micro
· Louis Pasteur- developed some of the first vaccines
· Discovered microbial basis of fermenting
· *oxygen doesn’t enable spontaneous generation of microbes
· Made and sold alcohol to fund his research
· “spontaneous generation”: all u need is air to create living things, like poof came out of thin air. he tried to disprove this
o An example could be mold, making it “spontaneous” but its not tho
· There were doubts about his exp
· John tyndall- wanted to find a discrepancy
· There was heat resistant bacteria in other people’s experiments, but Pasteur’s experiment didn’t have those kinds of bacteria, so they essentially burned out
· cells come from other cells
· The “golden age” came about after this theory came to light.
· People began studying microorganisms, they worked on viruses, many studies were conducted, to find out why people were getting sick. These findings came to initiate prevention and treatment of disease.
· Microbes have affected people both positively and negatively, like helping scientists discover more things, but also killing ppl
· Not all microbes are bad
o The good ones can be in probiotics
· Penicillin was discovered by Alexander Flemming this was an accidental discovery. By leaving a plate uncovered, or smth, and when he returned 5 days later, noticed a growth had appeared, it was mold. And there were no bacteria surrounding the mold, as if it makes smth inhibiting bacterial growth.
· Mary, the “mother of penicillin”, found a melon covered in mold, wanted to see if it had high amounts of penicillin, or if it could muster a high amount. She bought it, took it to lab, and discovered that it did yield a high amount.
· 3 branches to the phylogenetic tree of life that microbes fall under
o Bacteria, archaea, eucarya
o Prokaryotes can include bacteria and archaea
· Prokaryotes don’t have a membrane bound nucleus, in turn, eukaryotes have a membrane bound nucleus + organelles
· When clarifying the genus, it should be italicized. And the species should always be lowercase, ex. Shigella -> S. boydii
· Bacteria
o Most common type in human infection
o Prominent features; rod-shaped, spherical, and spiral
o Rigid cell-wall
§ Made up of peptidoglycan
o Responsible for cell shape
o They multiply by binary fission
§ One cell dividing into two, then 4, etc etc. each cell will be identical to the first
o Some bacteria are motile, like having flagella
· Archaea
o Some traits shared w bacteria and eukarya
o Have cell wall, but not made of peptidoglycan
o Found in extreme environments like ice cold, or boiling hot
o Not known to cause disease in humans
· Eukarya
o All members are eukaryotic
o Algae, fungi, and protozoa. Can be multi or single celled
o Protis are only single celled, and microscopic
· Algae
o Contain chlorophyll
o Diverse- both single and multi
o Found in surface of water because they need the sunlight for photosynthesis
o Can come in the color red
§ Like the ‘red tide’
o Rigid cell wall
· Fungi
o Diverse single celled and multi
§ Single celled= yeast
· Multiply via budding
§ Multi= mold
o Gains energy from organic materials
o Found wherever organic materials are present
· Protozoa
o Microscopic, only single-celled
o Found in water & land
o Complex
o Larger than a prokaryote
o Don’t have rigid cell walls, so shapes can vary
o Gain energy from organic matter
o Most are motile
o Amoebas are green even tho they don’t do photosynthesis, because they ate smth green. And it shows up
· Viruses, viroids, prions
o Non-living elements
o Called ‘agents’
§ Not organisms bc theyre not live
o Consists of only a few molecules found in living cells
o Contain protein coat surrounding nucleic acid
o Some can have an extra layer of lipids, called an envelope
§ Protein bag of nucleic acid
o Depend on hosts to replicate
§ Inactive outside of host
· Viruses
o All lifeforms can be infected by viruses, us, animals, plants, and bacteria…and aliens?
o They frequently kill host cells
o Some live harmoniously w host
· Viroids
o Simpler than viruses
§ Still needs a host to replicate
o Consist of a single short piece of RNA
§ Contain no protective protein coat
o Viroids r smaller than viruses
o Generally, cause plant diseases, spread by insects
· Prions
o Infections proteins
§ Contain no nucleic acid
o Responsible for at least 7 neurodegenrative diseases
§ Animal diseases
· Scrapie in sheep
· Mad cow disease in cattle
§ Human disease
· Kuru
· Creutzfelt-Jacob
o All humans have prions, theyre just protein, but healthy duh
§ An infectious prion is a diff shape and goes around changing good proteins to nasty ones by changing their shapes
o Some ppl are born w certain prions
Chapter 3
· To the naked eye, we can see up to ~150 micrometers
· Microscopes are the most important thing in microbio (duh)
o Light microscopy
§ Light passes thru specimen, then thru series of magnifying lenses
§ Important factors:
· Resolution
· Magnification
· Contrast
o Resolution: the shortest distance between 2 points that can still be distinguished as separate points
§ Wavelength of light sed is major factor in resolution //shorter wavelength -> greater resolution
§ Our eyes can’t see prokaryotic microbes, because of the distance of our cones and rods cells in our eyes
§ (resolution) Oil immersion reduces light refraction
§
o Refraction: when light passes thru a lens, and how it bends the light
§ Lens w/ 2 differently shaped convex sides, magnifies image
o Contrast: reflects the # of visible shades in a specimen
§ Higher contrast achieved for microscopy through specimen staining
o Bright-field microscopy
§ Produces a dark image against a brighter background
§ Has several obj. lenses
§ Parfocal microscopes remain in focus when objectives are changed
§ Total mag.
· Product of the magnification of the ocular x objective
o Interference microscopy
§ Causes specimen to appear 3D
· Depends on differences in refractive index
§ Mostly used in differential interference contrast (DIC)
§ Used to see live organisms, mostly their surface
§ Phase contrast microscopy is to see 3D internally
o Dark-field microscopy
§ Reverse image
· Specimen appears bright on a dark bg
· Like a photographic negative
· Achieves image thru a modified condenser
· Can see external structures that are very thin
o Fluorescence microscopy
§ Used to observe organisms that’re naturally fluorescent, or tagged w/ fluorescent dye
§ Or using an antibody because they stick to stuff. Like Spiderman’s webs
§ So like a fluorescent molecule sticking to the protein
· If u wanna do it ona live one, u can genetically add the fluorescent.
o GFP- green fluorescent protein. Comes from jellyfish
o Fluorescent molecule absorbs uv light and emits visible light
o Image fluoresces on dark bg
o Confocal scanning laser
§ Used to construct 3D image of thicker structures
§ Provides detailed sectional views of internal structures of an intact organism
o Laser sends beam through sections of organism
o Computer constructs 3D image from sections
o Used for both live and dead
o Electron microscopy
§ Doesn’t use light at all
§ To look @ smth as small as a virus
§ Always dead specimen, never live
o Transmission electron microscopy
§ Always a dead sample
§ Needs to be small and thin sample
§ Used to observe fine detail
§ Directs beam of electrons at specimen
· Electrons lass thru or scatter at surface
· 2D flat
o Scanning electron m.
§ 3D pic
§ Used to observe surface detail
§ Specimen coated in metal, usually gold
· Visible light has wavelengths 400-750nm
o Max resolution is 1 wavelength
· Magnification spreads light rays out
o To 150 micrometers, resolution of our eyes
§ Distance between photoreceptor cells
· Atomic microscope
o Capable of seeing single atoms
§ Works similar to those pin toys where you push your hand onto it, and keeps the shape
· Bacterial shapes and Arrangements
o Cocci – spheres
§ Come in bunches, chains, pairs, or quartets
o Bacilli – sticks/rods
§ Come alone or in chains
§ They chain horizontally, because vertically would cause replication
o Vibrios – bent rods
§ Typically singles
o Irregular
o Spiral
· Similarities between prokaryote and eukaryote
o Both have DNA
o Both have ribosomes
o Both have plasma membranes
o May contain cell walls
o May contain flagella
· Differences
o Nucleus in eukaryote, nucleoid in prokaryote
o Membrane bound eukaryote
o Ribosomes smaller in prokaryotes, larger in eukaryote
o Cell walls made differently
o Compostion and movement of flagella are different
o Eukaryotes have organelles (membrane bound)
o Cilia in eukaryotes (?)
· Cytoplasmic Membrane
o Membrane us embedded w numerous protein
o Is selectively permeable
§ Determines what molecule may or may not pass through
§ Water goes through low to high solute concentration (osmosis)
§ Bacteria would add as a plant cell when it comes to diffusion
o Evolution
§ It is theorized that mitochondria came from prokaryotes
§ Another endosymbiosis event happened, where cyanobacteria resulted in chloroplasts
· What makes these two differ is that they have their own DNA
· So, they express differently even though they both came from endosymbiosis
o Transport proteins
§ Directed movement across membrane
· Facilitated diffusion
· Active transport
· Group translocation
§ Facilitated
· Moves w/ a concentration gradient
o Flows from area of greater conc. To area of lesser conc.
o Active
§ Requires energy (ATP)
§ Proton motive force
§ Transporters allow protons into the cell
· Protons either accept or expel other substances
o Ex. Efflux pumps used in antimicrobial resistance
o Symport – same direction. But doesn’t mean concentration gradient is the same
o Antiport – opposite direction, protons go w/ their concentration gradient
o ATP binding cassette system (ABC transport)
§ Uses binding proteins to scavenge + deliver molecules to transport complex
· Ex. Maltose transport
o Group transport
§ Tricks concentration?
§ Transport mechanism that chemically alters molecule during passage
· Bacterial cell wall
o Made of peptidoglycan
§ Sugar chains wrapped in circles around cell
§ “glycol” = sweet
o Sugar chains linked to each other by short polymers of amino acids
§ Amino acids = peptide
o Rigid structure
o Surrounds cytoplasmic membrane
o Determines shape of bacteria
o Holds cell together
o Prevents cell from bursting
o Unique chemical structure
§ Distinguishes gram + from gram –
· Gram staining
o +.
§ Has a thick cell wall
· 9-amino acid crosslinks in peptidoglycan
§ thin periplasm
o –
§ Thin cell wall
· 4-amino acid crosslinks in peptidoglycan
§ Thick periplasm
· Flagella and pili
o Some bacteria have protein appendages
§ Not essential for life
· aid in survival in certain environments
§ they include
· flagella
· pili
· flagella
o long protein structure
o responsible for motility
§ use propeller-like movements to push bacteria
§ can rotate more than 100,000 revolutions/minute
· 82 mile/hr
o Some important in bacterial pathogenesis
§ H. pylori penetration through mucous coat
o Eukaryote flagella have microtubules, covered by cell membranes
§ The flagellum waves
o In prokaryotes, flagellum spin
§ Flagellin make the flagella
· Motility
o Motile thru sensing chemicals
§ Chemotaxsis
o If chemical compound is nutrient
§ Acts as attractant
o If compound is toxic
§ Acts as a repellent
o Flagella rotation responsible for run and tumble movement of bacteria
o They tumble & run
· types
o Monotrichous- one flagellum
o Polar flagellum- flagellum at end of cell
o Amphitrichous- one flagellum at each end of cell
o Lophotrichous- cluster of flagella at one or both ends
o Peritrichous- spread over entire surface of cell
· Pili
o Considerable shorter and thinner than flagella
o Similar in structure
§ Protein subunits
o Function
§ Attachment
· These pili called fimbre
§ Movement
· Seems accidental, growing pili might move the bacteria, or push off of each other, called twitching
· Conjugation- bacterial sex
o Mechanism of DNA transfer
· Prokaryotic growth
o Environmental factors
§ Temp
§ Oxygen
§ pH
§ water availability
§ and pressure
o temperature
§ growth rate increases w/ temperature
§ protein denature if temperatures are too high
· microbial proteins adapted to temperature range
§ mesophiles – 37 ish degrees C, so the human body
§ psychrophiles like 10-20 degrees C, like the arctic or the fridge/freezer
o water availability
§ all microbes require water for growth
§ high solute concentrations lead to plasmolysis
· increase internal osmolarity
o pumps ions inside
o synthesize osmolysis
§ halophiles- require high concentrations of NaCl
§ 2-4M (10x seawater)
§ Live in salt flats
o Variations in pressure
§ Barophiles
· Adapted to high pressures
o Up to 1,000atm
· Like bottom of ocean
§ Barotolerant organisms
· Grow at high but not very high pressure
§ Barosensitive organisms
· Die at high pressure
o Microbial nutritrion
§ Nutrients
· Must be supplied from environment
· Major elements
o C,H,O,P,K,N,S,Ca,Fe,Mg
o C. hopk’ns café is mighty good
· Trace elements
o Copper, zinc,manganese
§ Carbon source
· Autotrophy
· Heterotrophy
· Ex: chemolithoautotrophs
§ Energy source
· Phototrophy
· Chemotrophy
· Ex: chemoorganoheterotrophs
§ Electron source
· Lithotrophy
· Organotrophy
o Prokaryotic growth
§ Generation or doubling time is the time it takes for the population to double in number
§ Bacterial growth curve, depending on #of cells, you’ll need to apply x degrees to it.
§ Calculating # of bacteria over time
· Nt=N0 x2n
· Measuring bacterial growth: measuring biomass
o Spectrophotometer
§ Measures optical density (turbidity)
§ Cant tell if cells are DOA
§ More cells -> more light scattered -> less light detected
§ transmitted vs absorbed
o viable cell counts
§ counts only cells able to reproduce
· form colonies
o going from a dense concentration to a lesser one each time depending on #of colonies
o controlling microbial growth
§ physical agents
§ chemical agents
§ biological agents
o microbes die at logarithmic rate
o D-value = time to kill 90% of cells
§ 2 D-values = time to kill 99% of cells
§ Example: 20secs Lysol wipe to kill 90% bacteria. We get left w 10 bacteria, now 90% is gone
· Do it again, 2D, 20secs to kill 90%, were left w 1 bacteria, now 99% is gone
· 3 D-value, 20s to kill 90%, left w 0.1 bacteria, now 99.9% is gone
· Etc etc.
· 99.9999% six digits so 60 seconds? For Listerine to kill 1 million bacteria? To calculate, 1,000,000 for 10 seonds, 90% is gone so subtract. 900,000, becomes 100,000. Keep doing it until you get to 1 single digit. Omg it had to be less than one, so 70 seconds
· Food preservation
o Preventing growth + metabolic activities of organisms that cause spoilage and foodborne illness preserves food quality
o Preservation methods include the following physical agents:
§ Canning
§ Pasteurization
§ Cooking
§ Refrigeration
§ Freezing
§ Drying/reducing water availability
o Pasteurization- heating up at a high temp for x amount of time
§ Tuberculosis was the disease that started the process of pasteurization. Because tuberculosis came from cows
§ 63 degrees C for 30mins
§ Flash pasteurization
· Pasteurization treatments do not kill all cells
o Pasteurized food spoils eventually
o Leaves food tasting normal
§ UHT- ultra high temp
· Sterilizes
· Used for creamers
§ Cold temperature-refrigeration
· Slows growth, doesn’t kill all bacteria
· Freezing
o Temperature + pressure
§ Autoclave (high temp and high pressure) = steam cooker
§ Kills all bacteria
§ Kills endospores
· Clostridium botulinum (this toxin is used in botox)
o Botulism
· Bacillus anthracis
o Anthrax
o Other methods
§ Filtration
o Irradiation
§ UV, X-rays, y-rays
· Mutations -> death
· Causes formation of thymine dimers in DNA
o Thymines bind to each other instead of to adenine
· DNA damage can be repaired by several repair mechanisms
· Chemical agents
· Disinfectants- kills pathogens on surfaces
o Destroys eukaryotic cells as well
§ Cannot be used inside patients
§ Ex. Lysol, bleach (chlorine), soap, detergents etc.
· Antiseptics- kills pathogens on your body
o Ex. Mouth wash, Neosporin, peroxide, etc
· Antibiotics
o Must not affect humans
o Selectively kills microbes
§ May not work on all species
o Drug should affect microbial physiology
§ That doesn’t exist, or is greatly modified in humans
· Peptidoglycan
· Differences in ribosome structure
· Biochemical pathway missing in people
o Types of antibiotic compound
§ Bactericidal antibiotics kill the target organism
· many drugs only affect growing cells
o inhibitors of cell wall synthesis
§ only effective if organism is building new cell wall, like penicillin
§ must take antibiotic until all cells leave stationary phase
· bacteriostatic antibiotics
o prevent growth of organism
o cannot kill organism
o immune system removes infection
· biological agents
o probiotics
§ “good” bacteria
· Displace disease organisms from tissues
o Bacteriophages
§ “phages”
§ Viruses that attack bacteria
· Don’t harm eukaryotes