Lecture 1 - intro to microbes Flashcards
What constitutes a microorganism/microbe?
- A “living” organism that requires microscopy to be observed and studied.
- Broad term that encompasses many different organisms that are not necessarily phylogenetically related.
- High genetic diversity (ex: diverse metabolic pathways)
Example of microbial influence on human activity and history.
Pandemics, fermentation of food and beverages to prevent spoilage
Example of microbial influence on geology/ecology
Geochemical cycling impact agriculture (ex: nitrogen fixation)
Industrial and technological applications of microbio
Taking advantage of microbes that can survive in extreme conditions. Ex: taking enzymes from them to use for our own purposes like improving dish detergent efficiency
Typical size of microbes
Ranges from 0.2 micrometers (one thousand times smaller than our natural visual limit) to a few millimeters (viruses are usually 10-fold smaller)
What types of organisms are considered microbes?
Eubacteria, archaebacteria, fungi, protozoa, and viruses
Eubacteria vs archaebacteria
Eubacteria = true bacteria
Archaebacteria have similar features, but are not related to eubacteria
Are bacteria unicellular?
Yes, but they can exist in multicellular groups. In these groups, some bacteria will differentiate to serve different purposes (ex: cyanobacteria have some cells that generate carbohydrates and some that fix nitrogen)
Who is Robert Hooke?
Developed the first compound microscope. Low magnification, could see bread mold, human cells, and plant cells. Coined the term “cell”
Who is considered the first microbiologist?
Antonie van Leeuwenhoek. Cloth draper who build high power microscopes to see thread count of fabrics. First person to observe and describe single celled microorganisms.
Why is fermentation of food and beverages advantageous?
Helps make cleaner and more renewable waste. Prevents more dangerous microbes from growing because harmless microbes consume the material first. Helps food/beverage last longer.
What are lithotropic bacteria?
“Rock eating” bacteria that promote decay/solubilization of minerals. Facilitate bioleaching of metals from mineral ores (ex: harvesting Cu from circuit boards).
Why are bacteria important for geochemical cycles?
Bacteria are able to naturally fix nitrogen which is important for crop growth. Humans can only fix nitrogen by using energy-intensive industrial methods.
Who is Sergei Winogradsky?
- discovered lithotrophs
- developed enrichment cultures
- built the Winogradsky golumn which models wetland bacterial ecoystem
Why do lithotrophs “eat rock?”
Use inorganic materials instead of organic as the source of e- for metabolism.
How does the Winogradsky column work?
Bacteria self-segregate into niches depending on their metabolic pathways. Each niche supplies material for others.
How do bacteria generally arrange themselves in a Winogradsky column?
Aerobic on top and more anaerobic as you move down
What is bubonic plague caused by?
Yersinia pestis: bacteria that infects the foregut of the tropical rat flea
How does yersinia pestis get to humans?
- Infects foregut of tropical rat flea
- Causes flea to bite and regurgitate the bacteria
- Bacteria transferred to new host (rat)
- Rats infect humans
What were the effects of the bubonic plague on human demographics and cultural practices?
Demographic: 25-33% of the European population died
Cultural: Mass death caused society to question the need for moral behavior –> societal collapse
How did someone die of bubonic plague in modern times?
Infection via lab exposure to vaccine strain. Vaccine strain was attenuated by blocking the bacteria’s ability to produce iron. Person infected had an excess of iron in the bloodstream which allowed the bacteria to thrive
Hemochromatosis
Condition in which an individual’s organs accumulate an excess of iron
What causes tuberculosis?
Mycobacterium tuberculosis (bacteria)
Why is tuberculosis still a big problem?
Highly resistant to treatment, long treatment time, leads to chronic respiratory infection
How can tuberculosis be diagnosed?
Chest x-ray: mass found at the top of the lungs
Sputnum culture from a burst granuloma
Granuloma
A “walled off” area in the body that prevents bacteria from migrating within the body. If ruptured, it can lead to coughing and sputnum production
20th century microbial diseases
Influenza pandemic (1918) and AIDS caused by HIV
HIV + TB infection…
leads to longstanding respiratory infection. HIV makes someone more susceptible to TB
Immunization
Stimulation of a protective immune response by deliberate inoculation with an attenuated or inactive pathogen.
Who is Lady Mary Montagu?
Introduced the practice of smallpox inoculation to Europe in 1717.
Who is Edward Jenner?
Deliberately infected patient with matter from cowpox lesions to inoculate against smallpox.
Origin of the term “vaccination”
Cowpox is caused by the vaccinia virus (Latin vacca for “cow”)
Who is Ignaz Semmelweis?
Doctor who suggested that docs was their hands with calcium hypochlorite (antiseptic agent) to prevent puerperal (childbed) fever
What is an antiseptic?
Chemical agent that kills microbes; not specific to any specific to any type of microbe
What caused an increase in puerperal fever in the Wien maternity clinic?
Med students began taking pathological anatomy and dissecting cadavers without washing their hands afterwards. Onset of chlorine handwashing in 1849 caused a sharp decrease in fever.
Who is Joseph Lister?
Developed carbolic acid to treat wounds and clear surgical equipment. First person to introduce the idea of aseptic surgery
Why did Joseph Lister use carbolic acid?
Used in Scotland to decrease the smell of human feces.
When was aseptic surgery developed?
20th century. Made surgical environments completely microbe-free.
Who is Robert Koch?
- Founder of the scientific method of microbiology.
- Developed pure culture techniques
- 4 postulates to determine if a microbe causes a disease
What do Koch’s postulates do/require?
Criteria for establishing a causative link between an infectious agent and a disease. Requires a pure population of a microbe. Some of the postulate are not completely accurate.
What are the 4 Koch postulates?
- Microbe is always present in diseased host and absent in healthy individuals (not always true because some things are present in healthy individuals too)
- Microbe must be isolated from diseased individuals.
- Healthy individual becomes ill when pure microbe is introduced to host.
- Same microbe is re-isolated from inoculated individual.
What illnesses did Koch’s postulates help to identify?
Anthrax, vibriocholera, mycobacterium tuberculosis
Who is Barry Marshall?
Proved that H. pylori caused peptic ulcers by self-inoculating himself with H. pylori
Who discovered penicillin
Alexander Fleming
Who first purified penicillin?
Howard Florey and Ernst Chain; became the first commercial antibiotic to save human lives
Does penicillin treat gonorrhea?
It used to, but gonorrhea is now resistant to penicillin
Why are multidrug resistant bacteria often isolated from rivers?
Excreted antibiotics move into our water systems and into natural rivers
What is streptomycin?
Antibiotic made by the streptomyces bacteria that targets bacterial ribosomes
What is carbapenem-resistant enterobacteriaceae (CRE)?
Antibiotic resistant gut bacteria found on contaminated endoscopy equipment. Breaks down carbapenem
Spontaneous generation
Theory that living creatures arise from non-living matter. Prevalent view from ancient times and codified by Aristotle.
Example of “spontaneous generation”
It appeared that maggots came from rotting meat.
Equivocal generation
Theory that some organisms arise from dissimilar creatures.
Example of “equivocal generation”
Many arthropods have different developmental stage of life (ex: butterfly arising from cocoon)
Who is considered the father of western science?
Aristotle
Why did Jan Baptist van Helmont’s recipe for a mouse make sense?
1 piece of soiled cloth + 1/2 peck of wheat –> incubate for 21 days.
Gestational period of a mouse is about 20 days, mouse makes a nest in the soiled cloth and wheat
Evidence against spontaneous generation
Fancesco Redi (1660): Maggots in decaying meat were offspring of flies
Lazzaro Spallanzani (1760): Sealed flask of meat broth sterilized by boiling failed to grow microbes
Who discovered the microbial basis of fermentation?
Louis Pasteur
How did Louis Pasteur disprove spontaneous generation?
Swan neck flasks held growth medium and allowed access to air, but didn’t allow microbes in due to the shape of the neck. No microbial growth even with air
Who defined microbes as a separate category from plants or animals?
Erst Haeckel
- said that there was a third kingdom of life called Protista
What does the term Monera describe?
A phylum within kingdom Protista that contains bacteria
Who divided Protists into eukaryotic and prokaryotic?
Herbert Copeland
- eukaryotic: protozoa and algae, larger and more complex
- prokaryotic: bacteria
Who phylogenetically separated out fungi from plants?
Robert Whittaker
Who proposed the endosymbiotic theory surrounding mitochondria and chloroplasts?
Lynn Margulis
Mitochondria and chloroplasts evolved through endosymbiosis between prokaryotic cells and proto-eukaryotes
Why was endosymbiosis of prokaryotes and proto-eukaryotes beneficial?
- Prokaryotes received protection and metabolic products from prokaryotes.
- Proto-eukaryotes received high energy production capability. Some prokaryotes produced amino acids for proto-eukaryotes.
Why are mitochondria necessary for growth of large organisms?
Mitochondria are much more efficient in production of ATP due to oxidative phosphorylation
When did oxidative phosphorylation arise?
When O2 became prevalent in the air due to output from cyanobacteria
What were the two prokaryotes that are involved in the endosymbiotic theory
- Cyanobacteria (phototrophic) –> chloroplast
- Proteobacteria (respiring) –> mitochondria
Who identified archaebacteria?
Carl R. Woese studied 16S rRNA sequences of unusual prokaryotes found in hot springs. Study revealed that they were a distinct form of life
Why is 16S rRNA used for phylogenetic study?
16S rRNA is essential for the function of ribosomes and therefore for life. Evolves very slowly, so it can be used to find phylogenetic differences in bacteria.
Bacteria vs archaea cell wall
Bacteria uses peptidoglycan for their cell walls. Archaea don’t
How was molecular genetics (DNA manipulation) developed?
Discoveries in bacteria and bacteriophages pushed the discipline forward.
What allowed for recombinant DNA and molecular cloning to arise?
Restriction endonucleases and plasmids. Beginning of gene cloning.
Example of how recombinant DNA and molecular cloning are used
Manufacturing human proteins (ex: insulin or erythropoietin) in bacteria
How was the CRISPR-Cas system discovered?
Yogurt manufacturers realized that bacteria would sometimes get infected –> loss of yogurt. Looked for a way to protect bacteria against viral infection
What is CRISPR-Cas used for?
Site-specific genome editing in eukaryotic cells.
Example of CRISPR-Cas use
Curing cystic fibrosis in cells with specific insertion of a phenylalanine codon
How big are eubacteria and archaea?
0.4-10 micrometers. Most bacteria are between 1 and 3 micrometers.
How big are protozoa, algae, and fungi?
10 to 100 micrometers
Optical resolution
Distance at which two objects can be distinguished
What does optical resolution depend on?
- Adequate contrast between object and surroundings (most things that we need to observe are naturally colorless)
- Wavelength of the illuminating EM radiation (Abbe diffraction limit)
- Density of light detectors/photoreceptors
Detection
Ability to discern the presence of an object.
What is the resolution of the human retina?
Approximately 150 micrometers (0.15 mm)
What is the resolution of the retina limited by? Give an example.
Limited by quality and density of photoreceptors. Ex: bacteria that are a few microns apart will reflect photons so close together that they are perceived by the same photoreceptor. This means that we are unable to specifically detect each individual bacteria.
What is magnification?
Increases the apparent size of an object; can sometimes reveal more detailed info about the object.
Which forms of EM are most harmful to us and why?
UV, X-rays, and gamma rays are high enough energy to ionize water –> DNA damage
Why have we evolved to see the light spectrum that we do?
The visible spectrum is the part of the EM spectrum most emitted by our sun
Why is ozone important?
Absorbs light in the UV spectrum; greatly reduces the amount of UV light that reaches us
Which photopigments absorb light in areas where chlorophyll can’t?
Phycoerythrin and phycocyanin absorb light in the 500-600 nm range
Absorption
Photon energy is acquired/preserved by the absorbing object
Four types of light interaction with objects
Absorption, reflection, refraction, scattering
Reflection
Photon “bounces” off the surface of an object
Refraction
Speed of the photon is altered by a substance is passes through, thereby changing its direction
Scattering
Can occur when the wave front interacts with objects similar in size to the wavelength of the incident light
How is scattering used in microbial analysis?
Used to estimate the density of cells in a solution.
What wavelength of light is used to estimate bacterial concentration in a solution?
600 nm, approximately half the length of a standard bacteria
Which direction does light bend when it enters something with a higher/lower refractive index?
Higher: towards the normal
Lower: away from the normal
What are the refractive index value for air, water, and glass?
1, 1.3, 1.5
Why does refraction lead to magnification?
Light can be refracted in such a manner that our eye perceives the path of light to originate from a closer source
What is a compound microscope?
System of multiple lenses designed to correct or compensate for aberrations. Composed of an ocular lens (closer to the eye) and an objective lens (closer to the sample, designed to be parfocal).
What does parfocal mean?
A sample will remain in focus when moving between different objective lenses.
Total magnification of a compound microscope =
magnification of the ocular lens multiplied by the magnification of the objective lens
How are lens curvature and magnification related?
Greater curvature = greater magnification
Bright-field microscopy
Generates a dark image of an object over a light background
What is resolution of bright-field microscopy increased by?
- Higher numerical aperture
- Shorter wavelength light
- Higher contrast
How to increase numerical aperture
NA = n sin theta
- increase theta with wider lens closer to specimen
- increase refractive index (n) with immersion oil
How does immersion oil help increase resolution?
Immersion oil has a higher refraction index than air (usually similar to glass). When it fills the gap between the glass holding the specimen and the objective lens, there is greater refraction of light –> more light can be perceived by the objective lens
How does wavelength of light limit resolution?
Shorter wavelength = higher resolution. Wave fronts that are focused by a lens can interfere with one another –> rings of constructive/destructive interference of wave fronts appear around an obect
Abbe diffraction limit
Limit to how close two objects can be to each other and still be resolved. Different for every microscope.
What is the minimum distance that the human eye can resolve two separate objects?
250 nm
What is an airy disk?
Rings of constructive/destructive interference surrounding an image seen in a microscope
Cell fixation (microscopy)
Cells made to adhere to a slide in a fixed position
Cell staining (microscopy)
Cells imbued with pigment.
What do simple stains bind to? (microscopy)
Bind to cells, but not the surrounding media.
How do stains work?
Most stains have conjugated pi orbital systems, aromatic rings, and are cationic. This causes electrons to be held further from their nuclei –> easier for them to absorb energy.
What were John Tyndall’s experiments based on?
Used a box (Tynadall box) to establish link between dust in the air and growth of microbes. Box lined with glycerin to trap dust. Also used glass tubes to compare mountain vs cabin air.
Outcomes of John Tyndall’s box experiments?
- dust + growth media = growth
- no dust + media = no growth
- dust + media + heat = no growth
- dust from hay + media + heat = growth; microbes in hay dust must be heat resistant
Outcomes of John Tyndall’s glass tube experiments? (mountain vs cabin)
Fresh mountain air was more sterile; no growth when media exposed to it. Air from cabin + media –> growth. Air purity contributes to contamination
What is Tyndallization/fractional sterilization?
Serial heating of a sample for short periods of time to gradually kill off heat resistant microbes (doesn’t work for all microbes)
How does Tyndallization work?
First heating kills vegetative bacteria. Second heating kills remaining spores before germination
