1: Microbial Growth  How microbes grow  Methods measuring bacterial growth  Factors affecting microbial growth Part 2: Control of microbial growth  Terminologies  Physical methods  Chemical methods Part 3: Interactions between microbes & human  Normal microbiota  Pathogenic microbes  Mechanism of pathogenicity 51 Reference Tortora, G.J., Funke, B.R., & Case, C.L. (2018). Microbiology: An Introduction (13th ed.). Pearson.  Chapter 3 Observing Microorganisms Through a Microscope  Chapter 4 Functional Anatomy of Prokaryotic and Eukaryotic Cells  Chapter 6 Microbial Growth  Chapter 7 The Control of Microbial Growth  Chapter 14 Principles of Disease and Epidemiology  Chapter 15 Microbial Mechanisms of Pathogenicity 52