MICRB 265 lec 1
Microbiology studies microscopic life, including:
Acellular: viruses, viroids, prions
Cellular: bacteria, archaea, fungi, algae, protozoa
What is the difference between cellular and acellular life?
Cellular organisms contain the molecular machinery needed to replicate themselves.
Acellular agents must use another cell’s machinery to replicate.
What are viroids, viruses, and prions?
Viroid: Small RNA that replicates; no protein coat
Virus: DNA or RNA inside a protein shell
Prion: Misfolded protein that causes other proteins to misfold
All are infectious but not cells.
What defines a cell? (5)
All cells have:
DNA
Cytoplasm
Ribosomes
A membrane
They also perform metabolism, growth, and evolution.
What is the difference between prokaryotes and eukaryotes?
Prokaryotes: No nucleus, no organelles, dna in nucleoid, ex. bac, archaea
Eukaryotes: nucleus, organelles, dna in nucleus, ex. fungi, protozoa, algae
What shapes do bacteria have? (6)
Cocci (spheres)
Rods
Spirilla
Spirochetes
Filamentous
Stalked / budding
What is the difference between unicellular and multicellular organisms?
Unicellular: one cell does all life functions
Multicellular: cells depend on other cells
What is the RNA World hypothesis?
Life began when self-replicating RNA molecules were enclosed inside liposomes, forming the first primitive cells.
What is LUCA?
The Last Universal Common Ancestor — the first true cell from which all life evolved.
It was bacteria-like.
What is archaeoeukarya?
A common ancestor of archaea and eukaryotes that existed before they split.We know it existed because archaea and eukarya have very similar ribosomes, transcription, and metabolism.
What is endosymbiotic theory? 3 pieces of evidence..
Mitochondria and chloroplasts used to be free-living bacteria that were engulfed and became symbiotic inside eukaryotic cells.
Evidence:
They have their own DNA
They have bacterial-like ribosomes
They replicate independently
What is germ theory?
Many diseases are caused by specific microorganisms, not “bad air.”
Developed by Pasteur.
What did Pasteur disprove?
Spontaneous generation — life does not arise from non-living matter.
This led to pasteurization (heating to kill microbes).
What do Koch’s postulates prove?
They prove that a specific microbe causes a specific disease (causation, not correlation).
Why are Koch’s postulates limited? (4)
They fail when:
Organisms can’t be cultured (viruses)
Asymptomatic carriers exist
Immune status varies
Host specificity exists
How do modified Koch’s postulates work? (3)
They use DNA/RNA sequences to link microbes to disease:
Present when disease is present
Gone when disease resolves
Correlates with severity
What is phylogeny based on?
DNA sequence of 16S ribosomal RNA because it is:
Stable
Inherited
Present in all cells
What is LUCA on a phylogenetic tree?
The root of the tree — the first ancestor from which all lineages diverged.
Which is closer to Staphylococcus aureus: cyanobacteria or E. coli?
Cyanobacteria, because both are Gram-positive-related lineages, not proteobacteria.
What are the main bacterial phyla? and key traits (4x2)
Firmicutes--> Gram +, endospores(clostridia and bacilli)
Actinomycetota--> Gram +, decomposers (streptomycetes and mycobacteria)
Bacteroidetes --> Gram -, gut bacteria, major vertebrate colnizer, modulate host immjune responses (bacteroides)
Prteobacteria--> Gram -, many pathogens, very diverse (escherichia and yersinia)
What limits light microscopy?
Resolution is limited to 0.2 µm by wavelength of light.
Viruses (~0.1 µm) cannot be seen clearly.
Which microscope has the highest resolution?
Electron microscopes
(Shorter wavelength → higher resolution)
What does Gram staining show?
It shows cell wall structure:
Gram+ = thick peptidoglycan → purple
Gram– = thin wall → pink
Why does alcohol remove purple from Gram-negative bacteria?
Their thin cell walls let the crystal-violet-iodine complex leak out, unlike Gram-positive cells.
What does phase-contrast microscopy do?
Allows viewing living cells, showing internal structures and motility.
What does fluorescence microscopy detect?
Molecules that emit light after absorbing energy, such as DNA stained with DAPI.
What makes confocal microscopy special?
A laser focuses on one plane at a time, producing sharper 3D images.
Why must cells be sliced for TEM?
Electrons can’t penetrate thick samples, so thin sections are required to see inside cells.
What theory explains the origin of mitochondria and chloroplasts?
Endosymbiotic theory — mitochondria and chloroplasts were once free-living bacteria that were engulfed and became symbionts.
What are liposomes?
Small spherical lipid vesicles that likely enclosed RNA and were precursors to the first cells.
What is the Golgi apparatus?
An organelle in eukaryotes that modifies, sorts, and packages proteins after translation (not found in prokaryotes).
What is a viroid?
A small circular RNA that is infectious and has no protein coat.
What is a prokaryote?
An organism without a membrane-bound nucleus (Bacteria + Archaea).
What are Archaea?
Single-celled organisms that look like bacteria but are genetically closer to eukaryotes.
What kingdom contains yeasts?
Fungi
What are protozoa?
Free-living or parasitic eukaryotic microbes that feed on organic matter.
What is the shape of Escherichia coli?
Bacillus (rod-shaped)
What is anabolism?
Metabolism that builds large molecules from small ones (e.g., making proteins).
What is a ribosome?
A structure made of protein + rRNA that performs protein synthesis.
What is a coccus?
A spherical bacterial cell shape.
What is horizontal gene transfer (HGT)?
Gene transfer between organisms, not parent → offspring (e.g., plasmids).
What is the nucleoid?
The region of a prokaryotic cell that contains the chromosome.
What is evolution?
Change in heritable traits of populations across generations.
What is a spirochete? What is a stalk?
A spiral-shaped bacterium. A thin cell wall extension that helps bacteria attach to surfaces.
What is a ribozyme?
Catalytic RNA — RNA that acts like an enzyme.
define fungi, algae and protozoa
yeasts and molds and mushrooms, incredibly diverse
photosynthetic aquatic organisms
free living or parasitic eukaryotic microorganisms that feed on organic matter
define archaea and bacteria
similar in size and shape to bacteria, transcription/translation/metabolism more like eukaryotes
the original prokaryote, most have peptidoglycan cell wall and nucleoid
what is the origin of multicellularity
there are examples of unicellularity and multicellularity in both prokaryotic and eukaryotic organisms. some species can even display both uni and multicellularity depending on nutrient availability
what is rna world theory (4)
1. self replicating RNAs+ liposomes= prehistoric cells
2. these develop into DNA containing bacteria
3. Archeoeukarya arise
4. some develop endosymbitic relationship with mitochondria which leads to separation of archaea and eukarya
when disproving spontaneous generation, which liquid putrefies
the flask that is gently tipped so the the microbe laden dust mixes with sterile liquid
what are the steps of the 4 original kochs postulates
1. the suspected pathogen must be present in all cases of the disease and absent from healthy animals
2. the suspected pathogen must be grown in pure culture
3. cells from a pure culture of the suspected pathogen must cause disease in a healthy animal
4. the suspected pathogen must be reisolated and shown to be the same as the original
rank the taxonomic classifications
Life, domain, kingdom, phylum, class, order, family, genus, species
who wrote the first book dedicated to microscope observation (mold structures)? who discovered bacteria?
robert hooke
antoni van leeuwenhoek
define resolution. what affects it?
ability to distinguish adjacent objects and distinct and separate
affected by the wavelength light. longer wavelength= lower resolution.
define contrast
ability to see an object as distinct from its surroundings
what 4 types of microscopy is light microscopes used for
brightfield
phase contrast
differential interference contrast
fluorescence/ confocal
electron microscopes are used for
scanning electron microscopy and transmission electron microscopy
descrive bright field microscopy
1. can be hard to see
2. move in and out of fields of view with fluid dynamics
3. not a lot of useful information
how to do a gram stain (4)
1. heat fixed smear is penetrated with crystal violet for 1 min (turn cells purple)
2. iodine added for 1 min, causes crystal formation (still purple)~ need to be added for good amount of time
3. decolorize with alc for 20 sec, crystal solubilize and move out but in gram + thick peptidoglycan keeps them in. so only purple washes away in gram -. if left for too long, crystals can dissolve too much and get out of either.
4. safranin used as counterstain to dye everything, gram + still stays purple but gram - is now pink. if you leave for too long gram + can look pink
bright field vs phase contrast
bright field: sample contrast comes from attenuation of light in the sample
phase contrast: sample contrast comes from the interference of different path lengths of light through the sample
flueorescence microscopy
certain molecules can absorb photons, this causes electrons to excite and move into higher energy state, when the electrons relax they emit a photon. this photon is called fluorescence
electron microscopy (3)
uses electrons instead of light
shorter wavelength than photons
larger than photons
in electron micrscopy shorter wavelength than photons and larger than photons means.
shorter wavelength than photons --> higher resolution ( 1000x greater than light microscopes)
larger than photons--> cannot penetrate "larger" objects. proteins = good, cells= bad. to be able to see inside the cell you have to cut the cell.
the difference between cellular and acellular organisms is that?
cellular organisms contain the molcule machinery necessary for their own replication
endosymbitotic theory best explains
the difference between archaea and eukarya
timeline these events : spontaneous generation disproved, bacteria observeed, koch postulates, growth of bacteria on media, mold observed, germ theory of disease, community sampling of rRNA, kochs modified postulated, first genome sequenced
germ theory of disease
mold observed
bacteria observed
spontaneous generation disproved
growth of bac on solid media
koch postulates
community sampling of rRNA
first genome seq
kochs modified postulates
collectively what kochs postulates do
they demonstrated that particular microorganisms cause particular diseases
what is one issue with the original kochs postulates
organisms like pseudomonas aeruginosa only cause disease in immunocompromised patients
which microscope has the highest resolution for light microscopes? what about in general?
fluorescent confocal microscope
scanning electron microscope.