microbio exam #2

metabolism

The total of all chemical reactions and physical processes taking place within cells

catabolism

larger molecules are degraded or broken down into smaller molecules, usually with the release of energy

• degradation

• exergonic reaction —> ex: aerobic respiration

anabolism

larger molecules are built from smaller ones, resulting in formation of cell structures + requires energy input 

• biosynthesis

• endergonic reaction —> ex: photosynthesis

two types of energy

1. kinetic: energy of motion

2. potential: stored energy

• cells use chemical energy stored in chemical bonds

Gibb’s free energy

chemical energy in chemical bonds of molecules

• +ΔG: absorb energy, reaction is not spontaneous

• -ΔG: release energy, reaction is spontaneous 

ΔG

= G of products - G of reactants 

• indicates whether the reaction releases or absorbs energy

energy coupling

using an exergonic reaction to drive an endergonic reaction 

• overall delta G is -, together reactions are spontaneous

components of ATP molecule

1. nitrogenous base (adenine)

2. ribose sugar

3. three phosphate groups (where the high energy of ATP comes from due to repulsion of negative groups, ΔG= -7.3 kcal/mol)

aerobic respiration

seen in most organisms

• pathways involved: Glycolysis, TCA (citric acid cycle), ETS

requires/ produces: Glucose, O2/CO2 and H20

• ATP per glucose: 36 or 38

anaerobic respiration

seen in some prokaryotes, not seen in eukaryotes

• pathways involved: Glycolysis, TCA (citric acid cycle), ETS

requires/ produces: Glucose & salts/ CO2 and reduced salts

• ATP per glucose: between 10-25

fermentation

some prokaryotes & some eukaryotes

• pathways involved: glycolysis & fermentation

requires/ produces: Glucose & no O2/ Acid or alcohol & sometimes gas

• ATP per glucose: 2

glycolysis

• Occurs in the cytosol of both eukaryotes & prokaryotes 

• Has two phases: energy investment & energy payoff 

• Glucose, 2 ATP, 2 NAD+ – glycolysis → 2 pyruvates, 4 ATP, 2 NADH

pyruvate oxidation & tricarboxylic acid cycle

crucial aerobic respiration steps in the mitochondrial matrix, converting pyruvate from glycolysis into energy

• Yield per 1 pyruvate: 1 ATP, 2 NADH, 1 FADH2, 3 CO2

• Location of TCA cycle:

  • Eukaryotes: mitochondrial matrix 

  • Prokaryotes: cytosol

oxidation

loss of electrons & H+

reduction

gain of electrons & H+

NAD+ / NADH

essential coenzymes found in all living cells, acting as a pair in redox reactions to drive cellular energy production and metabolism

• Glycolysis and TCA load electrons onto NAD+ to form NADH (NAD: Nicotinamide Adenine Nucleotide)

• NADH delivers the electrons to the ETS and is oxidized back into NAD+

• FAD/FADH2 works the same way (FAD: Flavin Adenine Nucleotide)

what is the final electron acceptor in the electron transport system in aerobic respiration?

oxygen which gets reduced to water

chemiosmosis

the movement of ions (𝐻+ or protons) down their electrochemical gradient across a semipermeable membrane, powering ATP synthesis

movement of the protons across an ___ causes the formation of ATP

ATP synthase

__ATP made per NADH
__ ATP made per FADH2

3, 2

in prokaryotes, ETS proteins are located in ___

the cell membrane

in prokaryotes, protons are pumped into ___

the periplasmic space

what is the final electron acceptor in the electron transport system in anaerobic respiration?

inorganic salt
- the amount of ATP produced depends on the salt being used

glycolysis pathway

1 glucose to yield 2 pyruvate, 2 ATP, and 2 NADH

fermentation pathway

• stays in the cytosol

• uses pyruvate and NADH to yield NAD+, acids, or alcohols and sometimes gas

fermentation produces __ ATP per glucose

• 2

• … along with acid or alcohol and sometimes gad

carbohydrate fermentation media

• Contains specific sugars that can be fermented and converted to acids

• Contains a pH indicator (usually phenol red) to show the reaction 

• Inverted Durham tube inside helps capture gas if produced during fermentation of specific sugar 

• Used for differentiating and identifying bacteria based on which sugars they can or can’t ferment

genome

the total of all genetic material within an organism

chromosome

length of neatly packaged DNA containing genes

gene

fundamental unit of heredity responsible for a given trait + a specific segment of DNA that codes for a protein or RNA molecule 

genetics

the science that studies the inheritance of biological characteristics by living things 

DNA

(deoxyribonucleic acid) is the central molecule of genetics 

nucleotide

• the basic unit of DNA 

  • Made of three parts: phosphate group, deoxyribose sugar, nitrogenous base 

  • Bound by a sugar-phosphate backbone 

6 steps of DNA replication

1. Begins at the origin of replication (ORI)

2. Helicase unwinds the DNA double helix and forms a replication bubble 

3. An RNA primer is synthesized by primase

4. DNA poly III adds nucleotides in a 5’ → 3’ direction 

• The leading strand can be synthesized continuously because DNA poly III can add nucleotides in the 5’ → 3’

• The lagging strand is not synthesized continuously because DNA poly II must add short fragments (Okazaki fragments) that are 5’ → 3’ one at a time 

5. DNA poly I replaces the RNA primers with complementary DNA

6. DNA ligase seals gaps in the new DNA so it is identical to the parental strand 

DNA replication

• bidirectional

•  semiconservative because each replicated chromosome ends up with one new strand of DNA and one old strand 

DNA polymerase III

• is the primary replication enzyme responsible for deciphering and duplicating the DNA code

  • Can not add nucleotides to DNA without the presence of a primer inserted in the ORI site

role of primase in DNA replication

the enzyme responsible for making RNA primers to serve as starting points for DNA poly III

role of helicase in DNA replication

Helicase unwinds DNA double helix

role of DNA poly I in DNA replication

DNA poly I replaces RNA primers with complementary DNA

role of ligase in DNA replication

Ligase seals any gaps in sugar phosphate backbone of DNA 

role of DNA polymerase III in DNA replication

DNA poly III adds nucleotides in to 5’ → 3’ direction 

bacteria divide through:

asexual reproduction via binary fission: formation of two new cell of approximately equal size as the result of parent division  

vertical gene transfer

DNA is passed from one generation to the next 

horizontal gene transfer

transmit DNA between cells “of the same generation”

• Can be accomplished by conjugation, transformation, and transduction 

conjugation

• mode of genetic transmission in which a plasmid or fragment of chromosomal DNA is transferred from a donor cell to a recipient cell

• requires the attached of two related bacterial species and the formation of a bridge (sex pilus) that can transport DNA 

transformation

• uptake of small fragments of “naked” extracellular DNA by “competent” bacteria from their surrounding environment

• can occur naturally or artificially (electroporation or heat shock treatment) through recombinant DNA technology techniques 

transduction

• process by which a bacteriophage mediates transfer of DNA from a donor cell to a recipient cell

• a newly assembled phage (termed a transducing phage) accidentally incorporates a piece of the host bacterial DNA and later injects that DNA into another host cell

doubling time

• The length of time it takes for one cell to become 2 is called doubling time and varies between species and environmental conditions

  • # of bacteria x 2ⁿ

ways to measure bacterial growth

• spectrophotometer (measure turbidity quantitatively)

• cytometer (manual counting of cells) 

lag phase

none to little cell growth of bacteria as cells are adjusting to their environment 

exponential growth phase

bacterial cells are dividing at their maximal rate 

stationary phase

as nutrients are depleted, bacterial growth rate slows and equals the rate of cell death 

death phase

rate of bacterial death is faster, build up on toxins

environmental factors on bacterial growth: temperature

• Minimum: lowest temp that permits a microbe’s continued growth and metabolism

• Optimum: temp which promotes the fastest rate of growth and metabolism 

• Maximum: highest temp at which growth and metabolism can proceed

environmental factors on bacterial growth: pH

• Vast majority of organisms are neutrophils, prefer neutral pH environments (pH 6-8)

• Acidophiles: optimal pH below 6.0

• Alkalinophile: optimal pH of 8.0 or greater 

osmophiles

organisms that live in habitats with a high osmotic pressure (high solute concentration)

• Halophiles: live in high salt environments 

reducing medium

contains a substance that absorbs oxygen or slows penetration of oxygen into medium; used for growing anaerobic bacteria and determining oxygen preference of other bacteria 

microbes with the highest resistance

bacterial endospores —>  Clostridium, Bacillus, Sporosarcina

microbes with moderate resistance (5)

• Pseudomonas aeruginosa: thick slime layer

• Mycobacterium tuberculosis: has mycolic acid outer layer

• Staphylococcus aureus: very thick peptidoglycan layer

• Protozoan cysts: can survive in nature 

• Naked viruses (HAV, Norovirus)

microbes with least resistance (4)

• Most vegetative bacterial cells

• Protozoan trophozoites

• Enveloped viruses (HIV influenza)

• Fungal spores, hyphae, and yeasts

sterilization

 a process that destroys all viable microbes on inanimate objects, including viruses & endospores

disinfection

a process that destroys vegetative pathogens on inanimate objects, not endospores

sanitization

any cleansing technique that mechanically removes microbes from inanimate objects

degermination

reduces the number of microbes on the skin 

antisepsis

chemical agents are applied to the body to destroy or inhibit vegetative pathogens

influencing factors of microbial death

• Exposure time 

• Type of organism

• Microbial load: quantity of bacteria

• Agent’s action: microbistatic → stops microbes from growing, ex: refrigerator + microbicidal → kills microbes, ex: lysol  

• Organic matter: can slow down action of chemicals 

• Agent’s concentration: more concentration = more death

physical agents: heat —> moist heat

• uses hot water or steam 

  • Mode of action: denaturation of protein 

  • Hot water (65 deg C or greater) and boiling (100 deg C) achieves disinfection 

  • Autoclaving (steam under pressure) achieves sterilization (121 deg C/ 15 psi / 15 min)

pasteurization

• a technique in which heat is applied to liquids to kill potential agents of infection and spoilage + achieves disinfection not sterilization → 30 minutes at 66 deg C or 15 sec at 71.6 deg C (flash method)

physical agents: heat —> dry heat

• requires higher temperatures than moist heat but can also sterilize 

  • Dry ovens: 150-180 deg C → denatures proteins

  • Incineration: 800-1560 deg C → complete oxidation and combustion of cells

cold temperatures

Microbistatic: slows the growth of microbes (deadly for few microbes)

• Refrigeration: 0-15 deg C and freezing: <0 deg C

• Used to preserve food, media, and cultures 

dessication

• Dehydration leading to metabolic inhibition 

• Not effective microbial control, many cells will grow when water is returned 

• Used for increasing shelf-life of cultures

ionizing radiation

• Ionizing: deep penetrating power, breaks DNA

  • Gamma rays, X rays

  • Sterilizes medical supplies

  • Irradiated food products

nonionizing radiation

• Nonionizing: lower penetrating power, mutates DNA

  • Microwaves, UV rays

  • UV light allows disinfection of air, water and solid surfaces; causes thymine-thymine dimers which damages DNA

mechanical removal: filtration

• the physical removal of microbes by passing a gas or liquid through a filter with openings too small for microbes to pass through

  • Sterilizes heat and sensitive liquids and air in hospital isolation units 

    • HEPA: high efficiency particulate air filters remove particles ~ 0.3 microns

mechanical removal: sanitization

• mechanically removes microbes from inanimate objects to safe levels but not necessarily killing all pathogens

  • Can combine sanitization and disinfection to both kill and remove microbes

mechanical removal: degermation

• reducing the number of microbes on human tissue (ex: hand washing, toothbrushing, showering)

  • Can combine degermation and antisepsis to both destroy and remove microbes 

chemical control: halogens

Chlorine and Iodine

• The active ingredient in nearly ⅓ of all antimicrobial chemicals currently marketed

• Chlorine denatures proteins, microbicidal and can be sporadical at high amounts 

• Iodine denatures proteins,  microbicidal and can be sporadical at high amounts 

  • Iodophors: iodine attached to polymer which slowly releases iodine 

  • Tincture: chemically dissolved in alcohol

chemical control: phenolics

• Disrupt cell membranes and denature proteins, microbial but not sporicidal

• Phenol coefficient (PC): quantitatively compares a chemical’s antimicrobial properties to those of phenol 

  • PC= lowest concentration that kills in 10 min/ results for phenol 

    • PC > 1 indicates that the disinfectant is more effective than phenol 

chemical control: surfactants

Detergents and Soaps

• Detergents are soaps that act as surfactants 

• molecules that have a hydrocarbon chain

• Quaternary ammonia compounds (QACs) act as surfactants that alter membrane permeability of some bacteria and fungi 

chemical control: chlorohexidine

• Destroys cell membranes and denatures proteins

• Broad microbicidal activity but not sporicidal

• Exceptionally mild, low toxicity, and rapid action

chemical control: alcohols

Ethanol and Isopropanol

• Act as surfactants dissolving membrane lipids and denaturing proteins

• Microbicidal but not sporicidal

• Does destroy enveloped viruses 

chemical control: peroxygens

Hydrogen Peroxide 

• Weak (3%) to strong (25%)

• Produces highly reactive oxygen radicals that damage protein & DNA

• Broad microbicidal activity at low concentrations but strong solutions are sporicidal 

chemical control: heavy metals

Silver, Mercury

• Solutions of silver, copper, zinc, and mercury kill vegetative cells in very low concentrations by inactivating proteins 

• Broad microbicidal activity but not sporicidal 

chemical control: aldehydes

Formaldehyde, Gluteraldehyde

• Organic substances bearing a -CHO functional group on their terminal carbon

• Glutaraldehyde and formaldehyde kill by crosslinking to deactivate proteins and DNA

• Broad microbicidal activity and sporicidal at higher amounts 

chemical control: aldehydes

Ethylene Oxide 

• Strong crosslinking agent that inactivate proteins and DNA

• Sporicidal so they do reach sterilization 

• Used to sterilize many medical devices 

who discovered the first antibiotic?

Alexander Fleming in 1928

• He observed that the Penicillium fungus made an antibiotic: penicillin that killed Staphylococcus aureus

which bacteria and molds are commonly used in antibiotics?

• Bacteria in genera Streptomyces and Bacillus

• Molds in genera Penicillium and Cephalosporium

ideal antimicrobial drug

• selective toxicity

• long lasting potency

• easily delivered to the appropriate site of infection

• doesn’t disrupt the host’s health

• doesn’t encourage the development of antimicrobial resistance 

antibiotics

a naturally- produced chemical substance from one microorganism that can inhibit or kill another microbe 

synthetic drugs

antimicrobial compounds synthesized in the laboratory through chemical reactions 

targets of antibacterial drugs

1. Inhibition of cell wall synthesis 

2. Inhibition of protein synthesis 

3. Inhibition of nucleic acid synthesis, structure, or function 

4. Disruption of the cell membrane structure 

or function 

5. Blocks on key metabolic pathways

penicillin

• antibiotic

• Penicillins bind and block transpeptidase which cross-link the glycan molecules, causing breakdown of peptidoglycan and bacterial cell lysis 

• The original penicillins were limited to only gram - bacteria

  • Narrow spectrum: antimicrobials effective against a limited array of microbial types

• Newer, semi-synthetic penicillins (ampicillin, amoxicillin) are effective against both gram + and gram - bacteria

  • Broad spectrum: antimicrobials effective against a wide variety of microbial types  

• Non toxic but can trigger allergies

how does penicillin resistance occur?

• Many bacteria now produce enzymes that destroy the beta-lactam ring of beta-lactam antibiotics. These enzymes are referred to as beta-lactamases 

• Penicillinase- producing bacteria are resistant to some penicillins 

• Newer generation penicillins aren’t sensitive to penicillinase, so they are used for treating penicillin-resistant infections (ex: methicillin, oxacillin)

cephalosporins

• antibiotic

• Similar to penicillins, also have a beta-lactam ring 

• Inhibit cell wall synthesis 

• Newer generation cephalosporins have a broad-spectrum and effective against gram + and gram - bacteria

vancomycin

• antibiotic

• Inhibits cell wall synthesis (blocks NAG & NAM binding)

• Narrow-spectrum antibiotic most effective in treating staphylococcal infections in penicillin-resistant strains (ex: MRSA and other gram + bacteria)

• Can be very toxic (nephrotoxic, hepatotoxic, and ototoxic) and restricted to the most serious, life-threatening conditions 

• VRE is a growing nosocomial concern. Incidences of VRSA are now increasing 

bacitracin

• antibiotic

• Inhibits cell wall synthesis (blocks transport of peptidoglycan to cell wall)

• Narrow-spectrum antibiotic used against gram + bacteria

• Major ingredient in common drugstore antibiotic ointment for combating superficial skin infections by Staphylococci and Streptococci

polymyxins

• antibiotic

• Binds to LPS and has detergent activity that disrupts the outer and inner cell membrane 

• Narrow spectrum antibiotic used against gram - bacillus bacteria 

• Also a common ingredient in drugstore antibiotic ointments (ex: Neosporin)

aminoglycosides

• antibiotic

• A group of antibiotics that inhibit protein synthesis by binding to the 30s ribosomal subunit 

• Have a broad-spectrum range against both gram + and gram - bacteria 

• Examples: streptomycin, neomycin, gentamicin, kanamycin 

tetracycline

• antibiotic

• Inhibits protein synthesis by binding to the 30s ribosomal subunit 

• A very broad spectrum antibiotic used against gram + and gram - bacteria 

• Use can be limited by its possible side effects: vomiting, diarrhea, kidney problems, etc

chloramphenicol

• antibiotic

• Inhibits protein synthesis by binding to the 50s ribosomal subunit and blocking peptide bond formation 

• Very potent broad-spectrum antibiotic 

• Can be very toxic to humans and its uses are restricted