Microbiology Exam 2 Study Guide PDF

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Microbiology Exam 2 Study Guide Chapter 11 – Controlling Microbes 1. Approaches to Control Microbes a. Physical Methods i. Dry 1. incineration or dry oven  sterilization ii. Moist heat 1. Boiling water, steam under pressure  sterilization 2. Kills microbes by denaturing enzymes 3. Ex. Autoclaving iii. Radiation 1. Irradiation – bombardment with radiation 2. Ionizing – can penetrate a solid barrier, bombard a cell, enter it, and dislodge electrons form molecules a. x-ray, cathode, gamma  sterilization 3. Nonionizing – enters a cell, strike molecules, and excites them. Results? Mutation of DNA a. Note: non-ionizing radiation cannot penetrate a solid barrier b. UV  disinfection iv. Chemical Methods 1. Antimicrobial chemicals  disinfection or sterilization 2. Gases  disinfection of sterilization v. Mechanical Methods 1. Filtration of air and liquids  decontamination 2. Physical Controls a. Heat i. Moist heat – boiling water (100oC, 30 min) ii. Pasteurization – milk, fruit juices; flash method (~72oC, 15 sec) iii. Pressurized steam – autoclave (121oC, 15psi); used for surgical instruments, commercial canning (clostridium botulinum, problem in canning) b. Radiation i. Gamma radiation – DNA 1. Medical equipment, drugs, food-safe ii. Ultraviolet radiation – DNA iii. Microwave – heat

3. Factors in Treatment a. Situation – home, hospital, lab, factory i. Sepsis – the growth of microorganisms in the blood & other tissue b. Surface or medium c. Type and number of microorganisms i. Highest resistance = endospores ii. Moderate resistance = mycobacterium, S. aureus iii. Least resistance = non-endospore formers d. Environment e. Concentration of agent being used f. Mode of action 4. Mechanical Controls a. 2 kinds i. Fluid filtration ii. Air filtration – HEPA; hospitals b. Decontaminating Congress – letters containing Bacillus anthracis were opened in Hart Office of US Senate in 2001 i. B. anthracis is spore-forming – eradication is tough; building was heavily populated, needed to decontaminate heating/AC vents, carpet, furniture, office equipment, sensitive papers, artwork, personal belongings c. Decontamination Process i. Size/scope – samples taken from 25 buildings ii. Decontamination – vacuum with HEPA filter followed by tx with liquid chlorine dioxide (sterilant used for treatment of medical waste) or a decontamination foam; heavily contaminated areas used gaseous ClO2 5. Chemical Controls a. Factors for choosing appropriate germicide: storage/stability, residue, cost/availability, environmental risk b. Germicidal Chemical: i. Alcohols, hydrogen peroxide (H2O2), ethylene oxide (used on medical instruments, plastics; sugar, spices) 6. –static? or –cidal? a. Bactericide – a chemical that destroys bacteria except for those in the endospore stage b. Germicide/Microbicide – chemical agents that kill microorganisms c. Bacteristatic – prevent the growth of bacteria on tissues or on objects in the environment d. Fungistatic – inhibit fungal growth e. Microbistatic – antiseptics & drugs (used in the body) f. Degermination – reducing the number of microbes on the skin (ex. hand sanitizers)

7. How to determine Microbial Death a. Permanent loss of reproductive capabilities, even under optimum growth conditions, is the accepted microbiological definition of death b. Factors that influence death rate – time, concentration of the antimicrobial agent 8. Effects of Antimicrobial Agents – cellular targets of physical and chemical agents can be categorized as: a. Cell wall i. Wall maintains the structural integrity of bacterial and fungal cells, chemical agents damage the cell wall by blocking its synthesis, digesting it, or breaking down its surface ii. Cells without functioning cell walls become fragile and susceptible to lysis iii. Detergents and alcohols disrupt cell walls (especially in Gm- bacteria) b. Cell Membrane i. Membrane is composed of lipids and proteins, many viruses also have outer membranous envelopes ii. If a membrane is disrupted, leads to loss of selective permeability 1. Loss of vital molecules; damaging chemicals can enter the cell now iii. Surfactants: detergents that can work as microbicidal agents; polar molecules with hydrophilic and hydrophobic regions that can physically bind to the lipid layer; poke holes into cell membrane c. Cellular Synthetic Processes (DNA, RNA) and Proteins i. Like eukaryotic cells, prokaryotes rely on the central dogma ii. Antibiotics bind to the ribosomes of bacteria to prevent peptide bonds from forming; prohibits growth and metabolism iii. Similarly, for DNA, radiation, formaldehyde, and ethylene oxide interfere with DNA leading to mutations and inactivation 9. Effects of Cold and Desiccation a. COLD – slows down growth of cultures and microbes, some microbes are killed by cold temps but most will survive months in the cold b. DESSICATION – dehydration, die a few hours after air drying c. Note: lyophilization is the method of preserving microorganisms for many years (no ice crystals form, viable after freeze-dry)

10. Triclosan – mild and non-toxic, kills most pathogenic bacteria but NOT viruses and fungi, linked to cases of skin rashes due to hypersensitivity

a. many pathogens are naturally resistant to triclosan, including: mycobacterium tuberculosis, pseudomonas, e. coli, s. aureus b. Overuse of environmental microbicides will alter our normal flora; when bacteria become resistant to antibacterial agents, they simultaneously might become resistant to antibiotics c. to reduce, use “happy medium” i. don’t fill home with germicidal products (furniture, carpet, paint, hand sanitizer) ii. instead use traditional soaps and detergents that denature the bacterial cell to kill and bacteria will not become resistant to these

Chapter 9 – Microbial Genetics 11. Genetics and Genes a. Genetics – the study of heredity; heredity – the study of inheritance i. Transmission of biological properties (traits) from parent to offspring ii. Expression and variation of those traits iii. Structure and function of genetic material iv. How the genetic material changes b. Genome – sum total of the genetic material of an organism, composed of DNA (viruses are exception) i. Bacteria and some fungi contain tiny extra pieces of DNA called plasmids ii. Certain organelles of eukaryotes (mitochondria & chloroplasts) have their own DNA c. Chromosome – discrete cellular structure composed of a neatly packaged DNA molecule i. Eukaryotes – DNA wrapped around proteins called histones to form chromosomes (located in the nucleus) ii. Prokaryotes – condensed and secured into a packet by means of histonelike proteins (single, circular double-stranded chromosome) d. Gene – a fundamental unit of heredity responsible for a gene train in an organism i. 3 types: 1. structural genes – code for proteins 2. regulatory genes – control gene expression 3. genes coding for RNA machinery during protein production e. Size & Packaging of Genomes i. Viruses: 4-5 genes ii. Bacterium: one chromosome with 4288 genes iii. Human cell: ~30,000 genes on 46 chromosomes

12. DNA Structure – Nucleotides

a. Nucleotide – basic unit of DNA; chromosome consists of several million nucleotides linked end to end i. Composed of: 1. Phosphate 2. Deoxyribose (5-carbon) sugar 3. Nitrogenous base (joined by hydrogen bonds) a. Purines – double ring structure; adenine (A) & guanine (G) b. Pyrimidines – single ring structures; thymine (T)/ uracil (U) & cytosine (C) c. A  T; C  G ii. Nucleotides bind together in a specific fashion to form the sugarphosphate backbone of DNA; phosphate group binds on to sugar group of another nucleotide 1. Phosphate group called 5’ end because phosphate is attached to number 5 carbon on the sugar 2. OH (hydroxyl group) called 3’ end because hydroxyl group is on number 3 carbon on sugar iii. Joining Nucleotides 1. OH is removed from sugar and an H is removed from OH on the phosphate group forming a covalent bond between them a. Bond specifically called phopshodiester bond 2. Formation of bonds allow for synthesis of sugar-phosphate backbone iv. Once the 2 sugar phosphate backbones are made, nitrogenous bases of each nucleotide bend to each other via hydrogen bonds making them anti-parallel 1. 2 sugar-phosphate back bones come together in an opposite orientation v. DNA vs. RNA 1. Both are made of nucleotides (sugar, base, phosphate); both are also connected by covalent phosphodiester bonds between sugar and phosphate 2. DNA uses deoxyribose and RNA uses ribose; DNA is doublestranded whereas RNA is single-stranded; DNA contains thymine and RNA contains uracil 13. DNA Replication a. In replication, original DNA of cell is used as a template to make duplicate copy of DNA so the new cell has exact same sequence of DNA as original i. Cell wants to replicate DNA when it is dividing so that each new cell has cope of DNA identical to DNA of original cell b. Steps of replication: uncoil parent DNA molecule; unzip hydrogen bond between base pair to allow for separation of strands and expose bases to serve as

c. d.

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templates; synthesize 2 new strand by attachment of correct complementary nucleotides to each ss template Key enzymes: helicase, primase, DNA polymymerase III, DNA polymerase I, DNA ligase, Beginning of DNA Replication i. DNA replication is semiconservative (each old strand of DNA is used as a template for a new strand) so the 2 strands must be separated from each other so can be used as template ii. Replication starts at a specific site called origin of replication – at this site 2 strands of DNA are separated by enzyme called helicase (which essentially unzips the DNA molecule to separate strands) iii. Replication is bidirectional – can occur in 2 directions After DNA strand separation, replication proceeds i. DNA polymerase III is enzyme that adds nucleotides to growing nucleotide chains 1. Does so in 5’  3’ fashion; 5’ phosphate is added to 3’ OH on the sugar 2. DNA polymerase must have free 3’ OH to attach to new nucleotide in order to continue making sugar-phosphate backbone ii. When DNA are first separated, there’s no nucleotide chain for new DNA strand to bind to and build on 1. An enzyme called primase synthesizes a small piece of RNA (called RNA primer) that has a free 3’OH end 2. RNA sequence will have complementary bases of RNA to the DNA sequence it matches; however, it will have uracil not thymine so the new DNA strand can attach to it Synthesis of Leading and Lagging strand i. Since using both strands as a template, one is leading strand and one is lagging strand; mechanism for replication is little different for each strands 1. DNA polymerase III synthesizes in 5’ to 3’ direction, so since semiconservative, template must form 3’ to 5’ so DNA polymerase III begins at 3’ end of leading strand and synthesizes to 5’ ii. Enzyme helicase binds to origin of replication then unwinds strands by breaking hydrogen bonds between the nitrogenous bases iii. Primase makes an RNA primer then DNA polymerase III begins synthesizing the new DNA strand in 5’ to 3’ direction

iv. At this point, DNA polymerase iii has free 3’ OH end so continues to snythensize new DNA strand, however lagging strand does not have free 3’OH end, only has free 5’ end which wont work for DNA polymerase III

1. Therefor primase makes another RNA primer so now will ve free 3’OH end 2. DNA polymerase III will then synthesize another short fragment of DNA called Okasaki fragments v. When synthesis of both strands is complete, DNA polymerase I replaces RNA primer with a DNA sequence vi. For final step, DNA ligase seals the osasaki fragments together on lagging strand and seals gaps between new DNA that replace the RNA primer and the newly synthesized strand g. Steps to DNA replication i. Helicase binds to origin of replication; helicase unzips DNA by breaking hydrogen bonds; primase makes RNA primer to have free 3’ OH; DNA polymerase III synthesizes new DNA strand (complementary strand to template); DNA polymerase I replaces RNA primer with DNA; ligase seals DNA fragments together Chapter 9 pt. 2 14. Transcription and Translation a. Transcription – gene (DNA sequence) is transcribed into RNA (RNA sequence); a gene or several genes of DNA are transcribed to make mRNA i. Gene – short sequence of DNA that encodes a protein ii. Genome – DNA/chromosome of an organism consists of different segments of coding sequence called genes b. Translation – mRNA (RNA sequence) is translated into amino acid sequence; mRNA is used to make proteins for the cell c. Central Dogma of Molecular Biology – central body of thought for how the cell gets proteins; transcription and translation together lead to formation of protein 15....